nemat462abs 130..260 - M.MOAM.INFO (2025)

Journal of Nematology 46(2):130–260. 2014. Ó The Society of Nematologists 2014.

ABSTRACTS

ROOT KNOT NEMATODES: NEW INSIGHTS INTO PARASITISM SUCCESS. Abad, P. Institut Sophia Agrobiotech, UMR, INRA - CNRS - UNS, 400 route des Chappes, F-06903, Sophia-Antipolis, France. The root-knot nematode Meloidogyne incognita is a widespread and polyphagous obligate asexual endoparasite of plants that causes serious and growing problems to agriculture. This lifestyle implies dramatic changes of plant cells into specialized feeding sites, which are induced by secreted proteins by the nematode, so-called parasitism effectors. An integrated approach of molecular techniques has been used to functionally characterize nematode parasitism proteins. The complete genome sequence of M. incognita revealed that the assembled sequence consists of homologous but divergent segment pairs that might represent former alleles in this species. Based on comparative genomics, we identified in root-knot nematode genomes a set of genes preserved during the evolution of plant-parasitic nematodes, only shared with organisms having a plantparasitic lifestyle. These genes represent new targets to develop new methods to control plant-parasitic nematodes but harmless for the environment and consumer health. Another interesting feature of the M. incognita species is the paradox between its apomictic mode of reproduction and its potential host range encompassing more than 3,000 plant species. The adaptation of M. incognita to its environment (e.g., reproduction on resistant hosts) raises questions about genome plasticity leading to genetic variation and adaptive evolution. We reasoned that epigenetic mechanisms might in part be responsible for the generation of phenotypic variants that provide material for rapid adaptation. Thus, Meloidogyne spp. constitutes a unique model system to study the links between variation in genome structure, mode of reproduction, and adaptation to environment and hosts, in relation with parasitic success. PLANT-PARASITIC NEMATODE THREATS TO GLOBAL FOOD SECURITY. Abd-Elgawad, M.M.M. Phytopathology Department, National Research Center, EI-Tahrir St., Dokki 12622, Giza, Egypt. Plant-parasitic nematodes constitute a major constraint to agriculture. Estimates of their crop-loss are important for establishing research, extension, and budget priority. Regulatory policy action, pesticide impact assessment, resource allocation, and program prioritisation are usually contingent upon crop loss data. Recent questionnaire results of this study for world-wide estimates of crop losses due to plant-parasitic nematodes are presented herein. For the 20 life-sustaining crops, an average worldwide crop loss due to nematodes of 12.6% is estimated which equalled $215.77 billion annual yield loss. The other crops that represent a miscellaneous group important for food or export value have an average annual yield loss of 14.45% which equalled $142.47 billion. Losses for the total 40 crops average 13.5%. So, worldwide crop losses due to nematodes on 37 crops are estimated at $358.24 billion annually based on the 2010-2013 production figures and prices. These figures are staggering, and the real figure, when all crops throughout the world are considered, probably exceeds such estimations. Our current estimates far exceed previous ones probably because of challenging issues such as reduced number of effective nematicides available and limitation in their use due to environmental issues, increased adoption of intensive agriculture, climate change, occurrence of resistance-breaking pathotypes on economically important crops, and potential introduction of quarantine-nematodes. Basic and applied nematological research should be more oriented to provide better management of plant-parasitic nematodes in an economically and environmentally beneficial manner. NEMATOLOGY EDUCATION AND TRAINING IN EGYPT AND NEIGHBOURING COUNTRIES. Abd-Elgawad, M.M.M. Phytopathology Department, National Research Center, EI-Tahrir St., Dokki 12622, Giza, Egypt. During the second half of the twentieth century, pioneer nematologists from Egypt and neighbouring countries, taught nematology at universities in the Middle East for under and post-graduate students and few of them worked in the agricultural service institutions and governmental bodies for research, training and extension. In the last two decades, most Middle Eastern nematologists were taught by those pioneers or their students and graduated from local universities due to the high cost of education abroad. General Nematology courses are taught for undergraduates, but more specialized courses like Nematode Morphology, Nematode Physiology, Nematode Taxonomy, Nematode Management, Nematode Ecology, and Insect Nematology are taught for post-graduate students. In Egypt, infrastructure and training programs of nematology are based on three sectors namely: Egyptian universities (especially faculties of agriculture), the Agricultural Research Centre of the Ministry of Agriculture and Land Reclamation, and some institutes of the Ministry of Scientific Research. Similar governmental bodies are found in neighbour countries for nematode teaching, research and/or training programs. Currently, the numbers of Middle Eastern nematologists are almost enough to enhance the level of nematode education and training in their countries, but several drawbacks hinder the advance of applied nematology. These include lack of coordination and cooperation between specialists, insufficient awareness of nematode diseases and control especially by small scale farmers, limited financial resources, shortage in scientific instruments and equipment, and low activity of related agricultural 130

Abstracts 131 extension. The allocation of higher funding rates are proposed and assigned for specific nematological activities/sections to solve such problems. MICOFLORA ON ROOT-KNOT NEMATODES IN SOIL WITH DIFFERENT LEVELS OF SUPPRESSIVENESS. Adam1, M., A. Westphal2, J. Hallmann1 and H. Heuer1. 1Institute for Epidemiology and Pathogen Diagnostics , Julius Ku¨hn-Institut, Messeweg 11/12, 38104 Braunschweig, Germany; 2Institute for Plant Protection in Field Crops and Grassland, Julius Ku¨hn-Institut, Messeweg 11/12, 38104 Braunschweig, Germany. Infective stages of plant-parasitic nematodes dwell through the soil matrix before entering host plant tissues. Attachment of specific nematode antagonists on the nematode during this process has been documented. There is a lack of information of attachment of microbes, which are not known as parasites of nematodes. In a greenhouse experiment of three soils with Meloidogyne hapla, numbers of galls, eggmasses and eggs were lower in non-treated portions than the heated equivalents. The hypothesis of a biotic cause of this suppression was supported by the differences of the microbial communities and concomitant differences in level of suppressiveness among the soils. In baiting studies with second-stage juveniles (J2) of M. halpa, differences in PCR-DGGE of the fungal ITS or bacterial 16S rRNA genes were detected in DNA isolated from soilfree J2 after incubation in the soils. Especially in fungal DGGE fingerprints, many fungal species were detected on the J2 that were below detection limit in the surrounding soil. On J2 from all three soils, species related to the genera Davidiella and Rhizophydium were detected. J2 baited in the most suppressive soil were specifically associated with the genera Eurotium, Ganoderma, and Cylindrocarpon. Among the 20 operational taxonomic units of bacteria specific for J2 in suppressive soil six were closely related to infectious species like Shigella sp.; most frequent were Malikia spinosa and Rothia amarae. In conclusion, a diverse microflora adhered to M. hapla J2 that may interfere with female fecundity of the nematode. Exploiting functions of these microbes may be possible. COMPARISON OF LIFE HISTORY TRAITS OF THE ENTOMOPATHOGENIC NEMATODES STEINERNEMA FELTIAE AND STEINERNEMA RIOBRAVE. Addis1, 3, T., A. Teshome2, O. Strauch3 and R.-U Ehlers3. 1Faculty of Agricultural and Nutritional Sciences, Christian-Albrechts-University, Hermann-Rodewald-Str.4, 24118 Kiel, Germany; 2 Department of Biology, Ghent University, K.L. Ledeganckstraat 35, B-9000 Ghent, Belgium; 3e;nema, GmbH, Klausdorfer Str. 28-36, D-24223 Schwentinental, Germany. Life history traits (LHT) of entomopathogenic nematodes Steinernema feltiae and Steinernema riobrave were assessed at 258C using a hanging drop technique. The LHTs were studied using 5x, 10x and 20x 109 cells ml-1 of Xenorhabdus bovienii and Xenorhabdus cabanillasii for S. feltiae and S. riobrave, respectively, in semi-fluid nematode growth gelrite. The results indicated that increased food density had a significant positive influence on offspring production on both, S. feltiae and S. riobrave. Highest offspring production was recorded at bacterial food densities of 20 x109 cells ml-1 with 813/female for S. feltiae and 1,913/female for S. riobrave. Significant positive correlations of R = 0.90 and 0.76 were found between body length and offspring production for S. feltiae and S. riobrave, respectively. The lowest intrinsic rate of natural increase (rm) (1.1 days) was recorded for S. feltiae and the highest (1.4) for S. riobrave. A population doubling time (PDT) =0.6 days was recorded for S. feltiae and 0.5 days for S. riobrave. The life span of female nematodes, which ended the day following initiation of endotokia matricida, was not significantly different among the different bacterial food densities in both species. Significant differences in offspring production and population growth rate were assessed when comparing S. feltiae with S. riobrave. This result will be used to further investigate the optimal bacterial food density for mass production in bioreactors for maximum dauer juvenile recovery, synchronised population development and final yields of S. feltiae and riobrave. IMPROVING PRODUCTIVITY OF CITRULLUS LANATUS SUBSP. MUCOSOSPERMUS (EGUSI MELON) THROUGH IDENTIFICATION AND USE OF LOCAL NEMATODE RESISTANT VARIETIES. Affokpon1, 2, A., H.K. Baimey3, E.G. Achigan-Dako2, C. Tossou1, J.N. Lokossou1 and A.H. Bokonon-Ganta2. 1Institut National des Recherches Agricoles du Be´nin, Be´nin; 2Universite´ d’Abomey-Calavi, Be´nin; 3Universite´ de Parakou, Be´nin. Citrullus lanatus subsp. mucosospermus F. (egusi melon, Cucurbitaceae) is a traditional fruit vegetable with important nutritional, economic and socio-cultural values in West Africa. However, root-knot nematodes (Meloidogyne spp.) are a major production constraint, and a reason for drastic reduction of the egusi melon cultivated areas. To develop strategies for sustainable root-knot nematode management in egusi melon production systems, eight egusi landraces from Benin were assessed for resistance or tolerance to root-knot nematodes in naturally infested fields. Results indicated that responses to root-knot nematode attacks differ significantly among egusi landraces. The egusi landrace BEN 1-3-7 showed the greatest potential in nematode suppression with a final nematode population density of 236 nematodes per 25 g roots. In contrast, the landrace BEN 1-5-13 had the highest final nematode population density (13929 nematodes / 25 g root), with the highest average hulled grain yield (528 kg/ha), suggesting a good tolerance of this landrace to root-knot nematodes. Root galling varied significantly between egusi landraces, with an average gall index of 3.25 over 10. Correlation analysis indicated, in some cases, a negative correlation between galling index and seed weight, and between nematode densities and seed weight.

132 Journal of Nematology, Volume 46, No. 2, June 2014 The results provide evidence of genetic variability among West African landraces of egusi melon for reaction to root-knot nematodes. POTENTIAL OF MALDI-TOF MASS SPECTROMETRY AS A DETECTION TOOL TO IDENTIFY PLANT-PARASITIC NEMATODES. Ahmad1, F., O.O. Babalola1 and Hui-Fen Wu2,3,4. 1Department of Biological Sciences, Faculty of Agriculture, Science and Technology, North-West University, Mafikeng Campus, Private Bag X2046, Mmabatho 2735, South Africa; 2Department of Chemistry, National Sun Yat-Sen University, Kaohsiung 80424, Taiwan; 3Center for Nanoscience and Nanotechnology, National Sun Yat-Sen University, Kaohsiung 80424, Taiwan; 4Doctoral Degree Program in Marine Biotechnology, National Sun Yat-Sen University, Kaohsiung, 80424, Taiwan. Plant diseases caused by plant-parasitic nematodes substantially reduce economic crop production every year, resulting in massive economic losses throughout the world. Accurate identification of plant-parasitic nematodes is essential to plant pathogen diagnostics and, thus, plant disease management. The root-knot nematode, Meloidogyne incognita (Kofoid and White) Chitwood, is a sedentary endoparasite that retards growth and development of plants by attacking the root system, causing galling, stunting, and other adverse effects. We demonstrated the feasibility for the use of MALDI-TOF MS for rapid, direct and sensitive detection of adult root-knot nematode and their second stage juveniles (J2). We have proposed simple pretreatment protocols and have identified M. incognita with help of generated mass spectra of targeted samples. We also reported the differentiation between the adult and juvenile stages of the nematode based on mass spectrometric profiling, with the adult and juvenile stages exhibiting different marker peaks. However, common peaks to both stages were also found. The ability to record biomarker ions, in a broad m/z range, which are unique to and representative of individual nematode species, could form the basis of identification of nematodes by MALDI-TOF MS. However, more work needs to be done on other genera and species to know if identification between species of nematodes is possible. NEMATICIDAL ACTIVITY OF POKEWEED EXTRACTS. Ahmed1, S., P. Gerard2 and P. Agudelo1. 1School of Agricultural, Forest, and Environmental Sciences, Clemson University. Clemson, South Carolina 29634, USA; 2Department of Mathematics, Clemson University. Clemson, South Carolina 29634, USA. The nematicidal activity of pokeweed (Phytolacca americana) extracts on five plant-parasitic nematodes species was investigated. Extracts of leaves, stem, and roots were evaluated in vitro for their toxicity to Ditylenchus dipsaci, Pratylenchus penetrans, Apehlenchoides fragariae, Rotylenchulus reniformis, and Meloidogyne incognita. Extracts from the different plant parts obtained using different solvents (water, ethanol, methanol, and acetone) were also compared. The calculated LD50 for the extracts was used to determine the range of concentrations to be used in subsequent greenhouse studies. We report the results of greenhouse studies with root-knot and reniform nematode on soybean plants. Seedlings of soybean cv. Hutcheson were inoculated with 3,000 reniform nematodes or 2,000 root-knot nematode juveniles, one week after planting. One week after inoculation, selected extracts of leaves, stems or roots were added at a dose of 300 mg per plant, with the nematicide oxamyl (24% Vydate, 0.6 ml per plant) as positive control and untreated plants as negative controls. Each experiment was repeated twice and 20 replicates per treatment were included. Six weeks after inoculation, galls and egg masses per plant were assessed for the root-knot nematode experiments and reproduction factor for the reniform nematode experiments. Water extracts from pokeweed stems and leaves showed significant reduction of nematode reproduction and provided the same control as Oxamyl. These results suggest that pokeweed extracts are potential useful nematicides for management of root-knot and reniform nematodes. HOST STATUS OF THREE LOLIUM GRASS CULTIVARS TO SOUTH AFRICAN MELOIDOGYNE SPEICES. Ahuja1, P., H. Fourie1 and A.P. Nyczepir2. 1Unit of Environmental Sciences and Management, North-West University, Potchefstroom-2520, South Africa; 2USDA-ARS, SE Fruit & Tree Nut Research Laboratory, USA. Lolium grass is used extensively as a cover crop and for forage production in South Africa. The host suitability of two popular South African Lolium cultivars (Agrihilton & Raaigrass) and one (Jesup Max-Q) from the USA were evaluated for their host status to Meloidogyne incognita and Meloidogyne javanica under greenhouse conditions. Lolium and tomato seedlings (cv. Rodade) were inoculated with ± 2000 second-stage juveniles of the two Meloidogyne spp. The parameters recorded 56 days after nematode inoculation were i) number of egg masses and ii) number of eggs and second-stage juveniles/root system. Substantial, but insignificant variation existed among the three Lolium cultivars with regard to the parameters recorded. For M. incognita all three Lolium cultivars had Rf values > 1, indicating susceptibility to these nematodes. Conversely, Rf values < 1 were recorded for the three Lolium cultivars when evaluated against M. javanica, indicating resistance to the latter species. None of the evaluated cultivars of Lolium, however, were immune to either M. incognita or M. javanica. The susceptible tomato cultivar Rodade, had significantly higher numbers of egg masses and eggs/ root system compared to those recorded for the Lolium cultivars. Inclusion of these three Lolium cultivars in integrated nematode management systems where M. javanica causes problems can contribute towards managing this nematode pest effectively. However, in soils where M. incognita prevails none of these three Lolium cultivars is recommended to be cultivated.

Abstracts 133 POTENTIAL OF VETIVER (CHRYSOPOGON ZIZANIOIDES) GRASS ROOT EXUDATES AND EXTRACTS AS A TOOL TO MANAGE MELOIDOGYNE. Ahuja, P., M.S.A. Pretorius and H. Fourie. Unit for Environmental Sciences and Management, North-West University, Private Bag X6001, Potchefstroom, 2520. In 2013 Meloidogyne has been listed as the number one plant-parasitic nematode genus that damage agri- and horticultural crops worldwide. The withdrawal of synthetic nematicides from world markets necessitated studies aimed at evaluating various plant-derived products with nematicidal/nematostatic characteristics. Various concentrations of crude Vetiver root exudates and extracts, respectively, were evaluated for their effects on the motility of Meloidogyne javanica J2 as well as the reproduction of this pest in laboratory and greenhouse experiments. The 50, 75 and 100% root exudate concentrations reduced J2 motility significantly (p#0.05) from 48 h after the onset of the trial compared to the tap water control. For root extracts, from 53 to 94% of the J2 were non-motile for the 25, 50, 75 and 100% product concentrations, respectively, compared to that of 35% for the tap-water control at termination of the trial (94h sampling interval). In vivo reproduction data showed no significant differences for the number of egg masses as well as eggs and second-stage juveniles (J2)/root system 56 days after J2 that were exposed to the Vetiver root exudates were inoculated on roots of tomato seedlings. Although Vetiver root exudates inhibited the motility of J2, they were able to recover when removed from it and thus reproduced in tomato roots. A similar assay is currently in process for the root extract concentrations. Follow-up studies will focus on identification and refining of the chemicals that adversely affected J2 motility. Field testing of such products are also envisioned since this may render an alternative and environmentally safe product for future use to minimise damage inflicted by Meloidogyne in crops. INTRASPECIFIC VARIABILITY AND GENETIC STRUCTURE IN FRENCH GLOBODERA TABACUM POPULATIONS. Alenda, C., J. Montarry and E. Grenier. INRA, UMR IGEPP, Domaine de la Motte au Vicomte, BP35327, 35653 Le Rheu, France. The dispersal abilities and the population genetic structure of nematodes living in soils are poorly known. In the present study, we have pursued these issues in the tobacco cyst nematode, Globodera tabacum, which is responsible for significant yield reductions in the USA, France, Spain and Italy. In this study, ten microsatellites markers were used to analyse the patterns of its genetic structure in France and to compare the genetic diversity observed to some European and American populations. Large heterozygote deficiencies were observed at most loci. The limited active dispersal ability of larvae in soil, which favours mating between (half) siblings or a genetic structure at a sub-population scale, could be responsible for this pattern. Bayesian genetic assignments revealed two distinct genetic groups that matched with the geographic repartition of two agricultural cooperative societies. Very high FST values were observed between the overall pairs of French populations and analysis of partitioning of molecular variability showed that an important part of the genetic variability was observed among agricultural cooperative societies (34%) and among populations (39%), which support limited gene flows among the TCN populations. PLANT PARASITIC NEMATODES COMMUNITIES ASSOCIATED WITH OLIVE TREES IN MOROCCO. Ali1, N., E Chapuis1, J. Tavoillot1, M. Aı¨t Hamza2, A. El Mousadik2, A. El Oulkadi3, G. Besnard4, A. El Bakkali5, A. Moukhli3, B. Khadari6, C. El Modafar7, M. Ater8 and T. Mateille1. 1IRD, UMR CBGP, Montferrier sur Lez, France; 2UIZ-FST, Agadir, Morocco; 3INRA, Marrakech, Morocco; 4CNRS, UMR EDB, Toulouse, France; 5INRA, Meknes, Morocco; 6INRA, UMR AGAP, Montpellier, France; 7UCAM-FSTG, Marrakech, Morocco; 8UAE-FST, Tetouan, Morocco. Plant-parasitic nematodes significantly contribute to economic losses in the ten top olive producing countries in the world, especially in the Mediterranean basin (Spain, Italy, Greece, Tunisia, and Morocco). Diversity and structures of plant-parasitic nematode communities respond to evolutionary, environmental and anthropogenic forces. Instead of controlling the main pathogenic nematode species as usual, one of the innovative strategies to control plant-parasitic nematodes would be to manage diversity in communities in order to lead them to be less pathogenic. The present study aims at understanding the contribution of olive domestication and human impacts on the plant-parasitic nematode communities by analyzing the diversity of plant-parasitic nematodes in cultivated olive compared to wild olive in Morocco. Thus, 220 samples were collected in 2012 in several sites with cultivated and feral olive trees (i.e. wild olive resulting from cultivated olive) in the olive production areas located all along the Atlas foothills, as well as on wild olive Morphobiometric observations revealed a significant diversity of plant-parasitic nematodes, belonging to 12 families and 28 genera. Our results showed the presence of genera Pratylenchus and Meloidogyne in all sampled systems that are known harmful taxa for agriculture especially in nurseries. These two genera were more abundant in the cropped systems. Principal component analysis revealed no significant effect of olive systems on the diversity of PPN in communities but on the PPN community patterns Other factors such as olive genotype, soil physic-chemical characteristics, geo-climatic characteristics, associated plants with olive trees will also be discussed as major factors affecting the plant-parasitic nematode community patterns.

134 Journal of Nematology, Volume 46, No. 2, June 2014 EFFECT OF THE MECHANICAL WOUNDING OF GREEN BEAN ROOTS ON JOINT INFECTION WITH MELOIDOGYNE JAVANICA AND MACROPHOMINA PHASEOLINA. Al-Nadhari, S. King Saud University, Yemen. The effect of the mechanical wounding of the roots of green beans, Phaseolus vulgaris, on the development and severity of the disease complex caused by Meloidogyne javanica and Macrophomina phaseolina was investigated in the greenhouse (25 ± 28C). Five treatments with five replicates each were used in a randomized complete block design. The treatments included: 1) control, 2) M. javanica, 3) M. phaseolina, 4) M. javanica + M. phaseolina without root wounding, 5) M. javanica + M. phaseolina with root wounding. Root wounding was done by piercing (twinge) the root tips with very fine needle just prior to inoculation by the nematode and fungi. Results showed that root piercing has increased the severity of the disease complex. Plant growth decreased and gall and charcoal –root indices increased. Nematode reproduction also has increased. For example: plant weight has reduced (P # 0.05) almost 50%, compared to the control, where root galling and disease index were increased. M. javanica + M. phaseolina plus root wounding gave the highest number of eggs/g roots and the highest fungus colonization of the roots, compared to the same treatment, but without root wounding. DETECTION OF SUBSTANCES RELATED TO THE RESISTANCE OF COTTON GENOTYPE TX 25 TO MELOIDOGYNE INCOGNITA RACE 3. Alves1, G.C.S., S. da Costa Santos2, P.H. Ferri2, V.H.S. Barbosa2, M.R. da Rocha2. 1 Instituto Federal Goiano, Rodovia Geraldo Silva Nascimento Km 2,5, CEP 75790-000 – Urutaı´, Goia´s, Brazil; 2Universidade Federal de Goia´s, Escola de Agronomia, Caixa Postal 131, CEP 74690-900, Goiaˆnia, Goia´s, Brazil. Plants produce a wide range of secondary compounds that are part of the arsenal against pathogens. These substances may be pre or post formed, or may exist in the plant regardless of the presence of the pathogen. The purpose of this study was to detect the biochemical activity in resistant wild cotton genotype TX 25, as compared to a susceptible cultivar, to Meloidogyne incognita race 3. Plants were grown under greenhouse conditions and each plant was inoculated with 5,000 eggs and J2. At 8, 24 and 35 days after inoculation (DAI) root extracts were prepared for detection and quantification of total phenols, total flavonoids, analysis of phenols by high performance liquid chromatography (HPLC) and metabolite quantification by nuclear magnetic resonance of hydrogen (RMN). Total phenols were higher at 35 DAI on both resistant and susceptible genotypes. Flavonoid glycosides and gossypol derivatives were quantified by HPLC. A flavonoid glycoside was considered one of the secondary compounds related to the resistance because it was higher on TX 25 in all evaluations. In NMR it was found that sugars also confer an initial protection to genotype TX 25. PLANT PARASITIC NEMATODES: A SERIOUS THREAT TO AGRICULTURAL CROPS IN THE KINGDOM OF SAUDI ARABIA. Al-Yahya, F.A. Department of Plant Protection, College of Agriculture and Food Sciences, King Saud University, P. O. Box 2460, Riyadh 11451, Saudi Arabia. Meloidogyne spp were first reported on tomato in 1957 in the Eastern region of Saudi Arabia. Since then many other plant parasitic nematodes have been reported causing serious losses to many economically important agricultural crops. Until now several efforts have been made for the eradication of these nematodes both in the greenhouse and in the field, but success has not yet been achieved. For successful and target-oriented control of theses nematodes, a comprehensive nematological survey must be carried out to identify the various plant-parasitic nematodes based on their prevalence and economic importance. Thus we have recorded a total of 30 nematode genera from various regions of the kingdom. These genera have been classified into three groups. The first group includes the genera of the highest prevalence and economic impact (e.g. Meloidogyne spp., Heterodera, Pratylenchus and Tylenchulus). The second group represents the genera with relatively medium importance and prevalence (e.g. Rotylenchulus, Helicotylenchus, Rotylenchus and Tylenchorhynchus.) while the third group includes the nematodes of least prevalent and economic importance. We believe this information would be of great importance to lay out meaningful control strategies. NEMATODE INTERCEPTIONS IN INTERNATIONAL TRADE OF PLANTS FOR PLANTING. Anthoine1, G., B. Niere2, L. den Nijs3, T. Prior4, L. Pylypenco5 and N. Viaene6. 1ANSES, 7 rue Jean Dixme´ras, 49044 Angers Cedex 01, France; 2JKI-Plant Health Institute, Messeweg 11/12, 38104 Braunsweig, Germany; 3NPPO-NRC, Geertjesweg 15, 6700 HC Wageningen, Netherlands; 4FERA, 02FAO1/5, Sand Hutton, YO41 1LZ, York, United Kingdom; 5Instutite of Plant Protection-dept of Plant Quarantine, 33 Vasilkovskaya Str., 03022 Kiev 22, Ukraine; 6ILVO-Plant-Crop Protection, Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium. Trade of plants for planting and ornamentals in Europe has increased dramatically in the last few decades. For instance, numbers of Ficus imported to the Netherlands has increased from 1.6 million in 2006 to 3.8 million in 2010. Traded plants originate from different countries of which some play a major role at present (e.g. China). According to phytosanitary regulations (ISPM no 1 and the EU council directive 2000/29/EC) plants shall be free of harmful organisms or treated in such a way that no harmful organisms shall be present in order to prevent their introduction, spread and establishment. Absence of harmful organisms has to be checked by the importing country and when interceptions occur, appropriate measures should be taken. The latter is difficult to define. The findings should be officially notified by the European Union (EU) countries to the EU-commission via EUROPHYT. In this presentation we show what some countries (France, UK, Belgium, Germany,

Abstracts 135 Netherlands and Ukraine) have intercepted during the last decade. We will show the diversity of the nematode pressure in the framework of global trade and that different approaches in the countries on how to deal with inspections and treatment might be reflected in the data. A Dutch survey on import of 21 different countries over 3 years showed that around 20% of the samples contained EU-quarantine nematodes. Other nematodes of phytosanitary significance (based on PRA quick scan) were found in 11% of the samples. We will discuss what this data mean in terms of threats towards the importing countries. KNOCKING-DOWN MELOIDOGYNE INCOGNITA PROTEASES BY PLANT-DELIVERED DSRNA HAS NEGATIVE PLEIOTROPIC EFFECT ON NEMATODE VIGOR. Antonino-de-Souza Jr1,2, J.D., R.R. Coelho1,2, I.T. Lourenc xo1,2, 3 4 2 2 2 5 R.R. Fragoso , A.A.B. Viana , L.L. P. Macedo , M.C.M. Silva , R.M.D.G. Carneiro , G. Engler , J. de AlmeidaEngler5 and M.F. Grossi-de-Sa2,4. 1Graduate Program in Biology Molecular, Universidade de Brası´lia, Brası´lia, Distrito Federal, Brazil; 2 Embrapa Recursos Gene´ticos e Biotecnologia, Brası´lia, Distrito Federal, Brazil; 3 Embrapa Cerrados, Planaltina, Ditrito Federal, Brazil; 4Graduate Program in Genomic Sciences and Biotechnology, Universidade Cato´lica de Brası´lia, Brası´lia, Distrito Federal, Brazil; 5Institut National de la Recherche Agronomique, UMR 1355 ISA/Centre National de la Recherche Scientifique, UMR 7254 ISA/Universite´ de Nice-Sophia Antipolis, UMR ISA, Sophia-Antipolis, France. Meloidogyne incognita causes major damage and yield losses in numerous important crops worldwide. Examination of the M. incognita genome revealed a substantial number of proteases belonging to five different catalytic classes. Several reports indicate that M. incognita proteases could play essential roles in nematode parasitism, besides their function in digestion of giant cell contents. Nevertheless, the precise roles of these proteins during parasitism are still unknown, making them interesting targets for gene silencing to address protein function. We have knocked-down an aspartic (Mi-asp-1), a serine (Miser-1) and a cysteine protease (Mi-cpl-1) by RNAi interference to investigate the function of these enzymes during the plantnematode interaction. Tobacco lines expressing dsRNA for Mi-ser-1 (dsSER), Mi-cpl-1 (dsCPL) and for the three genes together (dsFusion) were generated. Interestingly, nematodes that infected plants expressing dsRNA for proteases produced a reduced number of eggs. In addition, nematode progeny matured in dsSER plants had reduced success in egg hatching, while progeny resulting from dsCPL and dsFusion plants were less successful to infect wild-type host plants. Quantitative PCR analysis confirmed a reduction in transcripts for Mi-cpl-1 and Mi-ser-1 proteases. Our data suggest that these proteases are involved in different processes during nematode development, like nutrition, reproduction and embryogenesis. A better understanding of nematode proteases and their possible role during a host plant-nematode interaction might help to develop novel strategies for phytonematode control. NATURALLY OCCURRING ENDOPHYTES ASSOCIATED WITH WHEAT AS BIOCONTROL AGENTS AGAINST CEREAL CYST NEMATODES. Ashrafi1,2,3, S., W. Maier2, G. Erginbas-Orakci1, M. Finckh3, and A. Dababat 1. 1 CIMMYT-Turkey, P.K. 39 Emek 06511, Ankara, Turkey; 2Julius Ku¨hn-Institut, Federal Research Centre for Cultivated Plants, Institute for Epidemiology and Pathogen Diagnostics, Messeweg 11/12, 38104 Braunschweig, Germany; 3University of Kassel, Faculty of Organic Agricultural Sciences, Ecological Plant Protection, Nordbahnhofstr. 1 a, 37213 Witzenhausen, Germany. Cyst nematodes (Heterodera and Globodera spp.) are distributed worldwide and have a great impact on crops. Within the Heterodera avenae complex, H. avenae, H. filipjevi and H. latipons are the most important cereal cyst nematode species (CCNs) which can cause significant economic losses. Despite the difficulties in controlling of cyst nematodes, biological control can be applied alone or in integrated management approaches to reduce the damages of nematode. Preliminary results obtained from a field experiment in Turkey indicated that there might be suppressive activities, potentially caused by fungi or bacteria, which can affect the nematode populations of CCNs. Therefore the present study is conducted with the following aims: i) isolation and identification of naturally occurring endophytic and parasitic fungi and bacteria from wheat and Heterodera filipjevi ii) evaluation of the biocontrol effects of the isolated fungal and bacterial species on H. filipjevi. To this aim a total of 175 bacteria and fungi associated with wheat and cysts of H. filipjevi were isolated from nematode infested fields. Isolates were purified and in vivo screening experiments will be conducted to evaluate their biocontrol potential towards the nematode. The total number of cysts extracted from both soil and the root system was counted and plant growth parameters including fresh root and shoot weight as well as plant height were measured. Preliminary results indicated that a total of 19 isolates including ten fungal and nine bacterial isolates are promising to be used in further detailed studies. PATHOGENICITY AND MANAGEMENT OF MELOIDOGYNE INCOGNITA ON PITTOSPORUM TOBIRA IN FLORIDA. Baidoo1, R., R.H. Stamps2, and W.T. Crow1. 1Entomology and Nematology Department, University of Florida, Gainesville, FL 32611; 2MidFlorida REC, University of Florida, Apopka, FL 32703, USA. Pittosporum tobira is grown primarily as a cut foliage crop in Florida. However, many P. tobira farms have been abandoned owing to severe root-knot nematode infestation and lack of effective management tactics. Hence, research is underway by the University of Florida to investigate the pathogenicity of root-knot nematode Meloidogyne incognita on P. tobira and to identify pesticides or bio-pesticides that could be used to manage these nematodes on perennial cut foliage crops, including P. tobira. This research includes on-farm, small plot, and greenhouse experiments. Pesticide and

136 Journal of Nematology, Volume 46, No. 2, June 2014 biopesticide treatments evaluated include commercial formulations of spirotetramat, furfural, and Paecilomyces lilacinus strain 251. Treatment applications were made during the spring and fall seasons according to manufacturer’s specifications. Efficacy was evaluated based on J2/100cm3 of soil, J2/g of root, and crop yield (kg/plot) or plant growth (canopy volume/ plant). Greenhouse results reveal that M. incognita causes significant growth reduction in P. tobira. It was also found that, M. incognita infection increases incidence and severity of leaf spot disease caused by Cercospora sp. on this crop. On-farm and small plot trial results indicate that furfural and P. lilacinus were effective in reducing population density of M. incognita on P. tobira. SURVIVAL AND INFECTIVITY OF THREE ENTOMOPATHOGENIC NEMATODES SPECIES ON TRINERVITERMES OCCIDENTALIS (ISOPTERA: TERMITIDAE) AS AFFECTED BY PESTICIDES, SOIL TEMPERATURE AND MOISTURE. Baimey1, H., L. Zadji1, L. Afouda1, M. Moens2 and W. Decraemer2, 3. 1University of Parakou. BP 123. Parakou, Route de l’Okpara, Benin; 2Ghent University, Ledeganckstraat 35, 9000 Ghent, Belgium; 3Royal Belgian Institute of Natural Sciences, Vautierstraat 29, 1000 Brussels, Belgium. Experiments were conducted to studythe survival of four Beninese isolates of entomopathogenic nematode (Heterorhabditis indica Ayogbe1, H. sonorensis Azohoue2, H. sonorensis Ze2, and Steinernema sp. Bembereke) in aqueous solutions of three pesticides i.e. fipronil (50g/l), sulfur (80%) and glyphosphate (41%). Secondly to test the infectivity of the isolates following fipronil treatment on the last instars Galleria mellonella. Thirdly to test the effect of temperature on the virulence of the EPN isolates applied on the termite Trinervitermes occidentalis and lastly to test the effect of soil moisture content on termites. The time at which 50% nematodes survived (ST50) after a pesticide treatment varied with nematode isolate from 1.38h (H. indica in sulfur at 2-fold rd) to 15.53h (H. indica in glyphosate at 0.5 fold rd). The higher the pesticide dose, the lower the ST50 value for each of the nematode isolates. The increase of both exposure time and fipronil concentration had negative impact on the efficacy of tested EPN isolates on G. mellonella. Termite mortality differed significantly between temperatures and soil moisture content for all tested EPN isolates. The greater the temperature at which termites and nematodes were exposed, the lower termite mortality with Azohoue2 giving highest mortality values at 278C (91%), 308C (69%), 338C (67%) and 358C (65%). However, the effect of soil moisture content on the EPN efficacy did not vary with the IJ concentration. CURRENT TRENDS IN PHYLOGENY AND CLASSIFICATION: OBSERVATIONS FROM A MORPHOLOGICAL PERSPECTIVE. Baldwin, J.G. and T. Pereira. Department of Nematology, University of California, Riverside, California 92521, USA. Challenges in phylogeny/classification are little changed from those articulated >25 years ago in ‘‘Vistas of Nematology,’’ but new priorities and tools point to major advances. These include increasingly rigorous molecular phylogenies using multiple genes, broader taxon representation, greater computer power and increasing analytical sophistication. Molecularbased phylogenies often point to errors in traditional classifications and raise related questions of morphological homology that are increasingly addressed with emerging microscopy tools supporting comparative digital imaging, analysis and reconstructions. Furthermore, genomics underlying phenotypic expression and evolution is adding a new dimension to recognizing homology, phenotypic plasticity and mechanisms/patterns of evolution. Often detailed morphological reconstructions first demonstrated with complex approaches can later be assessed with simpler techniques and broader taxon representation involving, for example, specific staining and confocal microscopy. Beyond phylogenetic classification increasing computational power and advances in digital microscopy/imaging have become important tools for assessing and vouchering biodiversity, including ‘‘reverse taxonomy’’ for efficient new species discovery. These tools have capability for improved global access to collections/metadata, vouchers and valuable rare specimens (e.g. types) that may include remote access to mounts on microscopes in distant collections. Twenty-five years ago challenges included how to approach inferring phylogeny, how to integrate traditional morphology with emerging molecular data, and how to express new findings in the inherently conservative process of classification. An overarching challenge was filling gaps in phylogenetic representation that demand progress toward globally inventorying and describing putatively more than 1,000,000 nematode species. These challenges remain, but emerging tools and techniques hold increasing promise toward further resolution. THE TANGLED WEB WE WEAVE: HUMAN MEDIATED SPREAD OF NEMATODES VIA TRADE NETWORKS. Banks1,2,3,4, N.C., M.E. Hodda1,2,3, S.K. Singh1,3,5, D.R. Paini1,2,3, K.L. Bayliss4. 1Plant Biosecurity Cooperative Research Centre, Bruce, ACT, 2617, Australia; 2CSIRO Biosecurity Flagship, Dutton Park 4102, Australia; 3CSIRO Ecosystem Sciences, Canberra, ACT, 2601, Australia; 4School of Veterinary and Life Sciences, Murdoch University, Murdoch 6150, Australia; 5Graham Centre for Agricultural Innovation, Wagga Wagga, New South Wales, 2678, Australia. Quarantine and hygiene are major measures for preventing damage from plant-parasitic nematodes, but there is little information on nematode spread, especially related to rapidly increasing trade associated with globalization. To investigate nematode movement associated with traded plant material, nematodes were sampled from markets of different sizes and types in several countries, together with the origins and destinations of the produce and volumes traded. Nematodes were

Abstracts 137 found consistently on various crops and associated soil. All trophic groups were represented. Markets differed in the distances produce travelled to get there, with some trading mostly local material, others trading mostly material from distant places, and others with a mix of the two. These patterns of movement can be analysed using network models and mapping software to suggest the speed, distance and amount of nematode spread using these pathways. This information will enhance biosecurity and quarantine through better targeting of surveillance and preventative measures. RELATIONSHIPS BETWEEN THREE MELOIDOGYNE SPECIES ON SUGARCANE. Barbosa1, N.M.R., E.M.R. Pedrosa1 and A. Chaves2. 1Universidade Federal Rural de Pernambuco, Rua Dom Manoel de Medeiros, s/n, Dois Irma˜os, Recife/PE, 52171-900, Brazil; 2Estacxa˜o Experimental de cana-de-acxu´car do Carpina, Rua Angela Cristina C. Pessoa de Luna, s/n, Carpina/PE, 55810-700, Brazil. The root-knot nematodes Meloidogyne incognita and Meloidogyne javanica cause high losses on sugarcane fields around the world. Reports of Meloidogyne enterolobii parasiting the crop in north-eastern Brazil worried producers because of the fast dissemination and extreme severity of this species in guava fields. This study had as objectives comparing reproduction of M. incognita, M. javanica and M. enterolobii in association or individually on sugarcane variety RB 867515. This involved evaluating M. enterolobii penetration (3, 7, 10, 17, 24, 31, 38 and 45 days after inoculation), development and life cycle on the variety RB 98710, using M. javanica as the control. The last objective being) evaluating effect of M. enterolobii initial density (0, 20,000; 50,000 and 100,000 eggs per plant) in relation to M. incognita on RB 98710 at 120 days after inoculation. RB 867515 was a good host for M. incognita and M. javanica, but not for M. enterolobii, permitting free reproductions for the former two species. Esterase phenotypes presented quantitative variations in the ratios of the three species recovered at the end of the experiment, indicating M. incognita prevalence over the other species. At 45 days, M. javanica complete the life cycle in contrast to the low number of vermiform and swollen juveniles and no adult female of M. enterolobii found in roots of RB 98710. Meloidogyne enterolobii initial population did not affect the early development of RB 98710 120 days after inoculation, presenting a reproductive factor always lower than 1. CELLULAR MECHANISMS IN PLANTS INFECTED BY CYST NEMATODES. Baum, T.J. Department of Plant Pathology and Microbiology, Iowa State University, Ames, Iowa, 50011, USA. Cyst nematodes secrete effector proteins into host root cells, and these proteins are instrumental in the formation of feeding cells and mediating susceptibility through the suppression of plant defences. Consequently, the identification of effector proteins and their functions is of high importance. We have developed an effector identification method in which whole nematode glands are purified and subjected to RNA extraction for transcriptomic analyses. In addition to effector identification, the characterization of their functions in plant cells is of highest interest. In one example that we studied, a cyst nematode effector protein is phosphorylated by a cytoplasmic plant kinase, which results in translocation of the effector into the plant nucleus. There, the effector interacts with a plant transcription factor, likely resulting in gene expression alterations. In another example we discovered that as a result of cyst nematode infection, and probably effector functions, microRNAs change expression in the developing syncytium. In particular, the microRNA396 regulatory network has strong impact on cyst nematode parasitic success. This and other microRNAs that we have shown to change expression in the syncytium, thus, represent powerful molecular targets for cyst nematode effectors to modulate plant cell development. Ongoing functional characterizations of microRNAs as well as identification and characterization of nematode effectors promise to provide the mechanistic understanding required to devise novel control mechanisms. OBSERVATIONS ON NEMATODE-SUPPRESSIVE SOILS LEADING TO IPM APPROACHES. Becker, J.O. Department of Nematology, University of California, Riverside, California 92521, USA. Biological nematode population suppression has been recognized in a number of cyst nematode-infested fields. A common denominator is often that susceptible crops are grown for years in monoculture or in narrow rotations of the same or similar plants. We have shown that in a Heterodera schachtii-suppressive field on the Agricultural Operations, University of California, Riverside, the suppressive agent is the ascomycete Dactylella oviparasitica. It attacks primarily late juvenile stages of sedentary sugar beet cyst nematodes and the young females. However its population appears to decline rapidly after discontinuing the monoculture. The fungus then requires at least one or two nematode population cycles on the next host to build up again to a population suppressive level. This leaves beets or cruciferous crops at the seedling stage exposed to unmitigated attack of invasive second-stage juveniles. Preventing this early nematode-caused damage as well as subsequent secondary microbial attack of the nematode-weakened tissues is difficult to achieve with an augmentation of nematodedestroying microorganisms before or at seeding. Consequently, this approach is most effective in conjunction with other integrated pest control modules such as resistant or tolerant host crops, and nematode-resistant cover crops. We report here on the utility of seed coating with the biorational nematicide Avicta (a.i. abamectin) that provides early broad-spectum seedling protection followed by castration or destruction of the nematode females and their eggs by seed treatment-compatible hyperparasites.

138 Journal of Nematology, Volume 46, No. 2, June 2014 SEED TREATMENTS AGAINST PLANT PARASITIC NEMATODES: WHEN A LITTLE GOES A LONG WAY. Becker1, J.O., and H.V. Morton2. 1Department of Nematology, University of California, Riverside, CA 92521; 2VIVA Inc., Greensboro, NC 27410. Expensive hybrid and genetically modified seed make healthy seedling establishment an economic priority. The increasing popularity of multi-component seed treatments to deliver seed and seedling protection against a wide-range of pests and diseases has been primarily driven by a multitude of regulatory and economic factors that resulted in user-safe, cost-effective and environmentally sound products. Nematicides are the most recent addition to seed treatment cocktails, following long after fungicides and insecticides. The nematicidal seed treatment Avicta in particular has proven its efficacy in seedling protection of many crops against various plant parasitic nematodes. Avicta has also been shown to protect against secondary fungal diseases and accelerated root senescence by microorganisms that opportunistically invade nematode-weakened tissues. Furthermore, its active ingredient abamectin, comprised of natural metabolites of Streptomyces avermectinius, has no known antimicrobial activity. It therefore allows for the combination with compatible microbial control agents to increase nematicidal efficacy, to extend the protective duration and to provide a sustainable solution to the control of plant parasitic nematodes. DETERMINATION OF THE EFFECT OF GROWING PARTIALLY RESISTANT POTATO VARIETIES ON THE POPULATION DENSITIES OF GLOBODERA PALLIDA. Been1, T., C. Schomaker1 and N. Matkaris2. 1Plant Research International, Plant Sciences Group, Wageningen University and Research Centre, P.O. Box 16, 6700 AA Wageningen, The Netherlands; 2Nematology section, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent Belgium. Twenty fields, where starch potatoes are grown every second year, were planted twice with the partially resistant potato cultivar Elles (RS = 25%) in 1993 and 1995. Four of these fields were resampled in 2012, to investigate the long-term effect of planting partially resistant potato cultivars on population densities of Globodera pallida. All fields were of 1 ha size, divided in plots of 10 x 16 meter (the latter in the direction of cultivation) and sampled to estimate population densities. The number of cysts (500 g soil)-1 during this first period of 4 years did not decrease. In 2012, three fields showed a marked decrease and field four, where resistant cultivars were alternated with susceptible ones, a moderate decrease. Ultimately, cyst densities decreased from 690, 1007, 405 and 1150 to 169, 260, 57 and 761 cysts (500 g soil)-1, respectively. The first use of Elles kept population densities either in check or caused them to decline and, therefore, was larger than the combined effect of natural decline and soil fumigation. Remarkable was the high natural decline (77%) observed in the non-host year of 1994. After the second crop, the nematode populations on two fields were more than twice as high as on the two other fields, probably due to the higher virulence of the local populations of the first two fields as estimated in a pot experiment. After 20 years, in all fields, population densities were comparable and had declined from 151, 110, 50 and 91 to 0.18, 0.10, 0.01 and 0.53 J2 (g soil)-1, respectively. EXPLORING THE CROP MICROBIOME FOR NEMATICIDAL ACTIVITY. Beilinson, V., D. Tomso, S. Uknes, E. Ward and F.M. Leach. AgBiome, 104 T.W. Alexander, Research Triangle Park, North Carolina 27709, USA. Plant parasitic nematodes remain difficult to control for most crops. Chemical solutions face increasing regulatory pressure, and genetic resistance is often limited in scope and durability. AgBiome is leading an effort to discover new solutions for significant agricultural pests by conducting systematic screens for microbes with activity against plant parasitic nematodes. We have assembled a large collection of microbes from the microbiomes of major row crops and are characterizing isolates for activity against nematodes. We have also sequenced the genome of thousands of microbial isolates and are using sequence data to guide both strain isolation and active strain discovery. Plant microbiome communities are diverse, and we show that populations vary across plant-associated environmental compartments. Many of these communities, including those from endophyte and rhizosphere compartments, are likely to exert significant influence on nematode infestation. Screening efforts and new isolates for nematode control are presented as a component of an integrated biologicals product discovery pipeline. NEMATODE COMMUNITY STRUCTURE IN CORN-BASED CROPPING SYSTEMS THEN (1995-1997) AND NOW (2012-2013). Bender, B., and A. MacGuidwin. Department of Plant Pathology, University of Wisconsin, 1630 Linden Drive, Madison, Wisconsin 53705, USA. Nematode community structure was studied in a long term trial of grain- and forage-based cropping systems in southern Wisconsin. Three systems were sampled during the corn phase in 1995-1997 and 2012-2013: 1) continuous corn with high pesticide and fertilizer inputs and chisel plow tillage, 2) corn grown with low inputs and rotary tillage in rotation with soybean/winter wheat, and red clover, and 3) corn grown with low inputs, manure, and rotary tillage in rotation with alfalfa for two years. A forage pasture system planted with a mixture of red clover, orchard grass, timothy, and brome that was rotationally grazed by cows was also sampled. Soil samples assayed for nematodes using a combination of sieving, centrifugation and incubation methods revealed 28 nematode families at the study site and nematode abundance ranging from

Abstracts 139 500 to 5000 nematodes per 100 cm3 soil. Corn yield increased in all three systems from 1989 to 2013, but system 2 consistently produced less yield than systems 1 and 3. In 1997, Maturity Indices distinguished system 1 from system 3. Maturity Indices were lower in all corn based systems as compared to pasture for 1995-1997. Preliminary faunal analyses of the current data set suggest that nematode community structure has not become more different in the systems over time. POPULATION DYNAMIC OF RING NEMATODE IN PEACH ORCHARD MANAGED WITH CASTOR BEAN CAKE AND MILLET CROP. Bernardo1, J.T., A.C. Krolow2 and C.B. Gomes2. 1PPGFS/Universidade Federal de Pelotas, Campus Universita´rio s/n C. P. 354, Pelotas/RS, Brazil; 2Embrapa Clima Temperado, Cx Postal 403, Pelotas/RS, Brazil. The Peach Tree Short Life syndrome associated with the ring nematode, Criconemoides xenoplax has been a problem in peach orchards in southern Brazil since the 1980s. Therefore, the influence of incorporating of castor bean cakes and soil cropping with black oat (Avena strigosa) and millet (Pennisetum americanum) was investigated in a peach orchard ’Rubimel’ naturally infested by C. xenoplax. During the spring/summer and autumn/winter seasons, 20 kg of castor bean cakes were incorporated into the soil at level of 10kg castor bean cake/m2 in plots with five peach trees. Just after the residue application, black oat or millet were seeded in the plots. Peach trees maintained without weeds were used as control. The experiment was carried out during 2012 and 2013 using a randomized design with six replications. Four months after the application of the treatments, soil samples were collected before and 30 days after plant incorporations for evaluation of C. xenoplax populations using regression analysis. Additionally, the chemical characteristics of peach fruits were also evaluated. Independent of the treatment, the C. xenoplax populations behaved similarly. In the spring-summer seasons the populations decreased in the soil and increased during the autumn-winter season. However, the nematode populations were lower in the pots where the plants received the residue associated with cover crops. Evaluating the fruit quality, there were no difference in the pH, total titratable acidity and ascorbic acid between the different treatments, but in the plots with castor bean cake and cover crops a significant reduction of total soluble solids was verified and it can possibly be explained by increased nitrogen from the castor bean cakes. PAECILOMYCES LILACINUS (=PURPUREOCILLIUM LILACINUM) AS A NEMATODE BIOCONTROL AGENT. S.D. Berry. BASF Agricultural Specialities (Pty) Ltd., 91 Clifton Canyon Drive, Gillits, 3610, South Africa. Nematodes are a constant threat to crop production worldwide. Various management strategies have been developed over the years, with most of the emphasis being on chemical nematicides and varietal resistance. However, with the demise of many of the traditional chemical solutions, the identification and management of these pests is becoming increasingly more important. In the last few years much interest and investment has gone into researching biological control of nematodes. A number of different biological control organisms have been researched and a few developed into commercial products (e.g. Paecilomyces lilacinus, Bacillus firmus, Pochonia chlamydosporia, Pasteuria penetrans). The aim of this talk will be to discuss the history, modes-of-action, nematode targets, crops, commercial products and efficacy of Paecilomyces lilacinus (also known as Purpureocillium lilacinum) as a nematode biocontrol agent. RECENT METHODOLOGICAL AND THEORETICAL ADVANCES FOR SPECIES DELIMITATION IN CONTEMPORARY NEMATODE TAXONOMY. Bert1, W., D. Slos1, T. Janssen1, P. Fonderie1, H. Steel1 and W. Decraemer1,2. 1 Nematology Research Unit, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium; 2 Royal Belgian Institute of Natural Sciences, Department of Invertebrates, Vautierstraat 29, B-1000 Brussels, Belgium. Only a small fraction of the estimated existing nematode species has been described. To grasp the overwhelming nematode diversity, some nematologists still have confidence in methods that date back to the 17th century, while others want to abandon species as fundamental entities of diversity. We will discuss the necessity and shortcomings of traditional and new approaches. It is obvious that nematode taxonomy is confronted with the challenge to fully incorporate new theory, methods and data from disciplines that study the origin, boundaries and evolution of species. These recent methodological advances hold promise for species delimitation methods that reflect ‘‘true’’ speciation events. However, these advanced methods have their limitations in daily taxonomic practise, and therefore it is evident that forwarding one best possible taxonomical method is currently impossible. We advocate the simultaneous use and efficient integration of slow integrative taxonomy and fast molecular operational taxonomical units. Examples of our own approaches, targeting a comprehensive and reliable description of nematode biodiversity, will be illustrated using case studies on free-living, plant-parasitic, and facultatively animal parasitic nematodes. These efforts are based on a combined acquisition of informative sequences, ecological and morphological data; and integrated in phylogenetic frameworks and supported by an appropriate databank. Finally, the link of our taxonomical work with other ongoing research projects will be highlighted. IS INTROGRESSION BREEDING ANSWER TO NEMATODE RESISTANT SUGARCANE IN AUSTRALIA? Bhuiyan1, S.A., B.J. Croft1, E. Wong1, P. Jackson2 and G.R. Stirling3. 1Sugar Research Australia, Woodford, Australia; 2 CSIRO Townsville, Australia; 3Biological Crop Protection, Brisbane, Australia. Root knot (Meloidogyne javanica) and root lesion (Pratylenchus zeae) nematodes are causing in excess of A$82 million losses to the Australian sugar industry annually. No commercial sugarcane varieties are resistant to these nematodes. A

140 Journal of Nematology, Volume 46, No. 2, June 2014 collaborative introgression program with Chinese institutes has used new sources of germplasm, Erianthus arundinaceus and Saccharum spontaneum clones to generate over 100 new families. In 2012, approximately 150 clones from different introgression families have been screened in a glasshouse for resistance to M. javanica and P. zeae .The initial nematode population used for inoculation of test clones (Pi) and final population recovered after 12 weeks (Pf), were used to determine the multiplication factor (MF) =Pf/Pi. For both nematodes, the lowest multiplication factors were observed on basic E. arundinaceus (MF = 3 - 7) and S. spontaneum (MF =1.8 - 6) clones, indicating moderate to high levels of resistance. Average levels of resistance tended to decrease with successive backcrosses between the wild species and commercial sugarcane. However, approximately 30% of backcross-three (BC3) populations of E. arundinaceus showed moderate resistance (MF Sequential use of a single R-gene introgressed in a susceptible background. We are currently analysing (1) the impact of agronomic practices on the parasite pressure in the soil and on ecological diversity including other nematode species, (2) the linkage between reduction of Meloidogyne populations in the soil and increase of R-genes durability, and (3) the sustainability assessment of such varietal and technical innovations. Multidisciplinary approaches were combined to foster synergistic and long-term goals. The diversity of partners and associated forces brought all the complementary expertise needed for answering specific short-term questions as well as generic mid- to long-term expectations. OCCURRENCE, DISTRIBUTION, AND IMPACT OF NEMATODES IN SOYBEAN FIELDS IN THE SOUTHERN UNITED STATES. Donald1, P., K. Lawrence1, T. Kirkpatrick2, B. Kemerait3, J. Bond4, D. Herschman5, C. Overstreet6, A. Wrather7, G. Lawrence8, S. Koenning9, P. Adugelo10 and C. Canaday11. 1Entomology & Plant Pathology, Auburn University, Auburn, Alabama, USA; 2Southwest Research & Extension Center, University of Arkansas, Hope, Arkansas, USA; 3Plant Pathology, University of Georgia, Tifton, Georgia, USA; 4Plant Pathology Southern Illinois University, Carbondale, Illinois; 5University of Kentucky Research & Education Center, University of Kentucky, Princeton, Kentucky, USA; 6Plant Pathology and Crop Physiology Louisiana State University, Baton Rouge, Louisiana, USA; 7 Plant Sciences University of Missouri, Missouri, USA; 8Biochemistry, Molecular Biology, Entomology & Plant Pathology Mississippi State University, Mississippi State, Mississippi, USA; 9Plant Pathology North Carolina State University, Raleigh, North Carolina, USA; 10School of Agricultural, Forest, and Environmental Sciences, Clemson University, Clemson, South Carolina, USA; 11Entomologyu & Plant Pathology, University of Tennessee. Jackson, Tennessee, USA. Changing crop management practices throughout the southern soybean production area stimulated a survey of plantparasitic nematodes present in soybean production fields with an eye toward the consequences of common crop hosts in the potential management strategies. Soil samples were collected from each state (150/state) over a two-year period. Data collected included genera of plant-parasitic nematodes, soil texture, and GPS. Root-knot, reniform and soybean cyst nematodes were identified as the most potentially damaging nematodes present in these fields. Distribution of these three nematode genera was not uniform between states nor within states. Soil texture data showed distribution of these nematodes across multiple soil types but higher levels in certain soil types indicating certain soil types were more conducive to higher reproduction. Microplot and field studies aimed at these three genera showed soybean varietal differences in yield as well as nematode reproduction.

Abstracts 155 PLANT ESSENCES AS TOOL FOR MANAGEMENT OF THE NORTHERN ROOT-KNOT NEMATODE (MELOIDOGYNE HAPLA). Douda1, O., M. Zouhar2, E. Nova´kova´2, R. Pavela1. 1Crop Research Institute, Division of Plant Health, Drnovska´ 507, 161 06 Praha 6 – Ruzyneˇ, Czech Republic; 2Czech University of Life Sciences, Faculty of Agrobiology, Food and Natural Resources, Department of Plant Protection, Kamy´cka´ 129, 165 21 Praha 6 – Suchdol, Czech Republic. The root-knot nematode, Meloidogyne hapla is a prominent root vegetable pest in the Czech Republic. Effects of plant essences from Pongamia pinnata, Tagetes erecta and powder botanical pesticide Presto for M. hapla management on carrot (Daucus carota) were tested. Tests were conducted on container-grown carrots maintained under greenhouse conditions. The delivery method involved absorbing the tested essences in perlite particles placed in a substrate in the case of plant essences. The results showed a statistically significant reduction (P$0.05) of gall numbers of the carrots when treated by Presto. Similar results were achieved with Pongamia pinnata, Tagetes erecta essences. However evaluation of fresh and dry root weights showed certain signs of phytotoxicity caused by tested substances. Based on these results the management of M. hapla using tested treatments seems to be possible, however further research is desirable to determine the appropriate dosage needed for efficient and most appropriate delivery method of these substances under field conditions. EFFECT OF HYDROGEN CYANIDE TREATMENT ON BURSAPHELENCHUS XYLOPHILUS MORTALITY IN WOOD. Douda1, O., M. Zouhar2, M. Manˇasova´2, M. Dlouhy´3, J. Lisˇkova´3 and P. Rysˇa´nek2. 1Crop Research Institute Prague, Division of Plant Health, Drnovska´ 507, 161 06 Prague 6, Czech Republic; 2Czech University of Life Sciences Prague, Faculty of Agrobiology, Food and Natural Resources, Department of Plant Protection, Czech Republic; 3Lucˇebnı´ za´vody Draslovka a.s. Kolı´n. The Pine Wood Nematode (Bursaphelenchus xylophilus) is one of the economically most important quarantine nematodes in Asia and Europe. Bursaphelenchus xylophilus originates from North America and its introduction into Japan, China, Korea and Portugal led to devastation of the local conifers from the genus Pinus. Direct protection of pine trees in affected countries is expensive and difficult. As wood transported within the international trade plays major role in B. xylophilus spreading into new areas, attention should be given to wood treatment. With methyl bromide phasing out new chemicals should be investigated as an alternative. Gaseous hydrogen cyanide (HCN) seems to be promising as a methyl bromide alternative. For that reason the main aim of this study was testing of HCN potential as a nematode killing agent for wood treatment. Data considering HCN concentration in gas chamber during test fumigations and inside treated wooden blocks were also acquired. Results show overall good efficiency of HCN treatment on B. xylophilus mortality, total mortality was achieved after 8 hours lasting fumigation with the target HCN concentration of 12.30 g.m-3. GLOBAL METRICS TO MEASURE THE IMPACTS OF NEMATODES: THEN WHAT? Duncan, L. University of Florida, IFAS, Citrus Research and Education Center 700 Experiment Station Rd., Lake Alfred, FL 33850, USA. Understanding the economic importance of plant parasitic nematodes is necessary for the rational allocation of research and management resources by public or private organizations and growers. This talk reviews some of the key methodological contributions to crop loss assessment in nematology and highlights some previous estimates of regional and worldwide damage to various crops caused by plant parasitic nematodes. Deterministic and stochastic algorithms that employ loss assessment to forecast optimum management systems, and methods by which these models are being used for site-specific management of nematodes are also described. Advances in multivariate analytical methods have improved the capacity to identify biotic and abiotic properties that modulate relationships between nematodes and hosts. As the effects of key environmental properties becomes apparent, it will be possible to improve the efficiency of site-specific IPM and to ask new questions, such as how soils can be modified to mitigate losses to plant parasitic nematodes, or how to conserve and materially enhance the services of agriculturally beneficial nematodes such as those that are pathogens of insect pests. NEMATODES: INDICATORS OF METAL POLLUTION IN THE WONDERFONTEIN CAVE (SOUTH AFRICA). du Preez1, G., H. Fourie1, P. Theron1, and A. Swart2. 1North-West University, Unit for Environmental Sciences and Management, Private Bag X 6001, Potchefstroom 2520, South Africa; 2ARC-Plant Protection Research Institute (PPRI), Biosystematics Division, Private Bag X134, Pretoria 0001, South Africa. Subterranean ecosystems (caves) are highly sensitive to anthropogenic influences, however, are mostly overlooked in the application of environmental management practices. Fortunately, nematode community structures can be analysed in order to serve as ecosystem health indicators. The Wonderfontein Cave, located in the Witwatersrand Basin mining area, suffer an influx of water from the highly polluted Wonderfontein Spruit. The objectives of this study were i) to analyse the nematode community structures present within soil, sediment, water, and guano samples from nine selected sites along a transect within the cave ii) and to relate nematodal data to the concentration levels of selected metals observed. Standard methodology was applied for the extraction and morphological identification of nematodes; substrate samples were subjected to total acid digestion and metal concentrations were measured by using an ICP-MS. The faunal functional guild analysis was used to calculate nematode community structure and enrichment indices of all substrates. CANOCO’s Principal Component

156 Journal of Nematology, Volume 46, No. 2, June 2014 Analysis (PCA) was applied to study correlations between nematode trophic groups and selected metals. Preliminary results revealed that Chromium, Copper, Zinc, and Nickel concentrations, of especially sediment samples, were significantly higher than acceptable levels as reported by aquatic invertebrate quality guidelines. Nickel for example was elevated 65 times above the guideline’s threshold effect concentration. The nematode faunal analysis revealed that only single substrates (sediment and soil) from five sites host both enriched and structured communities, while the PCA indicated that the structure of the Wonderfontein Cave nematode communities is influenced by metal concentrations. SUPPRESSIVENESS OF ORGANIC AND CONVENTIONAL FARMING SOILS AGAINST HETERODERA AVENAE. Eberlein1, C., H. Heuer2, J. Moos3, H.M. Paulsen3, and A. Westphal1. 1Institute for Plant Protection in Field Crops and Grassland, Julius Ku¨hn-Institut, Messeweg 11/12, 38104 Braunschweig, Germany; 2Institute for Epidemiology and Pathogen Diagnostics, Julius Ku¨hn-Institut, Messeweg 11/12, 38104 Braunschweig, Germany; 3Thu¨nen-Institute of Organic Farming, Trenthorst 32, 23847 Westerau, Germany. Heterodera avenae induces losses on cereal crops worldwide. The aim of this study was to test if biotic soil factors affect the development of female populations of H. avenae in soil of different locations, crop management and cropping sequence history. Soil samples from four fields were placed in root observation boxes of 1-L capacity either untreated or heat-treated (1348C, 10 min). Three of these were from current cereal fields, one from a field with a history of limited frequency of cereal in the rotation sequence served as a control. Each box was planted to six seeds of nematode-susceptible Hordeum vulgare ‘Hanka’ and inoculated with 10,000 J2 of H. avenae. With first appearance, nematode females visible on the root surface were counted weekly. Maximum female numbers were detected from the second week of observation. Thereafter, numbers remained at a similar level. At all monitoring times, the numbers of females in the untreated soil were less than those in heated soil, indicating that a biotic component negatively affected H. avenae females on the roots. The highest number of females was found in the control throughout the monitoring period. Soils from conventional or organic fields and from the cereal monoculture all had similar female numbers. These soils with some suppressive capacity had regular cropping of the host plant in common differentiating them from the control soil. This suggested that regular cropping of a host plant may increase suppressiveness. The cause of these differences in female populations warrants further investigation. LIFE CYCLE OF GLOBODERA ROSTOCHIENSIS AND GLOBODERA PALLIDA ON EARLY POTATO CULTIVARS: TIMES ARE CHANGING. Ebrahimi1,2, N., N. Viaene2,3 and M. Moens1,2. 1Laboratory for Agrozoology, Ghent University, Coupure links 653, 9000 Ghent, Belgium; 2Institute for Agricultural and Fisheries Research, Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium; 3Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium. Potato cyst nematodes (PCN) are the most economically important nematode pests of potato. Early harvesting is part of a preventive management approach for very early potato varieties in Belgium. Tubers are harvested before June 20 because it is assumed that no cyst formation occurs by that date. However, this assumption might not be valid anymore because of climate changes, the availability of new cultivars (with new traits), and the increasing prevalence of G. pallida. Therefore, pot and field trials were conducted to verify the date when first cysts appear in Belgian growing conditions. The development of different populations of both species on 3 potato cultivars, viz. Eersteling (susceptible to PCN), Premie`re (resistant to G. rostochiensis) and Ambassador (partially resistant to G. pallida) was monitored in a growth chamber with simulation of seasonal field temperatures. On Eersteling and Premie`re, males, females and cysts of G. pallida were found 8, 9 and 11 weeks (May 15), respectively, after soil infestation with cysts, when the accumulated degree-days (DD) were 261, 322 and 450. No G. pallida females were observed on Ambassador. Males, females and cysts of G. rostochiensis were found 10, 10 and 12 weeks (May 22) after inoculation, respectively, corresponding with 292, 292 and 398 DD. Observations in two fields and in microplots under prevailing weather conditions in 2013 revealed that cysts of both Globodera spp. were formed on June 12. Our observations show that both species of Globodera develop earlier than was assumed based on previous data. DEVELOPMENT AND VALIDATION OF A QUANTITATIVE ASSAY FOR VIABILITY TESTING OF POTATO CYST NEMATODES. Ebrahimi1,2, N., N. Viaene2,3 and M. Moens1,2. 1Laboratory for Agrozoology, Ghent University, Coupure links 653, 9000 Ghent, Belgium; 2Institute for Agricultural and Fisheries Research, Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium; 3Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium. The potato cyst nematodes (PCN) can be managed by a series of integrated management practices. However, knowledge of the survival of the nematodes in the field would make the evaluation of management strategies more accurate. This study aimed to develop and optimize a quantitative method to determine the number of viable eggs of PCN based on trehalose present in live eggs. Trehalose was extracted from cysts, a dilution series of eggs formed and quantified. Results showed that more trehalose was detected when cysts were crushed than when left intact. Reaction volumes needed to be adapted to the number of eggs because small reaction volumes hampered an accurate quantitative extraction of trehalose; optimally 11 eggs per ml reaction volume can be used. The detection limit was defined as 5 viable eggs. The trehalose method was compared with visual assessment and the hatching test, commonly used methods to determine cyst viability, by using cysts recovered

Abstracts 157 from soil after exposure to soil amendments (pig slurry, cattle slurry, farmyard manure, chitin compost, nitrogen fertilizer, wood chip compost alone and in combination with biochar). There was no difference between the visual assessment and the trehalose method. The visual assessment, however, is time consuming, requires expertise and could be subjective. Less viable eggs were measured with the hatching test than with the other methods. Hence, the viability assessment method based on trehalose seems to be a practical, sensitive, fast and cheap technique. THE CHANGING SCOPE OF CROP PROTECTION. Eckstein, K.E. Bayer CropScience, P.O. Box 143, Isando, 1600, South Africa. Consumers and food companies are demanding safer food which includes the requirement for crop protection products with a more favourable environmental profile, less active ingredient per ha and restriction on the Pre Harvest Intervals (PHI) of applications while increasing standards on food security and food quality. Historically nematicides have required considerable care in the application, storage, handling and transport of thereof. Furthermore the rates are high and are open for misuse. During the recent years Bayer CropScience has followed a WHO Class 1 phase out strategy resulting in the the global phase out of TemikÒ (aldicarb) and divestment of Mocap (ethoprophos) and Nemacur (fenamiphos). Bayer CropScience has made substantial investments in the Research and Development of nematicides and nematode suppressing compounds which are effective but at the same time meeting modern user, environmental and consumer requirements. The most recent development has been the introduction of Velum (fluopyram) as a nematicide in Zimbabwe. It not only offers superior control but also the benefit of a use rate of maximum 2 kg/ha product in comparison to 25 kg of aldicarb. Furthermore biological compounds in combination with chemical offers are becoming an important addition to integrated nematode control strategies. This includes the introducion of Votivo (Bacillus firmus) as the seed treatment in PonchoÒ VotivoÒ in the United States of America and South Africa. To ensure the supply of an increasing demand in safe and secure food Bayer CropScience is engaged in sustainable partnerships between the farming community, food companies and input providers such as seed and equipment companies. HOW TO MAKE EPN USE MORE COST EFFECTIVE? Ehlers, R.-U. e;nema, GmbH, Klausdorfer Str. 28-36, D-24223 Schwentinental, Germany. Biological control measures are usually higher in costs than conventional synthetic chemical products. Either biological control products find a niche to control pests for which synthetic compounds cannot provide a satisfying solution or they have to compete with chemical products. The production cost for EPN has shown a marked decrease over the last decade. Reduction of application density is one important parameter to reduce costs. Reduction in nematodes per ha can be achieved by improving application technology or use of more virulent strains or species. Important progress was made by exploiting economies-of-scale. With growing markets, production capacity has more than tripled during the last decade. Process stability is another important element to reduce production costs. Progress in production technology is reported. Other important factors are storage and transport cost. Nematode products are shipped at temperature of 4-108C. To maintain temperature smaller batches are packed with ice and transported by courier services. Once larger quantities are handled, the use of cooled containers is possible and can help to reduce transport costs. Beneficial traits of EPN define biological limits of the products and breeding programmes can target, for instance, enhancement of storage or production potential or virulence. Thus a combination of technical and biological tools is available to improve competitiveness of EPN products. ENTOMOPATHOGENIC NEMATODES IN BIOLOGICAL PLANT PROTECTION. Ehlers, R.-U. e;nema, GmbH, Klausdorfer Str. 28-36, D-24223 Schwentinental, Germany. Biological control provides environmentally friendly and effective plant protection measures. Acceptance is steadily increasing as well as market share. Entomopathogenic nematodes (EPN) are an important element of biocontrol. In the past they have mainly been used in niche markets. During the recent years biotechnical production and downstream processing has improved and production capacities have more than tripled. Product quality has improved as well as transport logistics enabling provision of viable and effective material at the farmer´s gate. Application technology and timing contributed to increase affectivity of EPN products. Today EPN are also applied in out-door markets. An overview will be given on major current and future EPN markets. BIOLOGICAL CONTROL OF WESTERN CORN ROOTWORM LARVAE (DIABROTICA VIRGIFERA VIRGIFERA) WITH DIANEMÒ (HETERORHABDITIS BACTERIOPHORA). Ehlers, R.-U. e;nema, GmbH, Klausdorfer Str. 28-36, D24223 Schwentinental, Germany. The biocontrol product DianemÒ contains entomopathogenic nematodes, Heterorhabditis bacteriophora. It is officially registered in Austria as a plant protection product to control larvae of the corn rootworm (Diabrotica virgifera virgifera). Field results from Hungary, Austria and Italy applying 2 x 109 nematodes/ha per ha obtained the same control as with chemical seed dressings with neonicotinoides or with application of granular insecticides containing the pyrethroid Tefluthrin. Adapted application technology has been developed to apply nematodes with 200 liters of water/ha to the seeds.

158 Journal of Nematology, Volume 46, No. 2, June 2014 Although the insect larvae occur approximately a month later, the nematodes persist long enough to control the pest. Insects penetrate into the roots where they are not easily reached by insecticides, whereas nematodes follow the insects into the galleries and kill the larvae 2-3 days after infestation. Latest field results which have used the novel application technology will be presented. Since the seed treatment with neonicotinoides was banned by the European Commission in 2013, the correct use of the product DianemÒ was demonstrated to more than 2000 farmers in Austria and nematodes will be used commercially for the first time on larger scale against this invasive maize pest. USE OF FLUORESCENT DYES TO LOCALISE NEMATODES WITHIN ROOT TISSUE. Ehwaeti1, M.E.M. and V. Blok2. 1Omar Al Moukhtar University, Elbedi, Libya; 2The James Hutton Institute, Invergowire, Dundee, DD2 5DA, Scotland, UK. Nematodes comprise a very small amount of root tissue once with the tissue and are difficult to locate, particularly in roots that have limited transparency such as potato. The aim of this work was to label juvenile nematodes with a fluorescent dye prior to root inoculation and then to examine the roots with a fluorescence microscope to identify their location with root tissues. The dyes PKH26 which fluoresces red and PKH67 which fluoresces green were used to label nematodes. Nematodes were soaked in diluted dyes for 2-4 hours prior to inoculating roots. Sprouts of three potato genotypes, Desiree, and JHI breeding lines 11415 and 12601, were placed in vermiculite for 2-3 weeks to allow the production of roots several centimetres in length. Desiree is susceptible to both Globodera pallida and Globodera rostochiensis. Breeding line 11415 has H1 resistance to G. rostochiensis (Ro1) and H3 quantitative resistance to G. pallida. Breeding line 12601 has H3 quantitative resistance to G. pallida, but is susceptible to G. rostochiensis. Roots were then cleaned and transferred to petri plates containing 20ml of 20% w/v Pluronic F-127 gel. The medium was then allowed to solidify. Roots were observed with a fluorescent microscope for up to 14 days to identify where nematodes were within the root tissue. Root pieces (0.7cm) containing nematodes identified by their fluorescence, were excised using a scalpel and immediately fixed. The fixed root sections were then embedded in wax. The wax blocks were sectioned, sections were collected on microscope slides and then wax removed. These sections were observed with the fluorescence microscope. Several examples of nematodes within root tissue were observed. COMPARISONS OF NEMATODE ASSAY LABS EFFICIENCIES AND RECOMMENDATIONS FROM TEN NEMATODE ASSAY LABS UTILIZING SAMPLES FROM TURFGRASS. Eisenback1, J.D. and Paula Agudelo2. 1Virginia Tech, Department PPWS, Blacksburg, Virginia 24061 USA; 2Clemson University, Clemson, South Carolina 29634 USA. Nematode advisory programs access the damage caused by plant-parasitic nematodes to diverse crops including turfgrass. The objectives of this study were to determine the efficiency of ten nematode assay labs and to compare the recommendations for their management. Three locations were selected including the Virginia Tech Turfgrass Research Center, Augustine Golf Course in Stafford, Virginia and the Philadelphia Country Club in Gladwyne, Pennsylvania. Nematode assay laboratories from ten states, Alabama, Florida, Georgia, Louisiana, Massachusetts, Michigan, Mississippi, North Carolina, South Carolina, and Virginia processed soil from each of three sites of the bentgrass greens. Soil was thoroughly mixed and divided into 30 samples of 500 ml each and three samples from each site were mailed to each lab. They were processed using standard operating protocols and recommendations were based on the results. South Carolina reported the highest (567) counts from the Virginia Tech Turfgrass Research Center and Louisiana reported the lowest (13). Only Alabama and North Carolina suggested a possible problem caused by root-knot nematodes, even though South Carolina, and Florida found higher numbers. Total plant-parasitic nematodes extracted from the Augustine Golf Course by Michigan were the highest (317) and the lowest from Louisiana (13). The highest total nematode counts for Philadelphia Country Club were reported by Massachusetts and Michigan (14,239, 11,800, respectively), and the lowest by Mississippi and North Carolina (849, 260, respectively). These nematodes were suspected to be damaging by Alabama, Massachusetts, Michigan, South Carolina, and Virginia but not Florida, Georgia, Louisiana, Massachusetts, and North Carolina. These results show that nematode counts vary from state to state, as well as recommendations regarding their potential damage to turfgrass. HETERODERA SCHACHTI: TRANSCRIPTOME AND EFFECTOROME. Elashry1, A., S. Habash1, N. Ahmadinejad2, H. Schoof2 and F.M.W. Grundler1. 1INRES Molecular Phytomedicine, University Bonn, Karlrobert-Kreiten-Str.13, 53115 Bonn, Germany; 2INRES Crop Bioinformatics, University Bonn, Katzenburgweg2, 53115 Bonn, Germany. The beet cyst nematode Heterodera schachtii is a biotrophic sedentary nematode, which depends on a multinucleate, syncytial feeding structure. The nematode secretes effector proteins into a specifically selected host cell to induce and maintain the syncytium. We analysed the H. schachtii transcriptome to identify its secretome and functionally annotate it using the presence of signal peptide and the absence of transmembrane domain as filters. In this way, we identified nearly 500 putative secretory proteins specific to plant-parasitic nematodes (P-PSP). Among them, we found 24 novel candidates supported by previously known H. schachtii ESTs. However, these ESTs had not been identified before due to their truncated sequences. We also used the Biana Interlog Prediction server (BIPs) to identify proteins in Arabidopsis thaliana which putatively interact with our P-PSP. In this second approach we identified 40 putatively interacting proteins. Some of them

Abstracts 159 may play a role in pathways of metabolites that are enriched, and/or genes that are differentially expressed in A. thaliana after infection with H. schachtii. In order to further characterize candidate effector genes, we analyzed their expression level and localization during different stages of parasitism by qPCR and in situ hybridization, respectively. In this way we identified a number of pioneer effectors and putative interacting proteins within the host plant. A detailed analysis of the effectorome will contribute to further understand the molecular processes involved in feeding site formation and to develop novel managing systems for the control of H. schachtii. DOES SOIL WATER POTENTIAL CAUSE ALLOPATRY AMONG CLOSELY RELATED STEINERNEMATID SPECIES IN FLORIDA? El-Borai1,2, F., N. Killiny1 and L. Duncan1. 1University of Florida, IFAS, Citrus Research and Education Center, 700 Experiment Station Rd., Lake Alfred, Florida 33850, USA; 2Plant Protection Department, Faculty of Agriculture, Zagazig University, Zagazig, Egypt. A geospatial survey of soil properties and native entomopathogenic nematodes (EPNs) in Florida citrus orchards revealed allopatric spatial patterns between some of the dominant species. Redundancy analysis showed that soil properties related to soil water potential explained significant variability in the EPN community structure. We studied behavioral and physiological responses to soil water potential of two closely related, allopatric Steinernema glaseri-group species. Steinernema diaprepesi is a dominant EPN in deep, well drained soils of the elevated central ridge region, whereas Steinernema sp. has been detected only off the central ridge in orchards with shallow ground water depth. In laboratory studies we monitored the survival of both species in a variety of soils, each at water potentials ranging from saturated to permanent wilting point. The persistence of Steinernema sp. was greater in saturated soil than in well-drained soil, whereas S. diaprepesi persisted better in well-drained compared to saturated soil. Thus, the behaviours of these species in controlled assays were congruent with those that could result in the observed spatial patterns in Florida. Comparative proteomic analysis revealed differences in protein expression by each species in well-drained and saturated soil. Detected proteins are being identified by LC-MS-MS to understand mechanisms of habitat adaptation. Our data suggest the plausibility of engineering soil moisture conditions in ways that favour more desirable (effective) EPN species. Conversely, these particular behavioural traits are likely to be useful in guiding the selection of EPN species for use in different ecoregions. STUDIES ON THE CEREAL NEMATODES, HETERODERA SPP. AND PRATYLENCHUS SPP. IN TURKEY. Elekcioglu1, I.H., M. Imren2, H. Toktay3, E. Yavuzosmanoglu-Sahin4, E.B. Kasapoglu1 and A. Dababad5. 1Cukurova University, _ Faculty of Agriculture, Department of Plant Protection, Adana, Turkey; 2Abant Izzet Baysal University, Faculty of Agriculture, Department of Plant Protection Bolu, Turkey; 3University of Nigde, Faculty of Agricultural Sciences and Technologies, Nigde, Turkey; 4Karamanoglu Mehmetbey University, Vocational School, Department of Plant and Animal Production, Karaman, Turkey; 5CIMMYT (International Maize and Wheat Improvement Centre), Wheat Improvement Program, Ankara, Turkey. Cereal cyst nematodes and root lesion nematodes are economically important in wheat production systems worldwide. Heterodera filipjevi is widely distributed in the Central Anatolia and East Anatolia whereas H. avenae and H. latipons were found in the eastern Mediterranean and south-eastern Anatolia region of Turkey. Pratylenchus thornei and P. neglectus are the predominant species of root-lesion nematodes in central, east and south-eastern Anatolia. Cereal cyst nematodes and root lesion nematodes cause yield losses between 5-50%. The effect of temperatures on the incubation duration of cysts of H. avenae was investigated under in vitro conditions. A number of variables were assessed to optimize P. thornei screening including initial nematode density, inoculation volume, soil type, tube size, reference cultivars, harvest time and watering regime. Growth room experiments showed clear differentiation between the resistance and susceptible cultivars after 9 weeks in sandy growth medium (70:29:1 sand, field soil and organic matter) with small tubes (15x100 mm) inoculated with 400 cultured P. thornei/plant. All H. avenae populations collected from south-eastern Anatolia were found to belong to Ha1 group Ha21 pathotype. The efficiency of some sources of resistance (CreR, Cre1, Cre2, Cre3, Cre7 and Cre8) in wheat against several H. avenae, H. filipjevi and H. latipons population was investigated in-vitro to resemble climate conditions in Turkey. Cre1, Cre3 and Cre7 genes induced resistance against both H. avenae and H. latipons populations. Plants with CreR and Cre8 genes revealed resistance against H. filipjevi. However, no gene was found to show resistance against all three nematode species together. STUDIES ON THE CITRUS NEMATODE, TYLENCHULUS SEMIPENETRANS, COBB IN THE EAST MEDITERRANEAN REGION OF TURKEY. Elekciog˘lu1, I.H., E.B. Kasapog˘lu1, H. Toktay2 and M. Imren3. 1University of C xukurova, Faculty of Agriculture, Department of Plant Protection, 01360, Adana Turkey; 2University of Nigde, Faculty of Agricultural _ Sciences and Technologies, Nigde, Turkey; 3Abant Izzet Baysal University, Faculty of Agriculture, Department of Plant Protection, Bolu, Turkey. Turkey is one of the most important citrus-producing countries in the Mediterranean region. The citrus nematode (Tylenchulus semipenetrans) was found an important parasite of citrus in this region and 90 % of citrus orchards were infested with this parasite. Population densities were mostly above economic threshold. Mediterranean biotype was found

160 Journal of Nematology, Volume 46, No. 2, June 2014 mainly in this region. Population dynamics of citrus nematode were studied in different citrus orchards in Adana condition, population reached the minimum level on December - January and the maximum level on July - August. Nematicide treatment resulted increase in orange yield (0.9 %), and the following year it reached 9.2 % although it was not statistically significant. Sour orange (Citrus aurantium L.) known susceptible to this pest is commonly used as rootstocks in this region. To determine the resistance of citrus rootstocks to T. semipenetrans different rootstocks were investigated in the collection orchard of University of Cukurova, Faculty of Agriculture and roots and soil from 34 sour orange clones (Tuzcu clones), C. obovoidea, C. ampullaceae, C. sulcata, C. taiwanica, C. volkameriana; Poncirus trifoliata, Troyer citrange, Carrizo citrange, Kleopatra mandarin and Yuzu rootstocks were examined. Citrus nematodes were found in different densities in all samples taken from roots and rhizosphere of the different rootstocks and were also found in the roots of Poncirus trifoliata, Carrizo sitrange and Troyer citrange which are known as resistant or tolerant, from previous studies. In order to reach more definitive conclusions on this subject, all of these rootstocks should be screened under controlled conditions. NEMATODES ASSOCIATED WITH GREENHOUSE BANANA PRODUCTION IN TURKEY. Elekciog˘lu, I.H., G. Yoraz and E.B. Kasapog˘lu. University of C xukurova, Faculty of Agriculture, Department of Plant Protection, 01360, Sarıcxam, Adana Turkey. Commercial banana production is well established in the subtropical regions of Turkey. The adoption of banana production under protection has increased production and productivity. In Anamur and Bozyazı, protected cultivation is common, with open cultivation widely used in Alanya and Gazipasa. In 2008, the production areas increased to 40 000 da and production increased to 200 000 tons, with accompanying high quality fruit. Root-knot nematodes and Helicotylenchus spp. cause economic losses in banana production. These nematodes cause damages by feeding directly on roots and also by inducing soil-borne diseases. Population dynamics of nematodes were examined in Bozyazı district of Mersin province in a greenhouse between 2011 and 2013. During this period, soil samples were collected each month, and the nematodes extracted. Plant parasitic nematodes were identified to species level and population densities determined. Helicotylenchus multicinctus, Helicotylenchus dihystera, Meloidogyne incognita and Meloidogyne javanica were identified, with H. multicinctus the most common. In general, the population density of H. multicinctus declined in December and January and increased in March. Helicotylenchus dihystera was not common in the study area with root-knot nematodes also not observed in high numbers. VBC-90017: A NEW BIORATIONAL NEMATICIDE FOR THE CONTROL OF PLANT-PARASITIC NEMATODES. Eldridge, R. Valent BioSciences Corporation, 870 Technology Way, Libertyville, IL 60048, USA. VBC-90017 is a new, novel, biorational nematicide under development by Valent Bio-Sciences Corporation. VBC-90017 has a number of user friendly features including a liquid EC formulation, application flexibility, and an excellent toxicity profile. Initial field efficacy studies show nematode gall reduction, population suppression and subsequent positive plant responses in potato, tomato, cucumber and tobacco. Field development trials conducted on a range of crops, and the unique features and benefits of this new material are discussed. ROOT-LESION NEMATODES, PRATYLENCHUS SPP., PARASITIZING POTATO IN PORTUGAL. Esteves1, I., C. Maleita2 and I. Abrantes1. 1IMAR-CMA, Department of Life Sciences, FCTUC, University of Coimbra, 3004-517 Coimbra, Portugal; 2CIEPQPF, Chemical Engineering Department, FCTUC, University of Coimbra, Rua Sı´lvio Lima, Po´lo II – Pinhal de Marrocos, 3030-790 Coimbra, Portugal. Root-lesion nematodes Pratylenchus spp., are widespread and economically important parasites of several cultivated plants, including potato. In Portugal, potato cyst nematodes, Globodera spp., are a frequent problem in main production areas and root-knot nematodes, Meloidogyne spp., have been found coexisting with potato cyst nematodes. Although the diversity and pathogenicity of potato cyst nematodes are well documented, little work has been done to assess the importance of Pratylenchus spp. to potato. This work aimed to assess the diversity of root-lesion and other plantparasitic nematodes in potato crops. Sampling encompassed fields in northern, central and southern regions of Continental Portugal. Nematodes were extracted from 42 soil and root samples, using standard extraction techniques, identified to genus level and population densities quantified. The identification of Pratylenchus spp. was based on the diagnostic morphological characters, species specific primers and D2D3 28S rDNA sequencing. Root-lesion nematodes were detected in all regions, except in the south, where nematodes were only found in the soil. Pratylenchus spp. were identified in 83% and 79% of soil and root samples, respectively. Meloidogyne spp. and potato cyst nematodes were also found in roots infected with root-lesion nematodes. Pratylenchus penetrans was the most abundant species followed by Pratylenchus neglectus, Pratylenchus crenatus and Pratylenchus thornei. The information obtained on the ubiquity and diversity of root-lesion nematodes reinforced the need for a careful monitoring of these nematodes in potato crops. Further studies are being conducted to assess the pathogenicity of the Pratylenchus spp. on common commercial potato cultivars.

Abstracts 161 CHARACTERISATION OF NOVEL SECRETED PROTEINS AND THEIR POTENTIAL ROLE IN CYST NEMATODE PARASITISM. Eves-van den Akker1,2, S., C.J. Lilley2, J.T. Jones1 and P.E. Urwin2. 1The James Hutton Institute, Invergowrie, DD2 5DA, Scotland; 2University of Leeds, Leeds, LS2 9JT, England. Plant parasitic nematodes comprise several groups; the most economically damaging of these are the sedentary endoparasites, causing losses to agriculture worth in excess of £75 billion per year. Sedentary endoparasitic nematodes are obligate biotrophs and modify host root tissue, using a suite of effector proteins, to create a feeding site that is their sole source of nutrition. Genes potentially involved in the feeding process of the potato cyst nematode, Globodera pallida, have been identified through analysis of life-stage specific transcriptome sequences mapped to its recently assembled draft genome. Differential expression analysis coupled with signal peptide and transmembrane prediction enabled identification of a range of novel, putatively secreted proteins. A role in cyst nematode feeding has been implicated for one gene family whose members were found to be highly diverse between individuals of the same population. In situ hybridisation was used to demonstrate expression of this gene family in the amphid sheath cells of parasitic stages. These cells service the amphids, structures that may be the origin of the feeding plug. Secretion of the encoded proteins has been confirmed using immunochemistry to localise them in host potato roots at the plant-nematode interface. Despite the variation in gene sequence between individual nematodes, in planta expression of double stranded RNA targeting a conserved region of the gene family results in approximately 80 % reduction in total nematode infection. Representatives of this gene family have been identified in other cyst nematode species but not in root-knot nematodes. MELOIDOGYNE INCOGNITA INFECTED CORCHORUS OLITORIUS: DRAMATIC EFFECT OF DIFFERENT EXTRACTS OF EUCALYPTUS OFFICINALIS. Fabiyi, O. Department of Crop Protection, Faculty of Agriculture University of Ilorin, Ilorin, Nigeria. This study investigated the nematicidal properties of various fractions from Eucalyptus officinalis obtained by different methods of extraction. The fractions were partially characterized using Fourier transform infra-red (FT-IR) and gas chromatography mass spectroscopy (GCMS) analysis. The constituent of E. officinalis as revealed by the GCMS result include eucalyptol (19.3%), a-pinene (3.8%), citronellol (4.2%), a-terpeneol (39.2%) and 1, 8-cineole (33.5%). The FT-IR spectral data obtained also supported the finding that 1,8 cineole and a terpeneol were the main constituents of the E. officinalis fractions. The fractions obtained from the methanol extract chromatographed on silica gel 100-120 mesh grade produced significantly (p1). Ac xaı´ (Euterpe oleraceae), atemoya (Annona cherimola), avocado (Persea americana), cashew nut (Anacardium occidentale), citrus (Citrus spp.), coconut (Cocos nucifera), grape rootstocks, jabuticaba (Plinia trunciflora), mango (Mangifera indica), mulberry fruit (Morus alba), passion fruit (Pasiflora spp.), sapodilla (Manilkara zapota), soursop (Annona muricata), starfruit (Averrhoa carambola), olive (Olea europaea) and strawberry (Fragaria x ananass) were considered as non-hosts or poor hosts to the nematode. These species may be planted in crop rotation in areas infested by M. enterolobii. HYPERSPECTRAL SENSOR TECHNIQUES AND POPULATION MODELLING OF HETERODERA SCHACHTII FOR ASSESSING THE SPATIO-TEMPORAL DYNAMICS OF NEMATODE INFESTATION IN SUGAR BEET VARIETIES UNDER FIELD CONDITIONS. Fricke1, B., K. Schmidt2, M. Daub3, H. Goldbach1. 1 INRES-Plant Nutrition, Faculty of Agriculture, Bonn University, Germany; 2 Nemaplot, Bonn, Germany; 3 Julius Ku¨hn Institute, Plant Protection in Field Crops and Grassland, Elsdorf, Germany. Crop losses in sugar beet due to Heterodera schachtii infestation are of major concern in many areas. Sustainable crop management provides resistant and tolerant varieties against pathogen infestation. The present study aims at monitoring the infestation process and to characterize specific phenotypic traits of different sugar beet cultivars related to nematode damage under field conditions. The application of hyperspectral sensor techniques allows non-invasive monitoring of the leaf canopy. Assessing hyperspectral signatures from canopy reflection is used to describe the stress response of the plants. A high spectral resolution, however, creates massive amounts of data. Thus, the classification of vegetation vitality by different spectral vegetation indices from ratio of selected wavebands is used to reduce the amount of sensor information. Choosing the most appropriate wavebands to correlate with plant physiological parameters may then facilitate data collection and handling. It bears, however, the risk of losing important information. In a novel approach, however, the entire spectral range of canopy reflection is considered. The classification by the NemaplotÒ model is based on two steps. First, the model is fitted to the hyperspectral signature by transforming the wavelength information to specific numerical parameters. Second, these numeric parameters are interpreted with a discriminant analyses in order to classify the signature which correlates best with nematode infestation. The stress answer of sugar beets over the course of the day and across a season was sampled and data are presented where hyperspectral information is used to describe Heterodera population dynamics. NEMATODES ASSOCIATED WITH EDAMAME, GLYCINE MAX L. (MERR.) IN ARKANSAS, UNITED STATES. Fultz, J., and T. Kirkpatrick. University of Arkansas, Fayetteville, Arakansas 72701, USA. Edamame, also known as vegetable soybean, Glycine max, was introduced to the United States from Japan in 1890 and has been growing in popularity as a high-fibre, low-sugar snack in recent years. In 2012, American Vegetable Soybean and Edamame, Inc. established the first commercial processing plant in the United States near Mulberry, Arkansas. About 2,000 acres of edamame are now being grown annually in the state. Since edamame is harvested as an immature seed, management practices vary from those used for conventional soybean. Plant-parasitic nematodes, particularly Meloidogyne incognita (root-knot) and Heterodera glycines (soybean cyst) known to be wide-spread in Arkansas, are pests of concern. The objective of this study was to determine the nematodes associated with edamame in Arkansas fields, and to evaluate new breeding lines for resistance to these nematodes. Production fields of 2013 were surveyed immediately following harvest to determine the presence, identity, and relative population density of nematodes. Where they were detected, Meloidogyne spp. second stage

166 Journal of Nematology, Volume 46, No. 2, June 2014 juveniles were collected and used to establish greenhouse populations on tomato and sequenced for further identification to species. Additionally, edamame lines developed by the Arkansas soybean breeding program were evaluated for host suitability to M. incognita and H. glycines in greenhouse trials. All lines were good hosts for H. glycines race 3, but M. incognita, race 3, reproduction was significantly lower in one of the lines. This work will aid producers and crop advisors in developing nematode management strategies for edamame production and provide insight on the potential risk of root-knot and soybean cyst nematodes to this emerging crop. DEVELOPMENT AND ASSESSMENT OF MICROSATELLITE MARKERS FOR THE SPECIFIC IDENTIFICATION OF THE BEET CYST NEMATODE, HETERODERA SCHACHTII. Gamel1, S., A. Letort1 and E. Grenier2. 1French Agency for Food, Environmental and Occupational Health and Safety (Anses), Plant Health Laboratory, Nematology Unit, Domaine de la Motte au Vicomte, BP35327, 35653 Le Rheu, France; 2INRA, UMR 1349 IGEPP, Domaine de la Motte au Vicomte, BP35327, 35653 Le Rheu, France. Heterodera schachtii is known as the sugarbeet cyst nematode due to the major agronomic damage observed on sugarbeet culture. Other plant families can also be affected by this polyphagous nematode such as Brassicaceae (i.e. oilseed rape, cabbage) and Solanaceae (i.e. potato, tomato). Despite H. schachtii’s global presence, the species is regulated in many nonEuropean countries. Consequently, imported crops have to be free from H. schachtii. In this context, the development of a reliable identification method sugarbeet cyst nematodes appears necessary both for improving agronomic strategies by preventing sugarbeet cultivation in contaminated fields and also for promoting agricultural trade of other crops. As H. schachtii belongs to the Schachtii group, the analytical method for a reliable identification of H. schachtii must be very specific. After sequencing numerous nematode populations and performing bioinformatic analysis, the study focused on the selection of specific and sensitive microsatellite markers for the identification the sugarbeet cyst nematode using conventional PCR. The findings showed that one primer set is specific enough when tested among 18 non target populations of Globodera and Heterodera genera of which 10 were from the Schachtii group (Heterodera betae, Heterodera. glycines, Heterodera trifolii, Heterodera daverty and Heterodera ciceri). The sensitivity of the test allowed the identification of a single juvenile of H. schachtii, making this method suitable for preplant analysis and import/export control. PHASE-OUT OF METHYL BROMIDE AND NEMATODE MANAGEMENT: ACHIEVEMENTS AND FUTURE CHALLENGES. Gamliel, A. Institute of Agricultural Engineering, ARO, the Volcani Center, Bet Dagan 50250, Israel. The phase-out of methyl bromide in Israel resulted in gradual reduction in methyl bromide use, until its total elimination in 2012. During the phaseout process, all the available and the registered soil fumigants were evaluated for the control of various soilborne pests and primarily nematodes. Additionally the combinations of fumigants were evaluated in order to enhance the performance of each fumigant alone. During these years few effective combinations, e.g. metham sodium and 1,3dichloropropenfor the control of soilborne fungi and nematode were registered and are commercially applied. The quest for new fumigants yielded only dimethyl disulfide, which has been registered and is still in commercial use. The commercial application of soil solarization underwent a considerable boost following the implementation of the methyl bromide phaseout. Solarization combined with nematicide fumigants provided effective control of root knot, in the production of high cash crops, e.g. strawberries and peppers. Bottlenecks were identified during the phase out of methyl bromide; these are focused in four major areas: application problems, low efficacy with specific soilborne pests, the cost factor, and the human factor. From a technology-diffusion perspective, certain alternatives such as application of new nematicides, and their combination with solarization, although comparable to integrated pest management, are complex, often more expensive than the equivalent chemical control methods. The phaseout of methyl bromide resulted also in gradual development of nematodes, pathogens and weed populations in fields which has been treated repeatedly with methyl bromide. Root knot nematodes have spread over vast area during the last years and require additional chemical and nonchemical treatments in order to provide a platform for a healthy and productive crop. More pests are yet to emerge, and the challenges in their effective control become a more challenging task. PASTEURIA NISHIZAWAE LIBRARY AND SCREENING THE BEST STRAIN FOR PRODUCT DEVELOPMENT. Gao, X., M. Doroh, S. Griswold, T. Hewlett, C. Marshall, A. Perret-Gentil and A. Santos. Syngenta Crop Protection, 12085 Research Dr., Suite 185, Alachua, Florida 32615, USA. We are developing a Pasteuria library that plays a vital role in the new product development and improvement of product performance against plant parasitic nematodes at the Pasteuria Bioscience site of Syngenta. Presently 106 strains of Pasteuria from 13 different genera of nematodes are available. Pasteuria nishizawae strains from the soybean cyst nematode Heterodera glycines, were used as a model for developing screening techniques. Nematode samples were collected in crop growing seasons. Pasteuria spores on the nematode’s cuticle or inside nematode bodies were examined under inverted microscopes (6403). Pasteuria spp. were isolated and cultured with the standard operating procedure. In-vitro spores were produced through fermentation. Strains were selected based on culture assays, 24-well infection bioassays and other screening experiments. Thirty four strains of P. nishizawae were tested against 10 selected soybean cyst

Abstracts 167 nematode-populations in the laboratory bioassays. Pn1, Pn3, Pn5, PnS38 and PnS46 were found to have high infection capacity. These isolates were tested in growth chamber experiments conducted in small containers. Each pot was filled to 2 cm from the top of the pot with sand. Three soybean seeds were sown per container, and one thousand H. glycines juveniles were inoculated per pot. Infected roots were stained with Red Food Colour for visualization of nematodes two weeks after inoculation. Results from cultural assays, laboratory bioassays and greenhouse tests indicated that Pn1, PnS79 and PnS84 were promising strains for new products. The screening systems established for testing P. nishizawae against H. glycines have been used for other Pasteuria spp. on other nematodes. STEINERNEMA - XENORHABDUS SYMBIOSIS AND YERSINIA PSEUDOTUBERCULOSIS: BROTHERS IN ARMS OR HACKING SYSTEM? Gengler1,2, S., A. Laudisoit3 and P. Wattiau1. 1Veterinary & Agrochemical Research Centre, Brussels, Belgium; 2Institute of Life Sciences, Universite´ Catholique de Louvain-la-Neuve (UCL), Belgium; 3School of Biological Sciences, University of Liverpool, United Kingdom. The capacity of soil invertebrates to act as intermediary hosts was the starting question of our study and entomopathogenic nematodes (EPN) were investigated in this respect. We wondered whether notorious mammalian pathogens taxonomically related to Xenorhabdus, were able to ‘‘hack’’ the symbiotic relationship associating Xenorhabdus and Steinernema EPN. Our previous results showed that Yersina pseudotuberculosis, that causes gastroenteritis in mammals, can survive during several generations of the EPN life cycle in vitro. Genetic determinants that allowed Y. pseudotuberculosis to survive inside EPN are currently under study. The potential implication of the recently discovered type 6 secretion system components as well as yplA phospholipase involved in survival within the insect host is systematically investigated. Deletion mutants obtained so far show promising results since an yplA mutant is unable to survive inside EPN. At the same time, we are investigating whether Y. pseudotuberculosis really plays a symbiotic role in EPN or if it just hacks the system. To deal with this question we obtained monoxenic IJs hosting Y. pseudotuberculosis. Our first results show that the capacity of Y. pseudotuberculosis to survive infection cycles inside EPN lacking Xenorhabdus is substantially reduced. Moreover, multiplication of EPN inside the insect cadaver seems to be affected and emergence of IJs from the cadaver is delayed. This strongly suggests that Y. pseudotuberculosis cannot maintain in EPN in the absence of Xenorhabdus. These findings, if they turn out to have an environmental significance, may reveal an unexpected biotic reservoir explaining the long-term persistence and dissemination of pathogenic bacteria in the environment. INTRASPECIES DIVERSITY IN THE ROOT KNOT NEMATODE MELOIDOGYNE ETHIOPICA. Gericˇ Stare, B., P. Strajnar, S. Sˇirca and G. Urek. Agricultural Institute of Slovenia, Hacquetova ulica 17, SI-1000 Ljubljana, Slovenia. The root-knot nematode Meloidogyne ethiopica are considered one of the most important emerging global plant pests. Meloidogyne ethiopica is considered a tropical, polyphagous pest that is able to parasitize at least 80 different host plants. However, this pest can survive winters in moderate temperate region with temperatures below freezing point. This species was reported from Africa (Tanzania, Kenya, Ethiopia, Mozambique, Zimbabwe, South Africa), South America (Brazil, Chile) and Europe (Slovenia, Greece, Turkey, Italy). We have compared populations of M. ethiopica from South Africa, Brazil and Europe (Slovenia, Greece, Turkey) and evaluated variability of biochemical markers used for identification (EST, MDH isozyme profiles) and several molecular markers (ITS, 18S and 28S rDNA and mtDNA) used in phylogenetic studies of genus Meloidogyne. The studied populations, determined as M. ethiopica based on morphological and biochemical characters, have displayed three different patterns of isozyme esterase. While some of Meloidogyne species have one speciesspecific esterase pattern, some species like M. arenaria, M. incognita and M. exigua are known to display several esterase patterns, and some patterns can be found in more than one species (e.g. A1). Sequences from our tested populations were combined with other Meloidogyne spp. sequences from public databases to assess phylogenetic relationship between different populations of M. ethiopica and closely related species. Great care should be taken when molecular marker is selected to study phylogenetic relationships or to confirm identification of Meloidogyne species as variability both at rDNA and mtDNA was observed in this and previous studies. ANALYSIS OF ENDOPHYTIC FUNGI AND PLANT-PARASITIC NEMATODES FROM IRRIGATED AND UPLAND RICE ECOSYSTEMS IN KENYA. Gheysen1, G., T. Kyndt1, F. Soraya de Carvalho2, M. Ho¨fte2, W. Bert3, T. Janssen3, R.K. Mibey4 and P.N. Njira1,4. 1Department of Molecular Biotechnology, Ghent University, Coupure Links 653 B-9000, Ghent, Belgium; 2Department of Crop Protection, Ghent University, Coupure Links 653 B-9000, Ghent, Belgium; 3Department of Biology, Ghent University, K.L. Ledeganckstraat 35, B-9000, Ghent, Belgium; 4Moi University, P.O Box 390030100, Eldoret, Kenya. Endophytic fungi are ubiquitously distributed in almost all plants species. They have been found to colonise single or multiple plant organs, often protecting their host plants from biotic as well as abiotic stresses. In the present study, the diversity of endophytic fungi and plant-parasitic nematodes from irrigated and upland rice ecosystems was analysed using DNA sequences and morphological characteristics. A total of 45 and 25 fungal species were identified from irrigated and upland ecosystems, respectively. In the irrigated ecosystem, a relatively high number of different fungi were isolated.

168 Journal of Nematology, Volume 46, No. 2, June 2014 Epicoccum nigrum was the most dominant species in this ecosystem. Species richness was lower in the upland ecosystem, where Fusarium oxysporum isolates were extremely abundant. Nematodes were found only in the upland ecosystem, and were identified as Pratylenchus zeae and Pratylenchus goodeyi. Previous records showed P. zeae to be a rice pathogen, but P. goodeyi is described here for the first time in rice, a remarkable observation especially in coastal low altitude areas characterised by high temperatures. Preliminary results from greenhouse experiments indicate that P. goodeyi can indeed infect and multiply in rice. Based on available information from other crops studied, Epicoccum nigrum and Talaromyces flavus were selected and are currently tested for their biological activities against these lesion-nematodes. DIVERSITY OF ENTOMOPHILIC NEMATODES AND THE THREAT OF CRYPTOGENIC INVASIVE SPECIES. Giblin-Davis1, R.M., N. Kanzaki1, 2 and K.A. Davies3. 1Fort Lauderdale Research and Education Center, University of Florida-IFAS, 3205 College Ave., Davie, Florida 33314, USA; 2Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687 Japan; 3Australian Centre for Evolutionary Biology and Biodiversity, School of Agriculture, Food and Wine, University of Adelaide, Waite Campus, PMB 1, Glen Osmond, South Australia 5064, Australia. Nematode fauna are often highly specialized for microniche utilization, i.e., many nematode species are highly habitat specific. For example, a transect survey of soil, litter, epiphyte and insect-associated nematode fauna in the same site yields different nematode community strata, and the communities do not necessarily overlap each other. Thus, the ‘‘insect-associated nematode community’’ can potentially be considered as a specific ‘‘habitat’’ of specialized and divergent entomophilic nematodes. Based on entomophilic nematode surveys conducted by the authors, correlation between the nematode assemblage pattern (e.g., feeding habitat groups) and the life history of their host/carrier insect (e.g., feeding resource, habitat preference and life cycle) was expected, and the number of entomophilic nematode species was estimated at somewhere between 40,000 and 500,000 species. Considering that the number of currently described species for the entire phylum is about 27,000, most of the entomophilic nematodes are hypothesized to be undescribed species. Biodiversity can be affected by many different kinds of environmental alternations. Here, we focused on the potential for cryptogenic invasive species being introduced by international trade as a threat to the diversity of entomophilic nematodes by introducing several contemporary examples. GENETIC IMPROVEMENT AND BREEDING OF EPN: THE RACE FOR THE ‘‘SUPER NEMATODE’’. Glazer, I. Department of Nematology, Agricultural Research Organization, the Volcani Center, Bet Dagan 50250, Israel. Entomopathogenic nematodes (EPN) which are used as commercial biocontrol agents were targeted for genetic improvement in recent years. Most studies were aimed at improvement of traits which were believed to contribute to the efficacy of these organisms under field condition. That included, enhancement of heat and desiccation tolerance, host finding and resistance to nematicides. In these studies substantial enhancement of the selected trait was demonstrated. The contribution of these traits to nematode efficacy under field condition was not yet demonstrated. In many cases the enhanced trait was not stable and rapid degradation was observed following relaxation of the selection pressure. Furthermore, reduction of fitness can occur during multiple reproduction cycles in large scale liquid culture of the nematodes. Modern genetic and molecular tools such as RNA-seq, transcriptomics can be utilised to follow and understand the molecular basis of a beneficial trait. Molecular markers and QTL can also help to develop new strategies for genetic improvement of EPNs. New techniques for genome editing (TALENs and CRISPR-Cas9) may be able to modify important traits. In the presentation, approaches to address the challenges concerning development of the ‘‘super nematode’’ will be discussed. CHARACTERIZATION OF MELOIDOGYNE SPECIES FROM IRRIGATED RICE IN SOUTHERN BRAZIL. Gomes1, C.B., R.R.D. Negretti2, V.S. Mattos3, L. Somavilla2, R. Manica-Berto2, D. Agostinetto2 and R.M.D.G. Carneiro3. 1 Embrapa Clima Temperado, C.P. 403, 96001-970 Pelotas RS, Brazil; 2PPGFS/Fac. Agronomia, Universidade Federal de Pelotas, C.P. 354, Pelotas RS, Brazil; 3Embrapa Recursos Gene´ticos e Biotecnologia, C.P. 02372, 70849-979, Brası´lia DF, Brazil. Southern Brazil has the largest cultivated area of irrigated rice in the country. Nonetheless, rice production may be limited by several plant pathogens, including root-knot nematodes (Meloidogyne spp.). Considering the importance of root-knot nematodes in Brazil, a survey was carried out in irrigated rice fields in Rio Grande do Sul (RS) and Santa Catarina (SC) states. Meloidogyne populations were characterized biochemically by esterase (Est) and malate dehydrogenase (Mdh) phenotypes and morphologically by the female perineal patterns. Fifty five Meloidogyne spp. populations were detected in 48% of rice samples, and a total of five esterase phenotypes were identified. In Rio Grande do Sul state Meloidogyne graminicola (Est VS1, Rm: 0.70), Meloidogyne sp.2 (Est R2, Rm: 0.81, 0.91) and Meloidogyne sp.3 (Est VS1-2, Rm: 0.61, 0.70) were identified, which corresponded to 80, 40 and 10% of samples, respectively. In Santa Catarina state M. graminicola, Meloidogyne javanica (Est J3, Rm: 1.00, 1.20 and 1.35), Meloidogyne sp.1 (Est R1, Rm: 1.02), Meloidogyne sp.2 and Meloidogyne sp.3, accounted for 93.7, 12.5, 62.5, 12.5 and 6.2% of samples, respectively. Meloidogyne javanica showed a N1 Mdh phenotype (Rm: 1.0), while other populations exhibited a N1a (Rm: 1.4) phenotype. Morphology of female perineal patterns allowed identification only for M. javanica and M. graminicola. To further characterize these tree atypical

Abstracts 169 populations, sequencing and phylogenetic analyses of internal transcribed spacer-rRNA (ITS) and D2-D3 segment of 28S rRNA have been done. Future phylogenetic studies involving these atypical isolates will be performed. USE OF NUTRITIONAL SUPPLEMENTS FOR THE MANAGEMENT OF ROOT-KNOT NEMATODE (MELOIDOGYNE INCOGNITA) INFECTING POTATO. Gondal1, A.S., N. Javed1, S.A. Khan1 and M. Shahid2. 1Department of Plant Pathology, University of Agriculture Faisalabad, Pakistan; 2Plant Pathology Research Institute, Faisalabad, Pakistan. A wide range of synthetic chemicals has been discriminately used as the major control measure against plant pathogens. Environmental pollution, degradation, insecticide resistance development and other agronomic concerns have prompted scientists to seek alternative disease management strategies. Present in-vitro studies were conducted to evaluate the efficacy of nutritional supplements including micro-power, humic acid and plant protectors containing benzoic acid against root-knot nematode (Meloidogyne incognita) infection on susceptible potato cultivar. Each treatment applied as single or in combined form significantly reduced the number of galls and egg masses and promoted overall plant growth as compared to unamended control. Application of aqueous solution of 4% plant protector + 4% micro-power + 2% humic acid enhanced the number of leaves, root and shoot developmentand tuber weight and decreased the root weight, with the minimum number of females, root galls and egg masses recorded. Nematodes fecundity was observed to be the highest for the control treatment resulting in poor plant growth and development of higher number of galls and egg masses. The significantly lower number of galls and egg masses and enhanced plant growth observed with the combined application of plant protector 4%, micro power 4% and humic acid 2% indicated this treatment to be superior. EFFECTS OF LONG-TERM CORN-SOYBEAN CROP SEQUENCES ON THE NEMATODE COMMUNITY. Grabau, Z.J. and S.Y. Chen. University of Minnesota Department of Plant Pathology, Southern Research and Outreach Center, 35838 120th Street, Waseca, Minnesota 56093, USA. The effects of long-term corn-soybean crop sequences on the nematode community were studied at a field site that was established in 1982 in Minnesota, USA. The crop sequences were: (i) five-year rotation between crops such that both crops are in years 1, 2, 3, 4, and 5 of monoculture every year; (ii) annual rotation with both crops planted each year; (iii) continuous monoculture of each crop; (iv) annual rotation between two cultivars, but crop monoculture of each crop. Since 1995, sequence IV was single-cultivar monoculture of each crop. Since 2010, soybean susceptible to soybean cyst nematode (Heterodera glycines) and Bt corn were used for crop sequences i, ii, and iii while monoculture of a H. glycines-resistant soybean cultivar or a non-Bt corn cultivar was used for sequence iv. Beginning in 2010, half of each plot was treated with granular nematicide. In 2013, the nematode community in each plot was assessed at planting, midseason, and harvest. Nematicide was effective against plant-parasitic nematodes, but also reduced fungivore population and shifted the nematode community to a lower ecological succession. Crop sequences strongly affected plant-parasitic nematodes, but also affected fungivores, bacterivores, nematode community diversity, maturity, and enrichment. Differences in the nematode community were most dramatic when comparing crops rather than length of monoculture within a crop. After dramatic change in the initial years of growing a crop, changes in the nematode community continued incrementally as years in monoculture increased. This study shows the agronomic and ecological impact of crop rotation. DIRECT AND INDIRECT CONSEQUENCES OF GLOBODERA PALLIDA SELECTION BY RESISTANT POTATO GENOTYPES. Grenier, E., S. Fournet, M.C. Kerlan, D. Eoche-Bosy and J. Montarry. INRA, UMR IGEPP, Domaine de la Motte au Vicomte, BP35327, 35653 Le Rheu, France. In this study, we have first explored the variability of resistance durability in different potato genotypes harboring the same resistance QTL but differing by their genetic background. The indirect consequences of the resistance adaptation in terms of local (i.e. genotype-specific) adaptation and cross-virulence were then also investigated. Following the virulence of the potato cyst nematode Globodera pallida in a long-term experimental evolution protocol, our results showed that nematode populations were able to adapt to the resistance of four potato genotypes carrying the QTL GpaV from Solanum vernei, and that the plant genetic background has an impact upon the durability of resistance. The pattern of local adaptation observed here revealed a trade-off between the adaptation to a resistant potato genotype and the adaptation to another resistant genotype differing in its genetic background. In terms of cross-virulence between potato genotypes derived from different resistance sources (Solanum sparsipilum and Solanum spegazzinii), we showed that the adaptation to resistance QTL GpaVvrn does not necessarily allow the adaptation to collinear GpaV loci. The results presented here will be useful for identifying durable strategies for resistance deployment. The virulent populations obtained from these experimental evolutions are actually used in order to identify, through a genome-scan approach, the genomic regions involved in that resistance breakdown. DEVELOPMENTS IN THE IMPLEMENTATION OF ENTOMOPATHOGENIC NEMATODES IN INTEGRATED PEST MANAGEMENT SYSTEMS IN NORTH AMERICA. Grewal, P.S. Department of Entomology and Plant Pathology, University of Tennessee, Knoxville, TN 37996, USA. Entomopathogenic nematodes (EPNs) have emerged as excellent alternatives to chemical pesticides. They have been evaluated against nearly 200 insect pest species almost all of which can be controlled under field conditions. Broad host range

170 Journal of Nematology, Volume 46, No. 2, June 2014 and ability to seek and kill insects in soil and in cryptic habitats such as plant roots and tree trunks, where most chemical pesticides fail to reach make EPNs especially attractive. Ease of application via standard pesticide spray equipment and via diverse irrigation systems has facilitated the adoption of EPNs in diverse ecosystems. Compatibility with numerous agrochemicals including insecticides, miticides, fungicides, herbicides, wetting agents, plant growth regulators, and spray adjuvants enhances their use in IPM. Some pesticides even enhance EPN pathogenicity via synergism. Although, EPNs are most widely used against soil-dwelling stages, applications against above ground pests have also been successful. Foliar applications in glasshouses have shown particular promise. Applications to insect galleries in tree trunks provide excellent control of wood boring insects. Slow release formulations and partially-desiccated EPN-infected cadavers hold promise in prophylactic EPN use particularly for delivery via growing media commonly used in greenhouses and potted plant industries. Successes have also been reported in the use of EPNs in traps designed to lure and kill cockroaches, houseflies, grasshoppers, molecrickets, cutworms, and weevils. EPNs have also shown promise for the control of human and animal pests including fleas, yellowjackets, ants, termites, lice, and ticks. This presentation will provide an overview of the progress in implementing the nematodes in integrated pest management systems in North America. QUARANTINE NEMATODES DETECTED IN NINGBO PORT, CHINA. Gu, J. Technical Centre, Ningbo Entry-Exit Inspection and Quarantine Bureau, 9 Mayuan Road, Ningbo 315012, Zhejiang, China. Since 2012, many seedlings or trees mainly from Japan and Italy were imported through Ningbo port, China. After laboratory examination and identification with morphological and molecular methods, many species on the Quarantine List of China were repeatedly found. Including Meloidogyne mali, Meloidogyne camelliae, Pratylenchus japonicus, Pratylenchus hippeastri, Pratylenchus pseudopratensis, Trichodorus viruliferus, Trichodorus japonicus, Longidorus martin Xiphinema rivesi and some more species still not identified. So Rooted plants in soil or growth medium could be an important pathway for the spread of nematodes. Phytosanitary treatment is necessary but problematics. Since morphological identification is not easy, we suggest that D2D3 region of 28S rDNA gene could be a DNA barcoding gene. EFFICACY OF TWO ABAMECTIN FORMULATIONS ON BELONOLAIMUS LONGICAUDATUS IN GREENHOUSE AND FIELD TRIALS. Gu, M. and W.T. Crow. University of Florida, Gainesville, Florida 32611, USA. Belonolaimus longicaudatus (sting nematode) is considered as one of the major destructive plant parasitic nematodes on golf course turf in Florida. However, since the cancellation of fenamiphos, effective management options for this nematode on golf courses are few. Our research evaluated two formulations of abamectin (Avid 0.15 EC and A12115I, an experimental 1.7% a.i. formulation) for ability to reduce population density of B. longicaudatus and to improve turf health. In separate trials Avid 0.15 EC at rates of 80, 40, and 20 g abamectin/ha reduced population density of B. longicaudatus on turf in pots while A12115I at rates of 35 and 70 g abamectin/ha did not. In a field trial on B. longicaudatus infested turf, the two abamectin formulations applied every 14 days at the rate of 35 g a.i/ha with and without applications of the plant activator acibenzolar-S-methyl were compared to each other and to untreated plots. Both abamectin formulations improved turf percent green cover and the Avid 0.15 EC formulation was slightly better on some dates. Applications of acibenzolar-Smethyl enhanced percent green cover from both abamectin formulations compared with either abamectin formulation alone. These studies indicate that abamectin may be a useful nematode management tool for B. longicaudatus on turfgrasses. Additional greenhouse and field trials evaluating abamectin formulations and combinations with other compounds are currently underway. MELOIDOGYNE INFECTIONS UNDER INFRARED SPECTROSCOPY (FTIR-ATR). Guerra2, M., N. Cubilla´n3, A.M. Casassa4, E. Portillo1 and E. San-Blas1. 1 Instituto Venezolano de Investigaciones Cientı´ficas, Centro de Estudios Bota´nicos y Agroforestales, Laboratorio de Proteccio´n Vegetal, Av. 8 entre Calles 79 y 80, Maracaibo, Venezuela; 2Unidad de Tecnologı´a La´ser y Optoelectro´nica, Instituto Zuliano de Investigaciones Tecnolo´gicas, Km11. Carretera Vı´a La Can˜ada, Maracaibo, Venezuela; 3Laboratorio de Electro´nica Molecular, Facultad Experimental de Ciencias, Universidad del Zulia, Maracaibo, Venezuela; 4Instituto de Investigaciones Agrono´micas, Facultad de Agronomı´a, Universidad del Zulia, Maracaibo, Venezuela. Infrared spectroscopy is a technique developed to observe vibrational aspects of the functional groups from the molecules of a given sample. Basically, a sample is irradiated with different wavelengths and a detector capable to register the vibrational mode of the molecules, builds a spectrum. These spectra can be compared and differences nsee if one sample is different to another one. Infrared spectroscopy was used to compare infected or not infected tomato plants. Thirty tomato seeds were sown in pots (1 l) and after 30 days, 15 of them were inoculated with root-knot nematodes p. and the remainder were used as control plants. After 6 weeks, the plants were removed from the pots and washed clean of the soil. Root and leave samples were taken and let dry for 24 h at room temperature. The air dried samples were placed in a Fourier Transformed Infrared Spectrometer (FTIR) with an Attenuated Total Reflectance (ATR) accessory and the resulting spectra collected. The total data was pretreated to eliminate CO2 and water interference, smoothed and a second derivative procedure was done. The main spectral contributions were assigned to proteins around 3277, 1533 and 1231 cm21 and lipids around

Abstracts 171 2924, 2855, 1737 and 1452 cm21. Carbohydrates were also detected; the major absorption was found to be in the 1000–1200 cm21 range. The samples from infected plants showed differences in their composition, comparedto those coming from healthy plants. This technique demonstrated its usefulness for investigating many aspects of the nematode-plant interaction. MORPHOLOGICAL AND MOLECULAR CHARACTERIZATION OF THE GENUS LONGIDORUS (NEMATODA: DORYLAIMIDA) IN CHINA WITH DESCRIPTION OF A NEW SPECIES. Guo, K., P.T. Hoa, and J. Zheng. Nematology Lab, Institute of Biotechnology, College of Agriculture and Biotechnology, Zhejiang University, Hangzhou 310058, China. The genus Longidorus was established by Micoletzky, 1922 with Longidorus. elongatus de Man 1876 as the type species. This genus contains migratory ectoparasitic nematodes that attack a wide variety of crops or trees. Some of the species in the genus are vectors of plant viruses. Given its possible vector role, the genus Longidorus has gradually received increased taxonomic attention in China. The geographical distribution and host associations of Longidorus in mainland China were determined from a comprehensive survey. Based on soil samples collected from 12 provinces of China. Eleven Longidorus species, including one new species, were identified based on morphological characters. These are L. henanus, L. litchii, L. hangzhouensis, L. pisi, L. macromucronatus, L. fangi, L. camelliae, L. jonesi, L. fursti, L. nagnaensis, and Longidorus n.sp. The new species is characterized by females with medium size (L = 4.9-6.6 mm), lip region 18-21 mm wide, guiding ring situated at 70-90 mm from anterior end, long odontostyle (142-168 mm), tail short, bluntly rounded. Ratio of males: females one to one, spicules 100-123 mm long, 10-13 ventromedian supplements, and three juvenile stages were observed. A phylogeny tree was constructed based on the D2D3 region of 28S rRNA of the Longidorus species found in the study. POTENTIAL USE OF COMPOST FOR MANAGING NEMATODES, SOIL QUALITY AND CARROT YIELD. Habteweld1,2, A., D. Brainard2, M. Ngouajio2, S. Kravchenko3, and H. Melakeberhan1,2. 1Agricultural Nematology Laboratory Michigan, State University, East Lansing, MI 48824, USA; 2Department of Horticulture, Michigan State University, East Lansing, MI 48824, USA; 3Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI 48824, USA. In the absence of nematode resistant carrot cultivars, human- and environmental-health driven restrictions of broadspectrum nematicides, developing multi-purpose soil amendments that suppress plant-parasitic and other soil-borne pests and diseases is one of the priorities identified in the US Midwest vegetable growers. A hypothesis that plant- (PC) and animalbased (AC) compost may suppress plant-parasitic nmeatodes, increase beneficial nematodes, soil conditions, and yield and quality of fresh market (Sugarsnax) and processing (Cupar) carrot cultivars was tested in a sandy clay loam field. PC and AC, at 1, 1.5 and 2 times the standard recommendation for nitrogen source, and urea and non-amended (as controls) were tested. Preliminary analyses indicate significantly higher Maturity index (MI) in compost-treated (2.1) plots than in the controls (1.6), and fertility index (FI) in compost-treated plots was 1.0 compared to 1.3 in the controls, suggesting improved soil conditions. Abundance of omnivore-predatory nematodes was positively correlated (r = 0.53) with root to shoot dry matter ratio in the processing carrot variety plots, implying improved soil condition and more carrot root than shoot growth. Average total abundance was about 50 nematodes/100 cc of soil, with herbivores and fungivores being most abundant and omnivore and predators least abundant. Multifactor analysis of the relationship among nematode guilds, trophic groups and soil physiochemical properties shows significant correlation (r = 0.68) between c-p 1 bacteriovores and NO3-N. Soil food web structure appears to mature with time. Overall, the results support the hypothesis that compost treatment may increase beneficial nematodes and improving soil conditions. NEW CHEMISTRIES, MODES OF ACTION, AND DIFFERENT FORMULATIONS FOR NEMATODE MANAGEMENT IN IDAHO. Hafez, S.L. and M.P. Pudasaini. University of Idaho, Parma Research and Extension Center, 29603 University of Idaho Lane, Parma, Idaho 83660, USA Experiments were conducted to determine the efficacy of new chemistries such as Movento, Nimitz (MCW-2) and several numbered compounds for the management of major nematode species affecting potatoes, sugar beets and onions in Idaho. Potatoes, sugarbeets or onions were planted in a silt loam field infested with Columbia root-knot (Meloidogyne chitwoodi), sugarbeet cyst (Heterodera schachtii) and lesion nematodes (Pratylenchus spp.) at damaging levels. Treatments were replicated five or six times in a randomized complete block design. Movento was initiated at sufficient foliage and repeated every two weeks. Nimitz was applied pre-and post-plant. Within one hour of applications, all plots were disked twice to incorporate Nimitz to a depth of 10-15 cm. Numbered compounds were sprayed at 2.5-5 cm or 15-20 cm bands in furrow at planting, at 15 cm plant height, or chemigated at first irrigation and at monthly intervals. Total potato, sugar beet, onion, and infected potato yield was determined. The results demonstrated that the fall application of Vapam followed by Movento at 56 and 70 days after planting, or pre-plant application of Nimitz at low rate, or application of systemic number compound at 15 cm plant height and chemigated 60 days after planting are promising treatments for M. chitwoodi management on potatoes in Idaho. Movento applied 14 and 28 days after emergence and numbered compound applied at planting increased sugarbeet yield

172 Journal of Nematology, Volume 46, No. 2, June 2014 12-36%. Two applications of Movento and numbered compound sprayed at pre-plant increased onion yield 12-14%. No standalone chemical or single application can control nematodes. Multiple applications in combination with these soft chemicals are promising nematode management alternatives. INTERACTION OF LESION NEMATODES AND FUNGUS (VERTICILLIUM DAHLIAE) IN MINT. Hafez, S.L. and M.P. Pudasaini. University of Idaho, Parma Research and Extension Center, 29603 University of Idaho Lane, Parma, Idaho 83660, USA. An experiment was conducted to study the interaction of root lesion nematodes (Pratylenchus penetrans and Pratylenchus neglectus) and fungus (Verticillium dahliae) on mint. Treatments included no nematode, no fungus, either species alone, or combination of nematode and fungus. Each treatment had five replications spread on the greenhouse bench in random complete blocks. One six-week old mint plant was transplanted into 1500 cm3 pot filled with sand and soil mix (1:1 by v/v) with 10 % peat moss. Inoculations were done with 20 micro-sclerotia of V. dahlia or 4 nematodes per cm3 of soil. The mint was allowed to grow until flowering stage (about 10 weeks), and cut at soil level and allowed to regrow again. Mint was cut four times and top dry weight was recorded each time. Data demonstrates that in all cuts fungus or nematodes significantly and progressively decreased the mint hay dry yield as compared to control check. Interactive effect of V. dahliae and lesion nematodes appears to be an additive on mint hay yield. Verticilium dahliae alone caused 44 % damage in mint hay yield. Root lesion nematode P. neglectus seems pathogenic to mint. A 23 % and 46 % reduction on mint hay were caused by P. neglectus alone or in combination with V. dahliae, respectively. Pratylenchus penetrans alone caused 44% yield reduction of mint hay while combination of P. penetrans and V. dahliae killed almost all plants. Population of P. penetrans increased by 41 fold, indicating that mint is an excellent host for P. penetrans. TOLERANT VARIETIES AND REDUCED RATE OF TELONE II FOR SUGAR BEET CYST NEMATODE MANAGEMENT IN IDAHO. Hafez, S.L. and M.P. Pudasaini. University of Idaho, Parma Research and Extension Center, 29603 University of Idaho Lane, Parma, Idaho 83660, USA. Three experiments; response of tolerant sugar beet varieties, in-row and broadcast fumigation of Telone II at reduced rate in-strip, and a combination of tolerant sugar beet varieties with low rate of Telone were conducted to determine the response of sugar beet yield and sugar beet cyst nematode at the Parma Research and Extension Center, Parma, Idaho. The experiments were carried out in a randomized complete block design with seven treatments including one control, each with five replications in silt loam field. Telone II was shanked to a depth of 30 cm. Temik 15G @ 20 at plant and 14.57 kg/ha at post-plant was applied. Nematode tolerant and susceptible (Hillshog 9036RR) sugar beet varieties were sown for these trials. Suager beets were harvested and weights were taken from all experiments. The results demonstrated that sugar beet yield was significantly increased in all tolerant varieties as compared to the susceptible variety. Final viable cysts were increased by four fold in the susceptible variety while there were no increases in tolerant varieties. The sugar beet yield was significantly increased in both in-row and broadcast application of Telone as compared to untreated control and Temik. All the rates of Telone in-row had significantly higher sugar beet yield as compared to broadcast applications. There were no significant differences in the sugar beet yield among the different rates in in-row or in broadcast applications of Telone. Sugar beet yield was increased in the susceptible variety when fumigated plots were compared with non-fumigated plots. Sugar beet yield of tolerant varieties were increased in fumigated plots compared to non-fumigated plots. SCOPE OF NEW NEMATICIDES AND NUMBERED COMPOUNDS FOR COLUMBIA ROOT KNOT NEMATODE MANAGEMENT IN IDAHO. Hafez, S.L. and M.P. Pudasaini. University of Idaho, Parma Research and Extension Center, 29603 University of Idaho Lane, Parma, Idaho 83660, USA. Experiments were conducted to determine the efficacy of Movento, Nimitz (MCW-2 15G), and numbered compounds against Columbia root-knot nematode (Meloidogyne chitwoodi). Ranger Russet potatoes were planted in silt loam fields with an average initial nematode population of 374 per 500 cm3 soil. Treatments were replicated five times in a randomized complete block design. Movento was initiated at sufficient foliage and repeated every 2 weeks. Nimitz was applied pre-and post-plant. Within one hour of applications, all plots were disked twice to incorporate Nimitz to a depth of 10-16 cm and then hills were formed. Numbered compounds were sprayed at 2-5 cm or 15-20 cm bands in furrow at planting, or at first irrigation and at monthly intervals. After harvest, total and infected potato yield was determined. The findings demonstrate that Vapam in combination with Movento resulted in the lowest infected tuber yield (2.5 %) and provided superior performance as compared to either product alone, Movento (38-42 %) and Vapam (17 %). Nimitz lowered the infected potato yield in all treatments (3.4 to 7. 3%) as compared to untreated control (14.5%). The low rate of Nimitz applied pre-plant reduced infected tuber as compared to post-plant application. The infected yield was lower in all new numbered compounds tested (< 1.2 %) as compared to the untreated control (9.2 %). In conclusion, the fall application of Vapam followed by two foliar applications of Movento at 56 and 70 days after planting, or pre-plant application of Nimitz at low rate appears to be a promising treatment for Columbia root-knot nematode management on potatoes in Idaho.

Abstracts 173 THE MANAGEMENT OF PLANT-PARASITIC NEMATODES IN ORGANIC HORTICULTURE. Hallmann, J. Julius Ku¨hn-Institut, Federal Research Centre for Cultivated Plants, Institute for Epidemiology and Pathogen Diagnostics, Toppheideweg 88, 48161 Mu¨nster, Germany. Plant-parasitic nematodes are a major threat to organically grown vegetables. High intensity of the production system allows the build-up of high nematode densities. The most commonly found plant-parasitic nematodes causing problems under temperate conditions in Europe are Meloidogyne hapla, Pratylenchus penetrans and Paratylenchus bukowinensis. Symptoms and economic losses are usually first seen on carrots and onions. In the past, organic farmers often used black fallow to reduce nematode numbers. However, as this will also diminish the organic matter content of the soil it is not a recommendable method. Over the past years, we developed rotation systems based on trap crops and non-host crops that reduce nematode numbers to a similar degree as black fallow but at the same time provide positive side effects like increasing organic matter content or nitrogen fixation. Good control of M. hapla can be achieved by growing fodder radish as trap crop during the season> If nitrogen fixation is desired, growing legumes over winter and incorporating them around mid-June just before the nematode cycle is completed. Best control of P. penetrans was achieved by growing Tagetes for three months. Growing fodder radish as green manure in late summer also helped to reduce P. penetrans significantly. Paratylenchus bukowinensis with its narrow host range comprising crops within the families Brassicaceae (e.g. fodder radish, rape) and Umbelliferae (e.g. carrot, celery, parsley) can be easily controlled by avoiding these host plants. However, a prerequisite for the success of all these measures is proper control of weeds, which are in general excellent hosts for M. hapla and P. penetrans. PRELIMINARY RESULTS ALLUDE TO POTENTIAL ROOTKNOT NEMATODE TOLERANCE IN INDUCED POLYPLOIDS OF PLECTRANTHUS ESCULENTUS. Hannweg1, 2, K., W. Steyn1, M. Daneel1, A. Sippel1 and I. Bertling2. 1 Agricultural Research Council – Institute for Tropical and Subtropical Crops, Private Bag X11208, Nelspruit, 1200, South Africa; 2Horticultural Science, School of Agricultural, Earth and Environmental Sciences, University of KwaZulu-Natal, Pietermaritzburg, South Africa. Plectranthus esculentus, a member of the Lamiaceae family, is an important tuberous vegetable in Africa. It occurs in dry forest areas in central Africa, and southwards to Angola, the eastern parts of Limpopo and Mpumalanga and KwaZulu-Natal coast of South Africa, and Swaziland. P. esculentus is reported to be highly susceptible to rootknot nematode, Meloidogyne spp., which substantially reduces plant yield. In order to develop selections with potentially improved nematode tolerance, several polyploid lines were developed in vitro using colchicine treated shoot cultures and verified using flow cytometry. Hardened-off diploid (control) and tetraploid plants were inoculated with 1000 J2 eggs of Meloidogyne species and evaluated after 56 days for the number of egg masses and number of eggs and J2 per root system. The tetraploid plants had significantly fewer egg masses, eggs and J2 per root system compared with the diploid controls. Further, the tetraploids, had far lower Rfvalues than the diploids indicating a higher tolerance for rootknot nematode infection. The results of a preliminary in vitro nematode challenge trial will also be presented. Further research concerning the mechanism of tolerance is planned. SITE SPECIFIC APPLICATION OF FUMIGANTS FOR CONTROL OF NEMATODES IN TEXAS. Haygood1, R., J. Woodward2, N. Foster3 and C. O’Hara1. 1Dow AgroSciences, Indianapolis, Indiana, USA; 2Texas A&M AgriLife Extension, Lubbock, Texas, USA; 3Texas Tech University, Lubbock, Texas, USA. Control recommendations for plant pathogenic nematodes are based on species present, soil types, nematicide efficacy, economics, and other variables. Research results show that the use of soil maps and ECa data can be used effectively to identify low and high risk, or ‘‘responsive’’ zones for root-knot nematodes (Meloidogyne incognita). As a result, input costs for the application of TeloneÒ II fumigant can often be reduced by 30 – 40% by using this technological approach to assist in nematode management. Advances in Geographic Information Systems (GIS), Global Positioning Systems (GPS), apparent Electrical Conductivity (ECa) data collection, applicator equipment, and an increase in user expertise has enabled broader adaption and investigation of cotton yield responses. These studies were conducted to further define the value of site specific applications in fields with variable nematode populations and soil types in Texas. BIOLOGICAL AND SYSTEMATIC IMPLICATIONS OF PHYLOGENETIC ANALYSES OF ; 2,800 FULL LENGTH SMALL SUBUNIT RIBOSOMAL DNA SEQUENCES. Helder1, J., P. Mooijman1, S. van den Elsen1, H. van Megen1, M. Vervoort1, C. Quist1, W. Bert2, A. Karegar3, G. Karssen4 and W. Decreamer2, 5. 1Laboratory of Nematology, Department of Plant Sciences, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands; 2 Ghent University, Department of Biology, Nematology Unit, Ledeganckstraat 35, 9000 Ghent, Belgium; 3Department of Plant Protection, College of Agriculture, Shiraz University, Shiraz, Iran; 4Plant Protection Service, Wageningen Nematode Collection, Geertjesweg 15, 6706 EA Wageningen,The Netherlands; 5Royal Belgian Institute of Natural Sciences, Brussels, Belgium. As compared to other ancient and basal Ecdysozoan phyla such as the Nematomorpha (; 350 known species), the Priapulida (;16 species) and the Kinorhyncha (;180 species), the phylum Nematoda stands out for being speciose

174 Journal of Nematology, Volume 46, No. 2, June 2014 (; 27,000 described species), highly abundant, and widespread in nearly all terrestrial, freshwater and marine habitats. We will present results of phylogenetic analyses of approximately 10% of the described nematode biodiversity (; 2,800 taxa; with underrepresentation of marine and tropical terrestrial species). We have tried multiple genes for phylogenetic reconstruction but so far the small subunit of the ribosomal DNA (; 1,700 bp) is the only gene that could easily be amplified and aligned for a wide range of nematode species. The resulting alignment including secondary structure information was analysed on the CIPRES Science Gateway (San Diego, USA), using RAxML, a maximum likelihood-based inference for large phylogenetic trees, and Bayesian inference, and the outcome of both analyses will be presented. Some relevant features of the resulting phylogenetic trees will be discussed, and attention will be paid as to whether or not such a single gene tree does provide us with useful information about organismal relationships. This talk will also be used to describe the positioning and the relationships between the nematode taxa that will be discussed in more detail in Theme 2: Morphology, taxonomy, phylogeny and classification. EFFECTS OF SUNN HEMP AND PIGEON PEA ON BENEFICIAL AND PLANT-PARASITIC NEMATODES IN THE TROPICS. Henmi, V.H. and S.P. Marahatta. Science and Math Division, Kaua’i Community College, University of Hawai’i Plant-parasitic nematodes such as burrowing nematode (Radopholus similis) and root-knot nematode (Meloidogyne spp.) are dominant in banana, Musa spp. ecosystems. Beneficial nematodes are also found in banana fields. A tropical cover crop, sunn hemp (Crotalaria juncea), can be used to suppress plant-parasitic and enhance beneficial nematodes. However, sunn hemp cultivation in Hawaii is under the threat of the flour beetle. Two experiments: Trial-I and Trial-II were conducted to compare the effects of another tropical cover crop, pigeon pea (Cajanus cajan)) with sunn hemp and no-cover crop control on Radopholus similis and Meloidogyne suppression and beneficial nematode enhancement. In both experiments soils infested with R. similis and Meloidogyne were sampled and amended with cover crop treatments or no-cover crop control and kept for two weeks. At the end of each experiment, nematodes were extracted through the Baermann funnel technique. The results of Trial-I and Trial- II showed that sunn hemp and pigeon pea did not reduce R. similis number (P > 0.05). However, Meloidogyne numbers were consistently reduced by sunn hemp and pigeon pea (P < 0.05). In both experiments sunn hemp increased beneficial nematodes number (P < 0.05). Pigeon pea increased beneficial nematode numbers in Trial -I (P < 0.05), but not in Trial –II (P > 0.05). However, the numbers of beneficial nematodes were consistently higher in pigeon pea compared to the no-cover crop control. Farmers could choose pigeon pea as an alternate to sunn hemp, as a cover crop for Meloidogyne suppression and beneficial nematode enhancement. PRISTIONCHUS SCRATCHPAD – AN ONLINE PLATFORM FOR TAXONOMY, SYSTEMATICS AND PHYLOGENY. Herrmann, M. and R.J. Sommer. Max Planck Institute for Developmental Biology, Department of Evolutionary Biology, Spemannstraße 37, 72076 Tu¨bingen, Germany. The utility of the nematode Pristionchus pacificus as a model system in evolutionary biology and comparative developmental biology builds on the availability of a sophisticated genetic and molecular toolkit with a resolved phylogenetic context. Specifically, surveys have recovered several new species of Pristionchus, which are particularly diverse in East Asia. In total, 29 Pristionchus species have been characterized by molecular, morphological and in some cases genetic tools. All of these species, often including independently collected wild isolates, are available as live and frozen stocks in the Tu¨bingen lab collection and can be distributed upon request. Similarly, a broad framework of 24 genera of the Diplogastridae family has been established, many of which are also available as live or frozen stocks. This includes among others, species of four newly described genera (e.g. Parapristionchus, Sudhausia), which have been characterized in recent years. To provide a transparent system for the taxonomy, biosystematics and phylogeny of Pristionchus and ultimately the Diplogastridae, we are establishing an online platform, Pristionchus Scratchpad (following the guidelines of scratchpads.eu), which can help researchers throughout the world to obtain an overview of published work and to request cultured isolates. Specifically, Pristionchus Scratchpad provides information on i) a species overview with published references and PDFs, ii) species distribution maps, iii) morphological characterization, iv) morphometrics and v) SSU rRNA and other molecular sequence tags. We hereby present Pristionchus Scratchpad as a model for an online platform of modern taxonomy, biosystematics and phylogenetic research in nematodes. COMPARISONS OF NEMATODE BIODIVERSITY USING MORPHOLOGICAL, MOLECULAR, ECOLOGICAL AND BIOLOGICAL CRITERIA. Hodda, M. CSIRO Ecosystem Sciences, GPO Box 1700 Canberra ACT 2601 Australia. There is an increasing trend towards assessments of nematode biodiversity based on molecular criteria only. This is understandable given the greater ease and automation possible assessing biodiversity using molecular techniques. Much greater expertise and resources are required to assess diversity using morphology in any sophisticated way corresponding to traditional taxonomy, or using ecological criteria such as host relationships, or using biological criteria such as interbreeding success. Even using ‘‘morphospecies’’, requires a considerable investment to assess nematode diversity in any meaningful way. Whatever their practicalities, when the results of these different measures of nematode biodiversity are compared, the

Abstracts 175 results are seldom exactly the same: diversity has been frequently underestimated by most methods, but that it may be overestimated by the new technique of ultrasequencing. However, these results depend on the species concept used, and the different species concepts may have different abilities to supply relevant measures of nematode biodiversity, depending on what diversity is being measured for. The different uses of nematode biodiversity are reviewed. NEMATOLOGY COMMUNICATION & EDUCATION: FROM FARMERS TO POSTDOCTORAL RESEARCHERS & EVERYONE BETWEEN. Hodda1, 2 , M., N. Tangchitsomkid3, K.A. Davies4, S.K. Singh5 and N.C. Banks1,6,7. 1CSIRO Ecosystem Sciences, GPO Box 1700, Canberra, ACT 2601, Australia; 2Australian National Insect Collection, GPO Box 1700, Canberra, ACT 2601, Australia; 3Thailand Department of Agriculture, 50 Phaholyothin Road, Chatuchak, Bangkok, 10900, Thailand; 4School of Agriculture Food & Wine, University of Adelaide, PMB 1, Glen Osmond, SA 5064, Australia; 5 Agricultural & Wine Sciences, Charles Sturt University, Locked Bag 588 Wagga Wagga, NSW 2678, Australia; 6CSIRO Biosecurity Flagship, 41 Boggo Rd Dutton Park, QLD 4102 Australia; 7School of Veterinary and Life Sciences, Murdoch University, 90 South Street, Murdoch WA 6150, Australia. Nematology is of necessity a very broad scienfic discipline, encompassing everything from the very practical (crop diseases) to the theoretical (species concepts); from the finely detailed (virulence genes) to the broad general (soil health); and from the local (species lists) to the global (biosecurity). This breadth should mean that interest is healthy, but too frequently the reaction is the opposite. Based on experience teaching everyone from farmers with no initial knowledge of nematodes to postdoctoral researchers, we suggest that there are several elements to successful education and communication about nematodes. First, a certain amount of basic knowledge is necessary as background. Second, information and communications need to be distilled into a few simple messages. This can be a considerable task. Third, that the details and exceptions and complicating factors are important, and recognizing how basic principles are modified in different situations is important in applying new knowledge about nematodes to particular situations. The skills to be able to recognize local variations are very important. Fourth, the practical uses of nematology knowledge need to be emphasized. Most people dealing with nematodes are non-specialists and require frequent updates, refreshers and a graded series of interactions, often best delivered over several presentations spaced some time apart to allow the information to sink in and avoid overload. SUBERIN AND LIGNIN – BARRIERS TO NEMATODE ENTRY? Holbein1, J., R.B. Franke2, F.M.W. Grundler1 and S. Siddique1. 1Rheinische Friedrich-Wilhelms University of Bonn, INRES, Molecular Phytomedicine, Karlrobert-Kreiten Str. 13, 53115 Bonn, Germany; 2Rheinische Friedrich-Wilhelms University of Bonn, IZMB, Ecophysiology, Kirschallee 1, 53115 Bonn, Germany. The essential function of a plant root is the uptake of water and minerals from the soil and their distribution to the aerial organs via the vascular bundle. Due to their vital function, these vessels are protected from external biotic and abiotic stresses by a tissue called endodermis. Special cell wall modifications in forms of biopolymer depositions enhance the barrier function of the endodermis. Primary walls of the endodermal cells are impregnated with suberin, while radial and transverse walls are additionally reinforced by belt like lignin structures, called casparian strip (CS). It has been proposed that suberin and casparian strip may act as insuperable barriers to nematodes invading the roots for feeding. In fact, the root knot nematode (RKN) Meloidoygyne incognita evade the endodermal barriers by first moving towards the root tip between cortical cells, and then once at the tip taking a U-turn to enter the vascular cylinder where they initiate feeding sites, whereas, the migratory nematode Pratylenchus penetrans feeds almost entirely from the cortex of roots suggesting an inability to cross endodermal barriers. To investigate the role of endodermal suberin and lignin during nematode infection, Arabidopsis lines and accessions were screened with altered levels of suberin and lignin against three different nematode species (Heterodera schachtii, Meloidogyne incognita and Pratylenchus spp.). These nematodes represent three different types of migration and feeding behavior. Our data showed that susceptibility of plants decreases significantly with increase in suberin content suggesting that root barriers play important role during plant-nematode interaction. Histochemical, analytical and molecular analyses have been performed to understand how enhanced suberin is mechanistically related to decrease nematode susceptibility. LABEL-FREE COHERENT RAMAN SCATTERING (CRS) IMAGING FOR METABOLIC PROFILING OF NEMATODES. Holden-Dye1, L., N. Dalliere1, P. Urwin2, C. Lilly2, V. O’Connor1, J. Pertek1 and S. Mahajan1. 1University of Southampton, Institute for Life Sciences, Highfield Campus, University of Southampton, Southampton, SO17 1BJ, UK; 2 Centre for Plant Sciences, School of Biology, University of Leeds, Leeds LS2 9JT, UK. Fundamental aspects of the biology of plant-parasitic nematodes are relatively underexplored however there is evidence to suggest that a major factor regulating their ability to infect the host crop is their lipid reserve. This is particularly important with respect to the phenomenon of field persistence whereby plant-parasitic nematodes that have thrived on a host crop deliver progeny with high lipid reserves, and thus high infectivity, back to the field. This positive feedback loop contributes to the persistence of crop infection and damage. Therefore there is an increasing appreciation that the key to identifying resistance breaking approaches and/or more efficacious ways of using existing practices will derive from improved understanding of the metabolic reserve of these agriculturally important parasites. We are deploying spectroscopic

176 Journal of Nematology, Volume 46, No. 2, June 2014 methodologies that can detail metabolic analysis and enhance metabolomics investigation. Coherent Raman Scattering (CRS) spectroscopy and microscopy are label-free techniques which allow high-resolution, non-invasive and non-destructive chemically specific imaging. Thus CRS can monitor key biochemical signatures in vivo and is well suited to such an investigation. We have preliminary data to show precise quantification of lipid stores and their tissue redistribution, responding to environmental and chemical manipulation. SPATIO-TEMPORAL DISTRIBUTION OF RENIFORM NEMATODE (ROTYLENCHULUS RENIFORMIS) AND RELATIONSHIPS WITH SOIL TEXTURE. Holguı´n1, C.M., P. Agudelo1, P. Gerard2, J.D. Mueller3 and A. Khalilian3. 1 School of Agricultural, Forest, and Environmental Sciences. Clemson University, Clemson, South Carolina 29634, USA; 2 Mathematical Sciences. Clemson University, Clemson, South Carolina 29634, USA; 3Edisto Research and Education Center, Clemson University. Blackville, South Carolina 29817, USA. Densities of reniform nematode, Rotylenchulus reniformis, can change in time and space within a field affecting sampling and management strategies. A three-year study was conducted in two fields in South Carolina to assess the horizontal and vertical distribution of reniform nematode and determine potential correlations with soil texture. Cotton-peanut and cottoncorn-soybean rotation systems occurred in fields 1 and 2 respectively. Each year, soil samples were collected in both fields at planting and after harvest. In field 1, 40 samples were collected at random locations. Sample cores were separated into four depths: 0-15 cm, 15-30 cm, 30-60 cm, and below 60 cm. In field 2, samples were collected from 80 4 x 4 m plots distributed within four sections of the field representing different soil electrical conductivity readings and textures. Reniform nematode densities within each field/plot showed high levels of spatial variability during the length of the study. Horizontal distribution analysis using the variance/mean ratios showed a significant clustered distribution at planting and after harvest in both fields during the three years. However, a significant neighbourhood structure was only detected in the first two years in field 2, with patches between 8 and 18 square meters. Vertical analysis showed the highest numbers of reniform nematode between 15and 30-cm deep. In some of the plots R. reniformis densities were significantly correlated with sand and clay content. In this study, the choice of sampling strategy, host as well as soil texture affected the spatial pattern of reniform nematode that was revealed. STYLETCHIP: A MICROFLUIDIC DEVICE FOR RECORDING HOST INVASION BEHAVIOR AND FEEDING OF PLANT PARASITIC NEMATODES. Hu1, C., J. Kearn2, P. Urwin3, C. Lilley3, V. O’Connor2, L. Holden-Dye3 and H. Morgan1. 1Faculty of Physical Sciences and Engineering, and Institute for Life Sciences, Bassett Crescent East, University of Southampton, Southampton, SO17 1BJ; 2University of Southampton, Centre for Biological Sciences, Bassett Crescent East, University of Southampton, Southampton, SO17 1BJ; 3University of Leeds, Centre for Plant Sciences, School of Biology, University of Leeds, Leeds, LS2 9JT. Plant parasitic nematodes (PPNs) infest the roots of crops and cause global losses with a severe economic impact on food production. Current chemical control agents are being removed from use due to environmental concerns and there is a need for new approaches in crop protection. A key feature of PPNs is a hollow stylet required for interaction with the host plant and feeding. This lance-like structure protrudes from the mouth of the worm and thrusts in a rhythmic manner to stab the host root. The underlying biology of stylet behaviour is poorly understood. We have addressed this by designing a microfluidic chip which traps the PPN Globodera pallida and permits the recording of an electrophysiological signal concomitant with stylet thrusting. The PDMS chip incorporates a precisely designed aperture to trap the nematode and valves for rapid application of test compounds and integral electrodes to facilitate acquisition of electrical signals. We show that stylet thrusting can be induced by controlled application of serotonin to the nematode. Each thrust and retraction produces an electrical waveform that characterises the physiological activity associated with the worm’s behaviour. The ability to reproducibly record stylet activity provides a new platform for nematicide screening that focuses on a behaviour that is integral to the parasite host interaction. This is the first report of a microfluidic chip capable of electrophysiological recording from nematodes other than Caenorhabditis elegans. This approach may also be applied to other species of economic or medical importance. MOLECULAR CLONING AND CHARACTERIZATION OF MJ-1-1 FROM MELOIDOGYNE JAVANICA. Hu1,2, L., K. Zhuo1,2, J. Liao1,2. 1Laboratory of Plant Nematology, South China Agricultural University, Guangzhou 510642, China; 2 Guangdong Province Key Laboratory of Microbial Signals and Disease Control, South China Agricultural University, Guangzhou 510642, China. Root-knot nematode, Meloidogyne, is one of the globally economically most damaging plant-parasitic nematodes causing great losses to many. Recently, some work has been done to explain the molecular mechanism involved in nematode infection. In the present study, a gene Mj-1-1, which encodes for a 149 aa protein with a predicted molecular mass of 17.982 KDa was isolated from Meloidogyne javanica. A real-time reverse-transcriptase polymerase chain reaction assay showed that expression of Mj-1-1 was upregulated in later parasitic stages. The highest transcriptional level of Mj-1-1 occurred in thirdstage juveniles, and this high expression persisted in parasitic fourth-stage juveniles. In planta RNA interference targeting

Abstracts 177 Mj-1-1 suppressed the expression of Mj-1-1 in nematodes and attenuated the parasitism ability of M. javanica. Meanwhile overexpression of Mj-1-1 rendered tobacco susceptible to M. javanica. It can be concluded that Mj-1-1 may play a role in parasitism of M. javanica, especially in later parasitic stages. ENTOMOPATHOGENIC NEMATODES AND ENDOPHYTIC BACTERIA: A NOVEL APPROACH WITH POTENTIAL FOR PLANT HEALTH. Hurley, M.J., D. Brazil and T. Kakouli-Duarte. Molecular Ecology and Nematode Research Group, EnviroCORE, Department of Science and Health, Institute of Technology Carlow, Kilkenny Road, Carlow, Ireland. Entomopathogenic nematodes (EPN) and bacterial endophytes have considerable potential in biological control and plant growth promotion, respectively. The use of EPN with various crop protection or improvement agents has been well documented. Similarly, there is evidence that bacterial endophytes can promote plant health and soil fertility. Although EPN and bacterial endophytes naturally function in a common habitat and have widespread agrobiological applications, the potential of synergy has yet to be explored. Mobilising the interaction of different nematode taxa with naturally occurring soil microbiota may result in increased plant growth and health, in addition to a reduction in the use of non-specific chemical fertilisers and pesticides. This project investigates the interaction between these two naturally occurring taxa. In preliminary experiments three EPN species, Steinernema feltiae, Steinernema carpocapsae and Heterorhabditis bacteriophora, were investigated for their interaction with a range of bacterial endophytes from our strain bank at EnviroCORE. In both laboratory and greenhouse trials the bacterial endophytes had no adverse effects on the ability of the EPN to kill Galleria mellonella hosts. In experiments where G. mellonella were exposed to increasing nematode doses, our results indicate that the presence of Pseudomonas fluorescens F113 and S118 caused H. bacteriophora to enter the insects at higher numbers. Experiments are currently ongoing in order to observe possible effects of the endophytes on S. feltiae and H. bacteriophora life cycle. SUSCEPTIBILITY OF PASSION FRUIT SELECTIONS FROM THE ARC-ITSC BREEDING PROGRAMME TO ROOT-KNOT NEMATODE (MELOIDOGYNE INCOGNITA AND MELOIDOGYNE JAVANICA). Husselman, J.H., M.S. Daneel and W.P. Steyn. Agricultural Research Council - Institute for Tropical and Subtropical Crops, Private Bag X11208, Nelspruit, 1200, South Africa. Passion fruit (Passiflora edulis) is a climber native to South America that is cultivated for its aromatic fruit in frost free areas around the world. In South Africa, production is primarily in Mpumalanga and Limpopo provinces with minor plantings in KwaZulu-Natal and the Eastern and Western Cape. The South African industry relies mainly on one cultivar namely ‘Ester’, a hybrid between the purple (Passiflora edulis f. edulis) and yellow granadilla (Passiflora edulis f. flavicarpa). Although viruses are the most important problem in the crop with no chemical cure available, leaf and soil diseases and nematodes also create considerable problems and selections from the breeding programme are screened for tolerance/ resistance to these pests and diseases. Breeding parents are selected for good horticultural characteristics and tolerance to the pests and diseases. A trial to test nematode resistance was conducted with seven selections and twelve replicates in a complete randomised design. The purple commercial standard Ester (Passiflora edulis f. edulis) was included as a susceptible control and yellow granadilla (Passiflora edulis f. flavicarpa) as a tolerant control. The selections were multiplied by air-layers, transplanted in 2 litre plastic bags fill with sterilized sand eight weeks later after which the plants were placed in a glasshouse to establish for four weeks before being inoculated with 3200 Meloidogyne eggs/bag. Evaluation done at 56 days after inoculation showed low infection rate and evaluation was extended. Results will be discussed but differences were observed with P. edulis ‘Ester’ and P. edulis f. edulis having the highest nematode numbers. ACTIVITY PROFILING REVEALS CHANGES IN THE DIVERSITY AND ACTIVITY OF PROTEINS IN ARABIDOPSIS ROOTS IN RESPONSE TO NEMATODE INFECTION. Hu¨tten1, M., M. Geukes1, J. C. Misas-Villamil2, S. Habash1, A. Elashry1, R. A. L. van der Hoorn2,3, F. M. W. Grundler1 and S. Siddique1. 1Rheinische FriedrichWilhelms-University of Bonn, INRES – Molecular Phytomedicine, Karlrobert-Kreiten-Straße 13, 53115 Bonn, Germany; 2 Plant Chemetics lab, Max-Planck-Institute for Plant Breeding Research, Carl-von-Linne´-Weg 10, 50829 Cologne, Germany; 3Plant Chemetics lab, Department of Plant Sciences, University of Oxford, South Parks Road, OX1 3UB Oxford, UK. Cyst nematodes are obligate, sedentary endoparasites with a highly specialised biology and great economic impact in agriculture. The development of a syncytium is aided by a cocktail of nematode effectors that manipulate host plant activities in a complex network of interactions by post-translational modifications. Previous transcriptome and proteome studies of syncytia generated a wealth of data that is based on abundance rather than activity of transcripts or proteins. Activity-Based Protein Profiling (ABPP) was recently introduced in plant sciences and has been proven highly useful to display differential enzymatic activities of proteins by using activity based probes (ABPs). ABPs are small molecular probes (biotinylated or fluorescent) that react with a specific subset of enzymes in an activity-dependent manner ruling out all those proteins, which are inhibited, inactive or lack cofactors. To better understand functional proteomics that lead to formation of syncytia, ABPP was conducted on syncytia induced by the beet cyst nematode Heterodera schachtii in Arabidopsis roots. This approach has identified genes and pathways that may play essential roles in feeding site development. Our data show that activity of several

178 Journal of Nematology, Volume 46, No. 2, June 2014 papain-like cysteine proteases (PLCPs) and proteasomal subunits that are involved in activation of plant immune responses after pathogen attack is specifically suppressed in syncytia. Moreover, we identified effector proteins from H. schachtii that may target and inhibit PLCPs and proteasome in host plants. Our research contributes to a broader framework in understanding of cyst nematode parasitism and provides a platform to develop novel solutions against these pathogens. GENE NETWORKS AND LIPID SIGNALS GOVERNING PLANT INTERACTIONS WITH THE ROOT-KNOT NEMATODE MELOIDOGYNE JAVANICA. Iberkleid1,2, I., N. Sela1 and S. Brown Horowitz1. 1Department of Entomology, Nematology and Chemistry units, Agricultural Research Organization, the Volcani Center, Bet Dagan, 50250, Israel; 2Department of Plant Pathology and Microbiology, the Faculty of Agriculture Food & Environment, the Hebrew University of Jerusalem, Rehovot, 76100, Israel. Plant parasitic nematodes produce a unique class of small helix-rich fatty acid and retinol binding proteins (FAR) with no counterpart in their plant hosts. Meloidogyne javanica’s Mj-FAR-1 is critical for the juvenile and sedentary development as well as the giant cell development presumably through compromising the plant defense by modulating plant lipid signals. For further insights into the role of Mj-FAR-1 protein in regulating disease development, we compared gene expression profiles of tomato hairy roots lines in which mj-far-1 is constitutively expressed and control roots at early and late stages after inoculation. Gene expression profiling revealed that 3970 transcripts were differentially expressed between the two lines; with 2069 up-regulated and 2205 down-regulated in the mj-far-1 overexpressing line (OE) compared with the control root line over all inoculated and non-inoculated samples. A total of 61 up-regulated and down-regulated genes were overlapped between all non-inoculated and inoculated OE roots. This list might provide initial understanding of the mj-far-1 associated increased susceptibility that occurs in mj-far1-1 root line. These included genes involved in fatty acid metabolism, a group of hormone signals including Jasmonic acid and Auxin related genes, biotic stress related genes, and genes of the phenylpropanoid pathway that were also consistently differentially regulated. Validation of the data was confirmed through the use of quantitative real-time PCR of representative differentially expressed genes and through the use of b-glucuronidase reporter gene downstream of candidate gene promoters. Overall, the transcriptomic analysis indicates that, at the early time points samples clustered predominantly in relation to the differential genetics while at later time points samples clustered in relation to temporal dynamics related to nematode infection effect. Our results will shed light on the role of lipid signals and mj-far1 effect in defining a common transcriptome that facilitate nematode infection. STUDIES ON THE EFFECT OF HETERODERA AVENAE (WOLLENWEBER, 1924) ON YIELD LOSSES IN WHEAT VARIETIES IN THE EASTERN MEDITERRANEAN OF TURKEY. Imren1, M., H. Toktay2, E.B. Kasapog˘lu3, A. _ Dababat4 and I.H. Elekciog˘lu3 1Abant Izzet Baysal University, Faculty of Agriculture, Department of Plant Protection 2 Bolu, Turkey; University of Nigde, Faculty of Agricultural Sciences and Technologies, Nigde, Turkey; 3Cukurova University, Faculty of Agriculture, Department of Plant Protection; 4CIMMYT (International Maize and Wheat Improvement Centre), Wheat Improvement Program, Ankara, Turkey. Globally Heterodera avenae (Wollenweber, 1924) is the most widely distributed and important species among cereal nematodes. The aim of the study was to determine yield losses caused by the H. Ha21 pathotype which was found in the Eastern Mediterranean region; one of the important spring wheat production centres in Turkey. Field trials were conducted during 2011 and 2012. The results of the study indicated that yield losses caused by H.avenae were between 4.36% - 25.7%, depending on the wheat variety. Bread wheat varieties, Adana 99 and Ceyhan 99, showed higher performance than the Silverstar wheat variety which is a known resistant variety against H. avenae. Although low nematode reproduction, high yield and less yield losses rates were found in Adana 99, high nematode reproduction rate, high yield rate and less yield losses were determined in Ceyhan 99. These findings suggest that Adana 99 is a moderately tolerant variety and Ceyhan 99 is a tolerant variety. FUNGAL PARASITES OF EGGS AND CYSTS OF GLOBODERA ROSTOCHIENSIS AS POTENTIAL BIOLOGICAL CONTROL AGENTS OF POTATO CYST NEMATODE IN INDONESIA. Indarti, S., Mulyadi, D. Widiayanto and J. Widada. Agriculture Faculty of Gadjah Mada University, Bulaksumur, Yogyakarta 55281, Indonesia. Globodera rostochiensis produces considerable damage in potato production areas in Indonesia, and the infested area continues to increase each year. Potato cyst nematodes (PCN) is difficult to control because the cyst protects the eggs from unfavourable environmental conditions allowing the eggs to survive for years, and hatch only in the presence of host root exudates. The aim of this research was to develop biocontrol agents using fungal parasites of eggs and cysts, isolated from soil found to be suppressive to PCN. The research was carried out in three stages: 1) identification of the soil that had the ability to suppress the development of PCN and analysis of antagonistic activity of fungi potentially involved in the suppression of nematode populations, 2) isolation of fungi and testing their ability to parasitize PCN eggs and cysts and 3) testing the effectiveness of the fungi to control nematode populations in laboratory and pot trials, thus at mesocosm as well as macrocosm levels. PCN infested soils with ability to suppress PCN multiplication were found in Central Java, Indonesia. Twelve fungal isolates with the ability to parasitise more than 50 % of the PCN cysts and with extracellular proteolytic

Abstracts 179 activity were isolated from these locations. These fungi gave effective control to PCN populations in laboratory and pot trials compared to untreated treatments. The best performing isolate of this group suppressed the PCN cyst multiplication by 2.4 times (30.07 cysts/ 100 g soil compared with 73.53 cysts/ 100 g soil on control or without fungal treatment). Tuber yield from treated plants were also 1.6 times higher than the control. PARTIAL SURVEY OF MELOIDOGYNE SPECIES ON COFFEE IN PARANA STATE, BRAZIL. Ito, D.S., A.C.Z. Machado, S.A. Silva and O.F. Dorigo. Insituto Agronoˆmico do Parana´, Rodovia Celso Garcia Cid, km 375, 86047-902, Londrina, PR, Brazil Parana´ is an important State of Brazil in coffee production. However, root-knot nematodes have been causing great losses for the coffee crop. The aim of this study was to carry out a survey on the distribution of root-knot nematodes in coffee crops in Parana´ State. For this, 50 samples of soil and roots were collected from coffee crops showing symptoms of nematode infestation, located in municipalities of Alto Parana´, Altoˆnia, Bela Vista do Paraı´so, Cambira, Carlo´polis, Cianorte, Corumbataı´ do Sul, Esperancxa Nova, Kalore´, Lupiono´polis, Moreira Sales, Munhoz de Melo, Paranavaı´, Perobal, Pe´rola, Pinhala˜o, Pitangueiras, Rolaˆndia, Santa Luzia, Sa˜o Jorge do Patrocı´nio, Sa˜o Tome´, and Xambre´. From the total of samples analyzed, 31 (62%) contained Meloidogyne paranaensis, nine (18%), M. incognita and two (4%), M. javanica. Four samples (8%) showed a mixture of M. incognita and M. paranaensis and four (8%) presented M. incognita and M. javanica. The main nematode species present in Parana´ State is M. paranaensis. These results will help in the mapping the distribution of nematodes in the coffee growing regions of Parana´, allowing to follow the dispersion of nematodes in the State and to manage more efficiently the populations present at each region. ON THE DIVERSITY OF NEMATODES PARASITIC IN INVERTEBRATES OF MINDANAO ISLAND, PHILIPPINES. Ivanova1, E.S., N.H.N. Sumaya2, A.B. Mohagan3 and S.E. Spiridonov1. 1A.N. Severtsov Institute of Ecology and Evolution, Russian Academy of Sciences, Leninskii pr., 33, Moscow, 119071, Russia; 2Biology Department, Mindanao State University-Iligan Institute of technology (MSU-IIT), Andres Bonifacio Ave., Tibanga, Iligan City, Philippines; 3Biology Department, University of Central Mindanao, University Town, Musuan, Maramag, Bukidnon, Philippines. Several representatives of interesting taxa of the nematodes of invertebrates were collected in the Island of Mindanao during a short field trip in November 2013. Large tropical millipedes of the order Spirobolida were found infected with a rich array of rhigonematids and ransomnematids inhabiting the hind gut of their host. For the first time since its description, predacious nematodes of the genus Zalophora Hunt 1994 were reported. Analysis of the D1D3 expansion segment sequences of Hethidae from Mindanao indicated close relationships with Indonesian species of Heth. Large tropical earthworms of the family Megascolecidae were collected in two habitats near Iligan City. The earthworms collected near Tinago Falls were found to be infected by the Synoecnema nematodes (Ungellidae). Phylogenetic analysis of LSU rDNA sequences has confirmed their independent position among other studied ungellids thus proving the validity of the genus Synoecnema. The earthworms collected in Initao-Libertad Protected Seascape and Landscape Forest were parasitized by nematodes of the genus Filiponema (nominal representative of the family Drilonematidae). These nematodes primarily found in the Philippines were also never molecularly characterised. Analysis of partial LSU rDNA sequence of Filiponema from Mindanao revealed their phylogenetic relationships with other tropical families of nematodes parasitic in earthworms (Ungellidae and Homungellidae), but not with Drilonematidae from temperate lumbricids (genus Dicelis). A mermithid from a spider caught on the Central Mindanao University campus, Musuan, Bukidnonwas characterised with LSU and SSU rDNA sequencing. NEMATODE ASSEMBLAGES AND MICROBIAL COMMUNITIES IN SOYBEAN CROPPING SYSTEMS. Jansen, C., S. Claassens and H. Fourie. Unit of Environmental Sciences and Management, North-West University, Potchefstroom 2520, South Africa. Soil is an important ecosystem that supports a wide variety of organisms such as arthropods, bacteria, fungi, and nematodes. This sensitive ecosystem is influenced by various factors of which intensive agricultural management practices are the most destructive. With the introduction of genetically modified (GM) glyphosate-resistant (Roundup Ready: RR) crops, herbicides such as glyphosate (Roundup) has been increasingly used. With the proximity that microorganisms and nematodes have with the roots of these plants, these organisms can be used as bio-indicators. The aim of this study was to compare microbial community structure and nematode diversity in soil samples from conventional- and Roundup Ready- soybean fields in soybean production areas of South Africa to that in adjacent, uncultivated grassland areas. Microbial community structure was determined by phospholipid fatty acid analyses. Nematode diversity was determined by extracting the nematodes from soil samples with standard methods and conducting faunal analyses. Results indicated a definite difference in microbial community structure between the various localities; however, no differences in community structure between Roundup Ready- and conventional soybean samples were evident. Soils of both Roundup Ready - and conventional soybean crops were primarily dominated by bacteria. Nematode analyses also indicated no differences between the soils of these two treatments. A complete faunal analysis will be discussed including correlations that might exist between microbes and nematodes associated with Roundup Ready and conventional soybean plantings as well as natural veld areas.

180 Journal of Nematology, Volume 46, No. 2, June 2014 NEMATODES FROM THE SEEKOEIVLEI NATURE RESERVE, MEMEL, SOUTH AFRICA. Jansen van Rensburg, C. and A. Mobara. Department of Zoology & Entomology, University of the Free State, P.O. Box 339, Bloemfontein, 9300, South Africa. In South Africa, the amount of taxonomic information for freshwater nematodes from wetlands is lacking, when comparing it to Europe. The Seekoeivlei Nature Reserve (45 km 2), situated in the north-eastern corner of the Free State Province near the town of Memel, is the largest protected area of wetlands on the Highveld and in South Africa. This marshland is a very important sponge area for the Klip River which is a tributary to the Vaal River, providing water to the highly industrialised and densely populated Gauteng Province. Nematode samples were collected from three localities within the reserve with the aims of determining the nematode genera present, taxonomically describing some of the species belonging to these genera, and to determine whether the nematodes present could give an indication of what the ecological status of the wetland is. Soil samples were collected using the core-syringe method and transported back to the laboratory at the University of the Free State, Bloemfontein campus where nematodes were extracted and prepared for light and scanning electron microscopy using standard techniques. A total of 45 genera belonging to 24 different nematode families were identified. Nematode genera data was used to determine the: Maturity Index (MI), Shannon-Wiener diversity index (H’), as well as Hill’s infinity (N‘). Results showed that nematode diversity was higher during the summer months compared to the winter months, with the Maturity index indicating that there may be some kind of ecological disturbance where the Klip River enters the Seekoeivlei Nature Reserve. Descriptions are currently underway for species belonging to the genera Brevitrobrilus, Chronogaster, Eutobrilus and Tobrilus. PROVIDING A LINK BETWEEN MORPHOLOGICAL DESCRIPTIONS AND DNA BARCODING TO STUDY SPECIATION OF THE GENUS PRATYLENCHUS IN A ROBUST PHYLOGENETIC FRAMEWORK. Janssen1, T., G. Karssen2, L. Waeyenberge3, M. Couvreur1 and W. Bert1. 1Nematology Research Unit, Ghent University, Department of Biology, Ledeganckstraat 35, 9000 Ghent, Belgium; 2Plant Protection Service, Nematology section, P.O Box 9102, 6700 HC Wageningen, The Netherlands; 3Institute for Agricultural and Fisheries Research, Crop Protection, Burg Van Gansberghelaan 96, B-9820 Merelbeke, Belgium. Root-lesion nematodes of the genus Pratylenchus are an important pest in economic crops. They have a worldwide distribution (except for the Polar Regions) and are poplyphagous. At the moment the genus consists of more than 70 species. The vast majority of these descriptions is based only on morphological and morphometric characters. However, it is well known as many authors have already stated that reliable identification of many species is impossible due to the lack of diagnostic characters, morphological interspecific plasticity and incomplete taxonomic descriptions. This often results in the impossibility to link original descriptions, based on morphological characterization, and contemporary molecular taxonomy. This task is further impeded as we demonstrated cryptic speciation in Pratylenchus goodeyi, providing additional evidence that cryptic speciation is common within the genus. To assess this species complex we investigated a new set of promising mitochondrial DNA markers for molecular species delimitation and barcoding. These new mitochondrial markers will be used to develop primers for mitochondrial genome sequencing. Finally, these mitochondrial genomes will be used to construct a robust phylogenetic framework to study speciation and allow an improved characterization and proper identification of Pratylenchus species. MOLECULAR INSIGHTS INTO THE INTERACTION BETWEEN RICE AND ENDOPARASITIC NEMATODES. Ji, H., K. Nahar, L. Bauters, G. Gheysen and T. Kyndt. Department of Molecular Biotechnology, Ghent University, Coupure links 653, 9000 Gent, Belgium. Rice (Oryza sativa L.) is one of the major staple foods in the world, and an interesting model monocot plant. Two nematodes, causing extensive crop yield losses, are Meloidogyne graminicola and Hirschmanniella oryzae. The response of rice upon infection with these nematodes was studied using transcriptome analyses on local and systemic tissues, at different time points and comparing the two different nematodes. The role of hormone pathways in the plant defence system was investigated using infection experiments after exogenous chemical treatments and in rice mutants/ transgenics altered in different hormone pathways. Our transcriptome data show a strong and fast reduction of the plant defence system upon rice infection with the root-knot nematode M. graminicola, not only in local but also in systemic tissues. The SA and JA pathways were both strongly suppressed inside the nematode feeding sites, but activation of these pathways could counteract nematode infection substantially. The brassinosteroid pathway promotes plant susceptibility for root knot nematodes by its antagonistic interaction with the classical defence pathways. The strong systemic defence suppression upon root knot nematode infection leads to a significantly enhanced susceptibility for rice blast (Magnaporthe oryzae) in the above-ground tissues. In case of H. oryzae infection, a systemic suppression of the SA pathway was detected, potentially caused by the nematodes secretion of two effectors: chorismate mutase and isochorismatase. Over-expression of these effectors in transgenic rice lines leads to a significant interference with genes involved in SA-production.

Abstracts 181 TOP 10 PLANT PARASITIC NEMATODES IN MOLECULAR PLANT PATHOLOGY. Jones1, J.T., A. Haegeman2, E. G.J. Danchin3, H. S. Gaur4, J. Helder5, M. G.K Jones6, T. Kikuchi7, R. Manzanilla-Lo´pez8, J. E. Palomares-Rius7, W. M.L. Wesemael9,10 and R. N. Perry8,10. 1James Hutton Institute, Invergowrie, Dundee, DD2 5DA, UK; 2Institute for Agricultural and Fisheries Research (ILVO), Plant Sciences Unit, Caritasstraat 21, B-9090 Melle, Belgium; 3INRA UMR 1355, CNRS UMR 6243, UNSA, 400 route des Chappes, F-06903 Sophia-Antipolis, France; 4Sardar Vallabhbhai Patel University of Agriculture & Technology, Meerut-250 110, India; 5Laboratory of Nematology, Department of Plant Sciences, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands; 6Plant Biotechnology Research Group, School of Veterinary and Life Sciences, WA State Agricultural Biotechnology Centre, Murdoch University, Perth, Western Australia, WA 6150, Australia; 7Division of Parasitology, Department of Infectious Diseases, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan; 8Department of AgroEcology, Rothamsted Research, Harpenden Herts, AL5 2JQ, UK; 9 Institute for Agricultural and Fisheries Research (ILVO), Plant Sciences Unit - Crop protection, Burg. Van Gansberghelaan 96, B-9820 Merelbeke, Belgium; 10Ghent University, Department of Biology, Ledeganckstraat 35, B-9000 Ghent, Belgium. A survey of researchers working with plant-parasitic nematodes was undertaken in order to determine a ‘‘top 10’’ list of these pathogens based on scientific and economic importance. Any such list will not be definitive as economic importance will vary depending on the region of the world in which a researcher is based. However, care was taken to include researchers from as many parts of the world as possible when carrying out the survey. The top 10 list emerging from the survey was composed of (1) root-knot nematodes (Meloidogyne spp.), (2) cyst nematodes (Heterodera and Globodera spp.), (3) root lesion nematodes Pratylenchus spp., (4) the burrowing nematode Radopholus similis, (5) Ditylenchus dipsaci, (6) the pine wilt nematode Bursaphelenchus xylophilus, (7) the reniform nematode Rotylenchulus reniformis, (8) Xiphinema index (the only virus vector nematode to make the list), (9) Nacobbus aberrans, and (10) Aphelenchoides besseyi. The biology of each nematode (or nematode group) is reviewed briefly. A review article based on this survey has been published in Molecular Plant Pathology as part of the ‘‘Top 10 pathogens’’ series. CANDIDATE SECRETED PROTEINS AND EFFECTORS OF PRATYLENCHUS SPP. Jones1, M.G.K., J.C.H. Tan1 and J. Fosu-Nyarko2. 1Plant Biotechnology Research Group, School of Veterinary and Life Sciences, Western Australia, Australia; 2State Agricultural Biotechnology Centre, Murdoch University, Perth, Western Australia 6150, Australia. Root lesion nematodes (Pratylenchus species) are a group of economically important migratory endoparasitic plant pathogens that attack host roots of major crops. We have used Roche 454 deep sequencing of RNA extracted from mixed stages of Pratylenchus thornei and Pratylenchus zeae to generate reads, which following assembly and annotation, have provided transcriptomic data on sequences of genes expressed by these nematodes. Using this data to provide target sequences we have shown that both P. thornei and P. zeae are amenable to gene silencing (RNAi). By applying bioinformatics tools we have interrogated the transcriptome data for the presence of sequences that encode candidate secreted proteins, and from these about 30 candidate genes for such proteins have been identified. Some of these are common to sedentary endoparasitic nematodes, in particular genes encoding wall degrading enzymes, some found in the sedentary endoparasites are absent, and some candidate sequences have no known function. In silico comparative analysis of proteins and putative effectors present in the migratory and sedentary endoparasites will be provided as well as functional characterisation of some identified effectors using RNAi. Since Pratylenchus spp. do not reprogram cell identity and metabolism to form giant cells or syncytia, it would be expected that classes of effectors involved in those activities by root knot and cyst nematodes would be lacking, whereas the types of proteins secreted to aid entry and migration of nematodes through plant tissues, and to suppress host defences would be present in all endoparasitic nematodes. DETERMINATION OF RACES OF ROOT-KNOT NEMATODES (MELOIDOGYNE SPP.) IN TURKEY. Kacar, G. and I. H. Elekciog˘lu. University of Cxukurova, Faculty of Agriculture, Department of Plant Protection, 01360, Sarıcxam, Adana Turkey. Root-knot nematodes are one of the most economically important pest species with an extremely wide host range and high damage potential worldwide. Nearly a 100 Meloidogyne species have been described. Meloidogyne arenaria, Meloidogyne chitwoodi, Meloidogyne hapla, Meloidogyne incognita and Meloidogyne javanica are the most common and the most important Meloidogyne spp. in Turkey. In this study 50 Meloidogyne populations were collected from different region of Turkey. Races of Meloidogyne species were investigated by using differential host tests. The results indicated that M. incognita race 1, race 2, race 5, race 6; M. arenaria race 2, race 3; M. javanica race 1, race 5 and M. chitwoodi race 1 and race 2 are present in Turkey. Meloidogyne incognita race 5, M. javanica race 5 and M. chitwoodi race 1 and race 2 constitutes the first record of these races in Turkey. ECOTOXICITY OF SILVER NANOPARTICLES SYNTHESIZED FROM AZADIRACHTAINDICA AND CURCUMA LONGA ON ROOT-KNOT NEMATODE MELOIDOGYNE INCOGNITA. Kalaiselvi1, D., P. Sundararaj and S.L. Hafez2. 1Department of Zoology, Bharathiar University, Coimbatore, India; 2University of Idaho, Idaho, USA. Experiments were carried out to establish the efficacy of green synthesized silver nanoparticles by using two different tropical plants Azadirachtaindica and Curcuma longa on the mortality of second stage juveniles of the root-knot nematode

182 Journal of Nematology, Volume 46, No. 2, June 2014 Meloidogyne incognita. Characterization of nanoparticles was carried out using scanning electron microscopy (SEM), X-ray diffraction studies, FT-IR techniques, DLS techniques, zeta potential studies and the nano size was confirmed. Initially, the formation of silver nanoparticle was confirmed by UV spectroscopy after adding the respective plant extract as a reducing agent to the 0.01 M silver nitrate solution. Second stage larvae of M.incognita hatched out from the eggs of adult female nematodes cultured on tomato roots were exposed to respective plant extract concentrations of 10 mg L-1, 25 mg L-1, 50 mg L-1 and 100 mg L-1. There was a significant difference in the mortality of second stage juveniles as a result of their exposure to the hextracts of A. indica or C. longa. Differences were also observed in mortality induced by different concentrations of both plant extracts and there is a linear positive relationship between the larval mortality and the concentration of plant extract. UNRAVELING MI-1 IMMUNE RECEPTOR PERCEPTION AND SIGNALING IN TOMATO. Kaloshian, I. Department of Nematology, Institute for Integrative Genome Biology, University of California, Riverside, CA 92521. The tomato (Solanum lycopersicum; Sl) gene Mi-1 confers broad-spectrum resistance to three species of root-knot nematodes (Meloidogyne spp.) and three phloem feeding insects including potato aphid (Macrosiphum euphorbiae). It is not clear how Mi-1 is able to recognize avirulence effectors from these diverse groups of pests and whether the detection of nematode and insect pests involve similar recognition complexes. Mi-1 encodes a nucleotide-binding leucine-rich repeat immune receptor with no subcellular localization signal. Surprisingly, using confocal microscopy and biochemical fractionation, we found that Mi-1 is localized to three subcellular pools including the plasma membrane, cytoplasm and the nucleus. Using forward genetics, we identified Somatic Embryogenesis Receptor Kinase 1 (SERK1) to be required for Mi-1-mediated aphid resistance but not for root-knot nematode resistance. SERK1 is a transmembrane protein localized to the plasma membrane. Co-immunoprecipitation experiments in both Nicotiana benthamiana, transiently expressing Mi-1 and Sl-SERK1, and in 35S-Sl-SERK1-HA resistant tomato cultivar Motelle showed that Mi-1 and Sl-SERK1 are present in a complex in the microsomal fractions. Using reverse genetics, we have identified among others, members of WRKY transcription factors, known regulators of plant immunity inducible transcriptional network. Sl-WRKY72a and Sl-WRKY72b are upregulated by both root-knot nematode and aphid feeding and required for resistance to both pests. Interestingly, Arabidopsis thaliana WRKY72 seem to regulate a network of genes independent of the known defense hormone salicylic acid. Our work suggests similar Mi-1-mediated defence responses to Meloidogyne spp. and aphids but distinct Mi-1 recognition complexes to these organisms. DIVERSITY AND TRANSMISSION OF SOIL TRANSMITTED STRONGYLID NEMATODES BETWEEN HUMANS AND OTHER PRIMATES IN THE WILD. Kalousova1, B., H. Hasegawa2, D. Modry1,3,4, M. Kitagawa2, K. A. Shutt5, A. Todd6, I. Profousova1 and K. J. Petrzelkova1,4,7 8. 1Department of Pathology and Parasitology, Faculty of Veterinary Medicine, University of Veterinary and Pharmaceutical Sciences, Palackeho 1-3, 612 42 Brno, Czech Republic; 2Department of Biology, Oita University, 1-1 Idaigaoka, Hasama, Yufu, 879-5593 Oita, Japan; 3Central European Institute of Technology, University of Veterinary and Pharmaceutical Sciences, Palackeho 1-3, 612 42 Brno, Czech Republic; 4Biology Centre, Institute of Parasitology, Academy of Sciences of the Czech Republic, Branisovska 31, 370 05 Ceske Budejovice, Czech Republic; 5Department of Anthropology, Durham University, South Road, DH1 3LE Durham, United Kingdom; 6WWF, Dzanga Sangha Protected Areas, Avenue des Martyrs, BP 1053 Bangui, Central African Republic; 7Liberec Zoo, Masarykova 1347/31, 460 01 Liberec, Czech Republic; 8Institute of Vertebrate Biology, Academy of Sciences of the Czech Republic, Kvetna 8, 603 65 Brno, Czech Republic. Humans have historically shared habitat with nonhuman primates. However, the dynamics of human-primate interactions have recently changed dramatically and increased as a result of forest fragmentation, habitat encroachment and tourism. Little is known about the diversity and transmission of helminthes between humans and other primates and the presence of potential zoonotic strongylids should be viewed as a major concern for both public health and primate conservation. Generic diagnosis of strongylids is traditionally facilitated by morphological examination of L3 larvae; however, coprocultures for the development of larvae are not commonly performed in routine diagnostics of strongylids in primates. The difficulties in diagnosing hookworms combined with their clinical relevance have driven the need for molecular research/diagnostic approaches. We therefore studied the diversity and transmission of soil transmitted strongylid nematodes between humans and other primates inhabiting Dzanga Sangha Protected Areas, Central African Republic and Campo Ma’an National Park, Cameroon. We developed the filariform larvae of hookworms from faeces by using Harada-Mori filter-paper culture and identified them based on morphology. We sequenced and compared the first and second internal transcribed spacers (ITS1 and ITS-2) of nuclear ribosomal DNA and partial cytochrome c oxidase subunit 1 (cox1) gene of mtDNA obtained from larvae to identify the molecular type. Our data revealed an as yet undocumented diversity of hookworms (Ancylostomatoidea: Ancylostomatidae) in western lowland gorillas, chimpanzees, agile mangabeys and humans and provided evidence for the transmission of some parasite taxa among primates and humans as well as the discovery of a novel nematode species affecting humans.

Abstracts 183 THE ROLE OF JASMONIC ACID AND ETHYLENE DURING ATTRACTION AND EARLY DEFENCE REACTIONS TO HETERODERA SCHACHTII IN ARABIDOPSIS THALIANA. Kammerhofer1, N., P. Dobrev2, R. Vankova2, M.A.J. Regis1, J. Hofmann1 and K. Wieczorek1. 1Division of Plant Protection, Department of Crop Sciences, University of Natural Resources and Life Sciences, UFT Tulln, Konrad Lorenz Strasse. 24, 3430 Tulln, Austria; 2Academy of Sciences of the Czech Republic, Institute of Experimental Botany, Rozvojova´ 263, 165 02 Prague 6 – Lysolaje, Czech Republic. The root penetration, intracellular migration and syncytium initiation of Heterodera schachtii were suggested to induce plant defence responses in their hosts that were impaired at later stages of syncytium expansion and nematode development. The potential induction of plant defence during nematode invasion and migration is however poorly studied so far even though it may determine host plant susceptibility at that stage. Accordingly, phytohormones were shown to play significant roles in feeding site induction and nematode development, but their effect on nematode attraction and invasion during the first 24 hours was only studied to a minor extent. Thus the aim of the present work was to elucidate stress hormone based defence pathways which occur at the onset of infection. First, the potential role of salicylic acid, jasmonic acid and ethylene during nematode migration and syncytium induction was tested by analysing hormone levels and also the expression of frequently applied marker genes in infected compared to non-infected plants. Next, hormones or their inhibitors were applied to the shoots of Arabidopsis to be transported towards the roots. The successful translocation and induction of plant responses was controlled by analysing hormone and transcript levels as before. After this system was proved successful the potential effects of hormone application on the attraction, infection and development of H. schachtii was studied. Results suggest a pivotal role of jasmonic acid and ethylene in attraction as well as early infection process of H. schachtii. Findings of this work can contribute in understanding plant – nematode interactions and could be integrated in future resistance breeding programmes. BURSAPHELENCHUS SP. (TYLENCHOMORPHA: APHELENCHOIDIDAE) A PHORETIC ASSOCIATE OF XYLEBORINUS GRACILIS (COLEOPTERA: SCOLYTINAE) FROM AVOCADO IN FLORIDA. Kanzaki1, 2, N., R.M. GiblinDavis1, D. Carrillo3, L. Duncan3 and R. Gonzalez1. 1Fort Lauderdale Research and Education Center, University of Florida/IFAS, 3205 College Avenue, Davie, FL 33314, USA; 2Forest Pathology Laboratory, Forestry and Forest Products Research Institute, Tsukuba, Ibaraki 305-8687, Japan; 3Tropical Research and Education Center, University of Florida/IFAS, 18905 SW 280 Street, Homestead, FL 33301, USA. During an experimental host-plant survey for the invasive red bay ambrosia beetle, Xyleborus glabratus, in Homestead, Florida, a native species of ambrosia beetle, Xyloborinus gracilis was found to be associated with a Bursaphelenchus species. This nematode species was isolated from X. gracilis and cultured on Monilinia fructicola or Botrytis cinerea for further study and was determined to be new to science and a putative sister species to B. kiyoharai because of two derived characters in males; possession of a tail spike versus the typical bursal flap, and the apparent absence of the P1 ventral single papilla, both typically plesiomorphic characters for the genus. Additionally, B. kiyoharai is associated with X. serriatus suggesting that the host and microbiome associations that are shared between these two species that are carried by ambrosia beetles may have ecological and biological significance in their evolution and lineage radiation. Molecular phylogenetic analyses of the nearfull-length small subunit (SSU: 18S) and the D2/D3 expansion segments of the large subunit (LSU: 28S) confirmed that B. pen˜ai n. sp. is very closely related to B. kiyoharai which is a member of the B. fungivorus clade which includes B. thailandae and B. willibaldi. The newly-found Bursaphelenchus sp. and B. kiyoharai both share very similar overall spicule morphology with the B. fungivorus clade. The new species is described and can be typologically differentiated from B. kiyoharai by the position of the hemizonid and its different geographical and host associations. DIVERSIFICATION OF TERMITE-ASSOCIATED NEMATODES IN RELATION TO EVOLUTIONARY PATTERNS OF DIVERSITY. Kanzaki1, 2, N., R.M. Giblin-Davis2 and K.A. Davies3. 1Forestry & Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687 Japan; 2Fort Lauderdale Research & Education Center, University of Florida/ IFAS 3205 College Avenue, Davie, Florida 33314 USA; 3Centre for Evolutionary Biology and Biodiversity, School of Agriculture, Food and Wine, The University of Adelaide, Waite Campus, PMB 1, Glen Osmond, South Australia 5064, Australia. Entomophilic nematodes are important subjects as indicators of biodiversity, potential genetic resources, and potential biological control agents. The nematode fauna associated with termites and their relatives were examined as predictors of nematode diversity. The termites were collected in the forests in the American and East Asian tropics and subtropics, and dissected to determine the nematode species associated with each termite species. In addition, as relatives of termites, some roach and woodroach species were also collected and examined for their associated nematodes. Information from the literature was also examined to complement deficiencies from our survey work. The termites and related insects were separated into four groups according to the pattern of associated nematodes. Roaches, phylogenetically the most basal group, were associated with external and internal parasites (e.g., Thelastomatids); woodroaches, intermediate between roaches and termites, were not found to be associated with any nematodes; lower termites (Kalotermitidae) were associated with some phoretic Poikilolaimus spp. and Halicephalobus spp. which were isolated only from kalotermitids; and the other more derived termite groups were associated with various phoretic rhabditids, diplogastrids and aphelenchoidids and a few

184 Journal of Nematology, Volume 46, No. 2, June 2014 parasitic species. Comparing the associated nematode assemblages and phylogenetic relationships of their insect hosts, the nematode association is hypothesized to have been lost by the common ancestor of woodroach/termites, i.e., the insects might have escaped from nematode associations, and then re-obtained and expanded the associated nematode fauna as the termite lineages radiated. DIVERSITY OF FIG-ASSOCIATED NEMATODES AS A MODEL SYSTEM FOR STUDYING BIOGEOGRAPHY, HOST SWITCHING AND COLONIZATION OF NEW HABITATS. Kanzaki1, 2, N., R. Giblin-Davis2 and K.A. Davies3. 1 Forestry & Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687 Japan; 2Fort Lauderdale Research & Education Center, University of Florida/IFAS 3205 College Avenue, Davie, Florida 33314 USA; 3Centre for Evolutionary Biology and Biodiversity, School of Agriculture, Food and Wine, The University of Adelaide, Waite Campus, PMB 1, Glen Osmond, South Australia 5064, Australia. Trees belonging to the genus Ficus, comprise a very interesting plant group. Fig trees are associated with their mutualistic pollinator fig wasps, and the patterns of their host associations with pollinator wasps, inquilines and nematodes are serving as an excellent model system for studying co-speciation and host switching. Several different lineages of nematodes are involved in this interaction and promise high species diversity. However with over 700 species of Ficus worldwide, the information about the diversity and biological interactions of this group and other fig-associated nematodes has not been sufficiently studied. Therefore, the authors have conducted several field surveys of fig-associated nematodes in Central America, Australia and the East Asian tropics and subtropics, as well as some African specimens deposited by previous investigators. During these surveys, many new species/genera have been observed, and new species (e.g., Parasitodiplogaster spp. and Schistonchus spp.), and a new genus (Teratodiplogaster) have been described. Furthermore, by comparing the nematode fauna from figs from the surveyed areas, region-specific nematode lineages were discovered. Also, several different confamilial genera (lineages) were confirmed with evidence of morphological and life history trait convergence, e.g., at least four clades of aphelenchoidids have converged around the general morphology and trait of plantparasitism of ‘‘Schistonchus’’, and three lineages of diplogastrids have converged around the traits of parasitism/necromeny of the ‘‘Parasitodiplogaster’’. The authors will summarize current knowledge of the diversity of fig-associated nematodes from field surveys. PARASITODIPLOGASTER SPECIES ASSOCIATED WITH PHARMACOSYCEA FIGS IN PANAMA. Kanzaki1, 2, N., R.M. Giblin-Davis1, W. Ye1,3, E.A. Herre5 and B.J. Center1. 1Fort Lauderdale Research and Education Center, University of Florida, 3205 College Ave, Davie, Florida 33314–7799, USA; 2Forest Pathology Laboratory, Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687 Japan; 3Present address: Nematode Assay Section, Agronomic Division, North Carolina Department of Agriculture & Consumer Services, 4300 Reedy Creek Road, Raleigh, NorthCarolina 27607, USA; 4Smithsonian Tropical Research Institute, Balboa, Ancon, Republic of Panama. Parasitodiplogaster species that are associated with figs in the Ficus subsection Pharmacosycea, i.e., Ficus maxima, Ficus yoponensis, Ficus insipida and Ficus glabrata were examined based on morphological characters and molecular sequences. Some nematodes isolated from the fig sycones were observed and recorded for their morphotype, and then digested for their DNA, while others were fixed in formalin-glycerin for additional morphological study. High resolution microscopic observation of the morphological materials yielded five morphotypes including two nominal species, Parasitodiplogaster maxinema and Parasitodiplogaster pharmaconema, and these morphotypes were distinguished from each other by male tail characters. In contrast, five and six genotypes were recognized by D2/D3 expansion segments of the large subunit (LSU) and near-full-length sequences of small subunit (SSU) ribosomal RNA genes, respectively. Except for the two nominal species, the genotypes and morphotypes were not clearly correlated because of limitations in the microscopic resolution in the initial morphotyping. Although the morphotypes and genotypes were not clearly paired, Pharmacosycea-associated Parasitodiplogaster species which are tentatively referred to as the ‘‘P. maxinema group’’ formed a monophyletic clade in both D2/D3 LSU and SSU analyses, and are morphologically characterized by their stomatal morphology, i.e., a tube-shaped stoma with two stick-like teeth and male tail morphology, presence of nine paired papillae and relatively slender spicule. A BURSAPHELENCHUS SPECIES HAS BECOME A PLANT-PARASITE IN FIGS AND CONVERGED IN APPEARANCE WITH SCHISTONCHUS. Kanzaki1, N., R. Tanaka2, R.M. Giblin-Davis3 and K.A. Davies4. 1Forestry & Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687 Japan; 2Division of Parasitology, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan; 3Fort Lauderdale Research & Education Center, University of Florida/IFAS 3205 College Avenue, Davie, Florida 33314, USA; 4Centre for Evolutionary Biology and Biodiversity, School of Agriculture, Food and Wine, The University of Adelaide, Waite Campus, PMB 1, Glen Osmond, South Australia 5064, Australia. Some morphological and biological characters of an undescribed Bursaphelenchus species are reported. The species was found from syconia of a Japanese native fig, Ficus variegata in Ishigaki Island, Okinawa, Japan. Although the Bursaphelenchus sp. shares an important typological character, male spicule possessing a strongly recurved condylus, with the

Abstracts 185 Bursaphelenchus eremus group and the Bursaphelenchus leoni groupof the genus, it was inferred to be monophyletic with the Bursaphelenchus fungivorus group based on the molecular sequences of ribosomal RNA genes. Further, the stylet morphology of the species is very unique in the genus, i.e., it is very long, thick and possessing long (ca 50% of total length) conus and large basal swellings and well-developed pharyngeal glands. The morphology is rather similar to that of the Schistonchus paraphyletic assemblage of the fig-floret parasitic group. Thus, the very unique stylet morphology found in the species appears to be an extreme example of convergent evolution in the nematode family, Aphelenchoididae inside figs. Whereas the other characters shared by the new species and its close relatives, i.e., lack of ventral P1 male genital papilla, female vulval flap, and papilla-shaped P4 genital papillae in males, corroborate the molecular phylogenetic inference. The unique biological character of obligate plant parasitism and highly derived appearance of the ingestive organs of Bursaphelenchus sp. expands our knowledge of the potential morphological, physiological and developmental plasticity of the genus Bursaphelenchus. STRUCTURE FUNCTION OF SOIL NEMATODE COMMUNITIES AFTER APPLE ORCHARD FLOOR MANIPULATIONS. Kapp1, C., S.G. Storey2 and A.P. Malan1. 1Department of Conservation Ecology and Entomology, Faculty of AgriSciences, University of Stellenbosch, Private Bag X1, Matieland, 7602, South Africa; 2Nemlab Diagnostic Laboratories, c/o R44, Anyswortelrug Road, Klapmuts, South Africa. As an essential non-renewable resource in all land-based ecosystems, it is imperative to develop methods in which to maintain the health, biological productivity and sustainability of our agricultural soil. The importance of this is indeterminate when considering the increase in the global human population and the consequent proliferation on the demand for food production. This has prompted research to develop uses of different biological indicators of soil health. Nematodes are one of the most abundant groups of Metazoa and form an integral part of the soil food web at several trophic levels, responding rapidly to any environmental changes. Characterization according to functional guild can impart valuable insight into the condition of the soil food web. A study was undertaken to determine the biodiversity and functionality of soil nematodes associated with the effects of various orchard floor manipulations in an apple orchard in the Western Cape province of South Africa. Samples were collected from the ridge of the tree row of each treatment plot. The nematodes present within each treatment were identified to family level, in addition to being quantified. Indices for the determination of ecosystem function were calculated for each treatment and a faunal analysis was done. Results will be discussed in terms of soil enrichment and structure as well as the increase or decrease in the levels of maturity over time. THE EFFECT OF DIFFERENT TOMATO GENOTYPE ON ROOT-KNOT NEMATODE-FUSARIUM WILT COMPLEX IN COASTAL KENYA. Kariuki1, P.M., F. Kariuki1, G. M. Kariuki2 and D. L. Coyne3. 1Department of Plant and Microbial Sciences, Kenyatta University, P.O. Box 43844-00100, Nairobi, Kenya; 2Department of Agricultural Science and Technology, Kenyatta University, Nairobi, Kenya; 3International Institute of Tropical Agriculture (IITA), c/o icipe, Kasarani, P.O. Box 30772-00100, Nairobi, Kenya. Tomato is one of the most popular vegetable crops grown worldwide, owing to its nutritive value and diversified use. The disease complex involving root-knot nematodes (Meloidogyne spp.) and Fusarium oxysporum is a common occurrence on tomato, and especially along the coast in Kenya. The effect of the disease complex was studied under screen house conditions in pots using five cultivars with varying levels of resistance to the two pathogens. The effects of both pathogens individually and sequentially were assessed on the five cultivars. Isolates of root-knot nematodes and F. oxysporum singly and concomitant caused significant reduction in both shoot and root growth on tomatoes over the un-inoculated control. The reduction was however, greater when both pathogens were inoculated together, as opposed to when root-knot nematodes were inoculated singly. Fusarium wilt inoculation alone had the least reduction in the growth parameters. Galling index, egg mass index and the number of juveniles in the soil reduced significantly while the Fusarium wilt index increased on sequential inoculation of the pathogens. Among the five cultivars tested tomato cv. Kilele showed the least galling index, egg mass index, juveniles’ population in the soil as well as the least Fusarium wilt index. It was followed by cv. Bt Okitu 101 and cv. T- R-3034-3-10-N-UG respectively. The most susceptible cultivars were Cal J and Hawai 7996. The study concludes that both pathogens interacts and are virulent against the tomato cultivars. SAFE AND EFFECTIVE NEMATODE AND OTHER PESTS MANAGEMENT STRATEGIES TO STRENGTHEN THE TOMATO VALUE CHAIN IN COASTAL KENYA. Kariuki1, G.M., L. Muriuki1, A.K. Thuo, J.W. Kibunja2, P.M. Kariuki,2 and D.L. Coyne3. 1Department of Agricultural Science and Technology, Kenyatta University, P.O. Box 4384400100, Nairobi, Kenya; 2Department of Plant and Microbial Sciences, Kenyatta University, Nairobi, Kenya; 3International Institute of Tropical Agriculture (IITA), c/o ICIPE, Kasarani, P.O. Box 30772-00100, Nairobi, Kenya. With rising population and urbanization in the developing world, peri-urban agricultural systems become more intensely cultivated and plagued by pests and diseases. Since individual pest control measures each have limitations, the integration of several tactics is often necessary. When available, host resistance should be incorporated as a first line defence mechanism to reduce pesticide reliance. Unfortunately, vegetable crops, particularly tomato are usually susceptible to a range of pests,

186 Journal of Nematology, Volume 46, No. 2, June 2014 including root-knot nematodes (Meloidogyne spp.), bacterial wilt, fusarium wilt and viruses. Resistance needs to be locally appropriate to species or strain variability and durable under multiple pest attack. To address this problem the current study is being conducted along the East African coast to: 1) determine the distribution and relative occurrence of virulent species, races and/or strains of begomoviruses, Ralstonia solanacearum and Meloidogyne spp. and fusarium wilt on tomato, 2) select tomato germplasm with multiple resistance against begomoviruses, R. solanacearum and Meloidogyne spp. and 3) explore and develop novel bio-pesticides for ttomato cultivation for action against begomoviruses (or vectors), R. solanacearum and Meloidogyne spp. Molecular techniques are being employed to attain accurate pest diagnostics and marker assisted identification of resistance and host interactions. The use of endophytic fungi Trichoderma spp. alongside moderately resistant varieties have been found to be effective against Meloidogyne spp. on tomato and also against disease complexes involving root-knot nematodes, Bacterial wilt and fusarium wilt. The beneficiaries of this study will be the small-scale farmers in coastal East Africa. NIMITZTM: A PARADIGM CHANGE IN NEMATODE CONTROL. Karmon, D. ADAMA Agricultural Solutions, Golan Street, Airport City 7019900, Israel. Worldwide damage to crops by nematodes is currently estimated to be greater than US$ 120 billion per annum. Existing measures taken by farmers to cope with nematode damage vary and include crop rotation, resistant varieties, sanitation, ‘‘suppressive crops’’, organic substances and chemical PPP’s (?). The availability of PPP’s has declined over recent years. Methyl bromide has been phased out while other soil fumigants like metham sodium & 1,3-dichloropropen are under increasing regulatory pressure. Specific nematicides belong mostly to organophosphate (OP’s) and carbamate groups are also under increasingly heavy regulatory pressure in many countries. ADAMA is currently in the process of registering NIMITZTM, which contains the active ingredient fluensulfone. Fluensulfone belongs to novel chemical class - Fluoroalkenyl (-thioether). NIMITZTM is a true nematicide, with irreversible activity, causing the death of nematodes, as compared to OP’s and carbamates that only have reversible nematostatic activity. NIMITZTM‘s novel mode of action affects the different developmental stages of nematodes in a variety of ways. NIMITZTM has toxicity and ecotoxicity profiles which are much better than the existing nematicides. NIMITZTM registration is expected shortly in several leading countries with the main target crops being the families of Solanaceae and Cucurbitaceae. Additional crops will be added to the registration at a later stage.Global field trials program proved the excellent efficacy of NIMITZTM in controlling root knot nematodes (Meloidogyne spp.) as well as a variety of other economically important species. Trial results are presented here. INVESTIGATING THE NEUROPHARMACOLOGICAL REGULATION OF STYLET BEHAVIOUR IN GLOBODERA PALLIDA. Kearn1, J., A. Dorey1, P. Urwin2, C. Lilley2, V. O’Connor1, L. Holden-Dye1. 1University of Southampton, Centre for Biological Sciences, Bassett Crescent East, University of Southampton, Southampton, SO17 1BJ, UK; 2University of Leeds, Centre for Plant Sciences, School of Biology, University of Leeds, Leeds, LS2 9JT, UK. All plant parasitic nematodes possess a stylet; a, protrusible mouth spear. The stylet is essential in the life cycle as it is required for the emergence of second stage juvenile (J2) from the egg, the penetration of host roots,or migration through the host and feeding. The thrusting of the stylet when the nematode is in contact with the host root mechanically pierces or separates the host cells and enables the first step in the life cycle. Due to the importance of the stylet in plant parasitic nematodes, its behaviour presents an attractive target for agrochemical control of plant parasitic nematodes. The neurobiology of this behaviour is however relatively uncharacterised. Here, we have used the potato cyst nematode Globodera pallida to study the effects of neurotransmitters, neuromodulators and pharmacological inhibitors on stylet thrusting and movement to gain a greater insight into how the behaviour of this crucial structure is regulated and controlled. Our data suggest that there are similarities between the neuropharmacological regulation of stylet behaviour in plant parasitic nematodes and the regulation of feeding in other nematode species. Using our data we have begun to develop a putative model of the neurotransmitters and neuromodulators involved in this behaviour and we look to develop this model further and to study the pharmacology of the stylet in other plant parasitic nematodes. MODE OF ACTION STUDIES ON FLUENSULFONE, A NEW NEMATICIDE. Kearn, J., E. Ludlow, V. O’Connor, L. Holden-Dye. University of Southampton, Centre for Biological Sciences, Bassett Crescent East, University of Southampton, Southampton, SO17 1BJ. Plant parasitic nematodes present an unacceptable burden to global food security, with nematode infection directly responsible for $125 billion of yield losses every year. The nematode problem is exacerbated by a lack of nematicides, as many chemicals have had their use restricted due to concerns over their non-selective toxicity. Fluensulfone is a new nematicide that is selectively toxic towards nematodes and thus has reduced potential for environmental damage. The mode of action of fluensulfone is however not known. Here, we have performed mode of action studies, utilising the model organism Caenorhabditis elegans, and the cyst nematode Globodera pallida. Caenorhabditis elegans has been previously used as a model organism in mechanism of action studies on anthelmintic compounds, due to the wealth of knowledge concerning its neuropharmacology and its experimental tractability. Globodera pallida was used to determine the relevance

Abstracts 187 of these observations to the activity of fluensulfone against plant parasitic nematodes. We have found that fluensulfone has complex and pleiotropic actions on C. elegans, including effects on locomotion, development, egg laying, egg hatching and feeding behaviour. These observations and analysis of C. elegans mutants suggest a mode of action that is distinct from other nematicides and anthelmintics. The effects of fluensulfone on the feeding behaviour of C. elegans and G. pallida are complex, and suggest an interaction with neural signalling pathways, a hypothesis that is supported by further pharmacological studies. Further studies on C. elegans and G. pallida are in progress to delineate the molecular mechanism(s) underpinning these responses to fluensulfone. INVESTIGATING STEINERNEMA FELTIAE AS A SUITABLE INDICATOR ORGANISM FOR HEAVY METAL TOXICITY ASSAYS. Kelly, E. and T. Kakouli-Duarte. Molecular Ecology and Nematode Research Group, EnviroCORE, Department of Science and Health, Institute of Technology Carlow, Kilkenny Road, Carlow, Ireland. Heavy metals occur in the environment naturally, but through mining and industrialization they can accumulate to toxic levels. This work explores the suitability of Steinernema feltiae as a sentinel species for heavy metal pollution. Nematodes are widely used as environmental indicators. Steinernema feltiae is a widespread soil-dwelling nematode, normally parasitizing insects to complete its life cycle, and possessing an age-synchronized developmental stage particularly suitable for toxicity testing. Here we report on laboratory based toxicity assays where third stage juveniles of the nematode were used as biomarkers for the effects of cadmium, lead and zinc. Nematodes were exposed to specific levels of heavy metals for different time periods. Cadmium and lead had a most profound negative effect on nematode life span in a time dependent manner. Experiments were also conducted in sand columns and on agar plates, in order to observe the effects of heavy metals on nematode motility and attraction to insect hosts. The host seeking ability of the nematodes was not negatively affected by heavy metals in sand, but their movement was compromised by heavy metals on agar so they did not respond directionally to the hosts. When nematode development in larvae of the moth Galleria mellonella was investigated, lead appeared to delay J1 and J2 formation and cadmium was detrimental to fecundity and J3 emergence. These results constitute a first report of metal toxicity end points that may be developed to use this nematode as a sentinel for heavy metal pollution. VARIABILITY IN RESPONSE OF WHEAT AND AEGILOPS GENOTYPES TO MEDITERRANEAN CEREAL CYSTS NEMATODE (HETERODERA LATIPONS). Kherfan1, W.I., F. C. Ogbonnaya2,3, L. Al-Banna1. 1Department of Plant Protection, University of Jordan: University of Jordan Amman-11942-Jordan; 2International Center for Agricultural Research in the Dry Areas (ICARDA), PO Box 5466, Aleppo, Syria; 3The Grains Research & Development Corporation, PO Box 5367, Kingston, ACT 2604, Australia. The Mediterranean cereal cyst nematode, Heterodera latipons, is one of the major pests that affect wheat production in Jordan and other countries in the Mediterranean region. To successfully deploy genetic resistance as a control measure, the reaction of bread wheat cultivars, synthetic hexaploids of wheat and Aegilops genotypes to a Jordanian isolate of the Mediterranean cereal cysts nematode, H. latipons were investigated. Furthermore, the impact of different sources of genes for resistance to this cyst nematode was compared. The examination of stained roots harvested after two, three and four weeks of inoculation from all genotypes, showed that the penetration of J2s occurred in all the genotypes. However, the total number of the penetrated J2s varied among lines and were less in Aegilops genotypes. The presence of different developmental stages of the nematode were higher in number and developed faster in the susceptible and moderately susceptible wheat genotypes than in the resistant ones. Interestingly, some Aegilops genotypes did not support the development of J2s to further stages. The development of J2s to J3s and J4s were less in wheat genotypes carrying the Cre3 resistant gene than those genotypes carrying Cre1 and Cre8 resistant genes. These findings suggest that different pre- and post-penetration resistance mechanisms may be associated with the differential response of the genotypes. Results from this study confirmed that the previously identified cereal cyst nematode resistance genes (Cre1, Cre3 and Cre8) confer resistance to the Jordanian pathotype of H. latipons used in this study. IN VITRO EVALUATION OF EXTRACT AND FRACTIONS OF MAERUA ANGOLENSIS AND TABERNAEMONTANA ELEGANS ON MOTILITY AND HATCH INHIBITION OF MELOIDOGYNE INCOGNITA RACE 2. Khosa1, C., A. Mc Donald2, M. Daneel1, D. De Waele3, G. Fouche4, M. Tselanyane4 and F. Calitz5. 1ARC-Institute of Tropical and Subtropical Crops, Private Bag X11208, Nelspruit 1200, South Africa; 2North-West University, School of Environmental Sciences and Development, Plant Protection, Private Bag X6001, Potchefstroom, 2520, South Africa; 3Laboratory for Tropical Crop Improvement; Catholic University of Leuven, Kasteelpark Arenberg 13, 3001, Heverlee, Belgium; 4Biosciences, Council of Scientific and Industrial Research, P.O. Box 395, Pretoria, 0001, South Africa; 5ARC-Central Office, P.O. Box 8783, Pretoria, 0001, South Africa. Plant-derived metabolites have attracted the most attention of all prospective alternatives to pesticides, either as plant extracts, formulated phytochemicals or as organic amendments to soil. Several plant species have been reported to contain metabolites with nematoxic activity and there is continuous effort in discovering new plant sources with such characteristics. Plant extract and fractions of Maerua angolensis and Tabernaemontana elegans were tested for nematicidal activity in terms

188 Journal of Nematology, Volume 46, No. 2, June 2014 of nematode second-stage juvenile (J2) motility and egg hatch inhibition at 10 different concentrations (0.1-1.0 mg ml-1) each in 96-well test plates. Four independent trials for each plant extract concentrate were arranged in a randomized-complete block design, with the eight treatments in each test replicated four times each. In vitro bioassay studies confirmed that extract and fractions of varying polarity of both plant products M. angolensis and T. elegans might be toxic to J2 stages of the rootknot nematode Meloidogyne incognita. All extract/fractions of M. angolensis tested caused immobility of J2, whereas only three extract/fractions of T. elegans affected mobility of J2 adversely. Methanol/dichloromethane (B) and evaporated dichloromethane (D) from both plants were the only extract/fractions with potent egg hatch inhibition activity at concentration levels 0.9 to 1.0 mg ml-1 after 21 days of exposure. This information provided further insight into the usefulness of these materials in root-knot nematode control in small-scale farming communities, domestic gardens and commercial farming. THE USE OF TRICHODERMA SPP. ALONGSIDE RESISTANT TOMATO VARIETIES FOR ROOT-KNOT NEMATODE MANAGEMENT UNDER FIELD CONDITIONS IN COASTAL KENYA. Kibunja1, J.W., J.K. Birgen1, G.M. Kariuki2 and D.L. Coyne3. 1Department of Plant and Microbial Sciences, Kenyatta University, P.O. Box 43844-00100, Nairobi, Kenya; 2Department of Agricultural Science and Technology, Kenyatta University, Nairobi, Kenya; 3International Institute of Tropical Agriculture (IITA), c/o icipe, Kasarani, P.O. Box 30772-00100, Nairobi, Kenya. The endophytic activity of Trichoderma spp. strains were assessed for their potential biological control against root-knot nematode (Meloidogyne spp.) on both resistant and susceptible tomato varieties in the humid tropical conditions of coastal Kenya. Screenhouse and field experiments were carried out at Pwani University. Trichoderma spp. was evaluated alongside tomato cv. Hawaii, Okitu, Nug (AVRDC) and Kilele and Cal J (local varieties). One week after sowing seedlings were drenched with a suspension of Trichoderma spp. which had been obtained in the course of this research and not yet characterized and T. harzianum T-22 (sold under trade name Trianum-P in Kenya) in the nursery bed. Seedlings were transplanted three weeks after planting and inoculated with 500 J2s. The trial was laid out in randomized complete block design (RCBD) replicated four times. Experiments were terminated at 60 days (greenhouse) and 90 days (field). There was no significant difference between treated plants and the control on growth parameters. However there were significant differences for galling and egg mass index, J2 density, and reproductive index between the treatments and the control. The study revealed a considerable significant variation in response against Meloidogyne spp. infection among the germplasm tested when used alongside the Trichoderma spp. Interaction was observed between the varieties and the treatments. The cultivar Cal J was highly susceptible to RKN followed by Hawaii. The cv. Kilele which is commonly used in Kenya had less galling and egg mass index with a low reproduction rate indicating its ability to suppress the adult female reproduction. The Trichoderma spp. used significantly suppressed the disease severity caused by root-knot nematodes. The results of this study points to a combined use of locally available Trichoderma spp. alongside resistant varieties in the humid tropics where temperatures are relatively high as part of integrated pest management against Meloidogyne spp. CHARACTERIZATION OF NEMATODE PESTS OF ENSET (ENSETE VETRICOSUM WELW. CHEESMAN) AND THEIR MANAGEMENT. Kidane1, S., D. Coyne1 and S. Haukeland2. 1International Institute of Tropical Agriculture, P.O. Box 30772-00100, Nairobi, Kenya; 2Bioforsk, Plant Health and Plant Protection, Norway. Enset (Ensete vetricosum Welw. Cheesman) is an important starch staple crop, cultivated primarily in southern and south west Ethiopia. Related to the banana family, enset is similarly infected by plant-parasitic nematodes. From previous survey studies Pratylenchus goodeyi appears to be the dominant nematode pest, which is believed to contribute to reduced productivity of enset. However, while surveys have demonstrated high P. goodeyi infection levels, there is relatively scant information on how damaging the nematode is to enset production. There is also little information on the variability of the nematode pest in terms of levels of pathogenicity on enset and if so, how this may relate to variability in climate and temperature zones under which enset is grown. Our study is being undertaken to assess the possible damage of nematode pests, with emphasis on P. goodeyi, and in relation to the presence of other diseases and how climate and agro ecology may affect this. Specifics of the study are presented. EFFICACY OF BIOACT WG (PAECILOMYCES LILACINUS STRAIN 251) TO CONTROL ROOT-KNOT NEMATODES (MELOIDOGYNE SPP.) IN HIGH VALUE CROPS. Kiewnick, S. Agroscope Competence Center for Nematology, Institute for Plant Production Sciences, IPS Schloss 1, CH-8820 Waedenswil, Switzerland. Root-knot nematodes (Meloidogyne spp. cause severe losses in high value crop production systems. As plant resistance or chemical control options are either not available or being phased out, a management strategy based on the biological bionematicide BioAct WG (active ingredient Paecilomyces lilacinus strain 251) was tested in on-farm and semi-commercial trials for efficacy in reducing root-knot nematode damage and nematode population densities in soil. It could be demonstrated that a pre-planting soil treatment followed by repeated post-plant applications significantly reduced the root damage in commercial greenhouse production systems. In contrast to chemical soil disinfestation, root-knot nematode soil population densities remained at low levels after application of BioAct WG. In greenhouses where Meloidogyne enterolobii had caused up to 50% yield loss, application of BioAct WG reduced the damage to a minimum and ensured high yield levels. Studies on

Abstracts 189 the interaction between P. lilacinus strain 251 and M. enterolobii revealed particularly high levels of egg parasitism when compared to other RKN species which explains the high efficacy in controlling this extremely aggressive species. In addition, semi commercial trials revealed a significant reduction in root damage and increase in tomato fruit yield following a BioAct WG application. The combination of a BioAct WG application with measures to reduce the nematode population densities in soil further enhanced efficacy of this root-knot nematode management strategy. DEVELOPMENT OF A STRAIN SPECIFIC SYBR GREEN I BASED REAL-TIME PCR ASSAY FOR ENVIRONMENTAL MONITORING OF PAECILOMYCES LILACINUS STRAIN 251. Kiewnick, S., S. Wolf, A. Lehmann and J. Frey. Agroscope, Institute for Plant Production Sciences, IPS, Schloss 1, CH-8820 Waedenswil, Switzerland. Monitoring of nematode antagonists after application allows for better understanding of the tri-trophic interactions between target nematode species, the host plant and the fungal biocontrol agent. Previous molecular assays were based on the internal transcribed spacer (ITS) region of the fungal genome, but only specific to species level. Therefore, other gene regions were tested to develop molecular assays for the detection and identification of Paecilomyces lilacinus strain 251 (PL251). Ribosomal RNA genes were evaluated for containing strain specific single nucleotide polymorphisms (SNPs), but, comparison of all sequence data obtained for individual P. lilacinus strains revealed no strain specific SNPs. Furthermore, a RAPD-analysis was conducted, but no specific fragments were detected. Therefore, a new approach using GS Junior 454 sequencing technology was used. 22 RAPD primers were used to generate amplicons for different P. lilacinus strains. All amplicons were sequenced and a total of 34 MB sequence data generated. Contiguous sequence of strain 251 was concatenated with addition of 100bp N linkers, to facilitate read mapping. Raw reads of the other libraries were then mapped against reference PL251. The consensus sequence of the other strains was aligned (MUSCLE, Geneious Pro v. 6.1.3). For the multiple alignment 8 contiguous sequences were found to be singleton. For these sequences, primer pairs were generated with Geneious Pro v. 6.1.3 and 2 combinations were suitable for use in SYBR Green I assays. Initial testing demonstrated a very high specificity for PL251 and no cross reaction with other closely related strains or biocontrol fungi. NEW CYST NEMATODES FROM SOUTH AFRICA: A VALUABLE RESOURCE FOR THE STUDY OF THE EVOLUTION AND BIOGEOGRAPHY OF THE GROUP. Knoetze1,2, R., A. Swart3 and L.R. Tiedt4. 1Department of Conservation Ecology and Entomology, Department of AgriSciences, University of Stellenbosch, Private Bag X1, Matieland 7602, South Africa; 2Directorate Inspection Services, Department of Agriculture, Forestry and Fisheries, Private Bag X5015, Stellenbosch 7599, South Africa; 3National Collection of Nematodes, Biosystematics Division, ARC-Plant Protection Research Institute, Private Bag X134, Queenswood 0121, South Africa; 4Laboratory for Electron Microscopy, North-West University, Potchefstroom Campus, Potchefstroom 2520, South Africa. The theory that South America or Africa is a centre of origin for Globodera prompted the initiation of a survey for the presence of indigenous cyst nematodes in The Cape Floristic Region of South Africa. The ITS-rDNA regions of four indigenous isolates of cyst nematodes, discovered in the survey, were amplified by PCR, sequenced and aligned with selected sequences from the Heteroderidae. Phylogenetic analyses of the aligned sequences were conducted using the Maximum Parsimony (MP) method. The morphology of the cysts was also studied using both light microscopy (LM) and scanning electron microscopy (SEM). Phylogenetic analysis of the sequences from three of the isolates, places them in a clade of Globodera species that parasitise non-solanaceous plants, confirming their membership of the genus. LM and SEM studies confirmed that these specimens can be classified as Globodera on the basis of their morphological characteristics. Morphometric data and phylogenetic analyses indicate that they are unique species, previously undescribed. Another isolate forms a monophyletic group with Heterodera species in the MP tree, but the morphology of these specimens is more consistent with that of Betulodera. However, unique labial patterns, as well as the presence of a vulva and functional gonads in some of the juvenile specimens, set them apart from both Betulodera and Heterodera. The discovery of new Globodera species in Southern Africa supports the theory of a Gondwanaland origin of the genus and the theory that divergence of the two main Globodera lineages might have occurred subsequent to the break-up of Africa and South America. NEMATODE MANAGEMENT IN FLORIDA VEGETABLE AND ORNAMENTAL PRODUCTION. Kokalis-Burelle1, N., F.B. Iriarte2, D.M. Butler3, J.C. Hong1 and E.N. Rosskopf1. 1USDA-ARS, US Horticultural Research Lab, Ft. Pierce, Florida, USA; 2Department of Plant Pathology, Kansas State University, Manhattan, Kansas, USA; 3Department of Plant Sciences, University of Tennessee, Knoxville, Tennessee, USA. Restrictions on fumigants necessitate development of new options for plant-parasitic nematode management in vegetable and ornamental crop production. USDA-ARS researchers are developing new nematode control tactics, including low-risk chemicals (SPK), steam, and anaerobic soil disinfestation (ASD). These approaches are integrated into site-specific management plans. SPK is a unique formulation of organic acids with efficacy against root-knot nematodes (Meloidogyne spp.), as well as soilborne plant pathogenic fungi and bacteria. Steam is effective for broad-spectrum soilborne pest control, however, logistics and expense of application may limit adoption. ASD uses a combination of solarization, organic amendments, and soil saturation to raise soil temperatures, stimulate microbial activity, and create anaerobic conditions in

190 Journal of Nematology, Volume 46, No. 2, June 2014 soil covered with polyethylene mulch. In a double-crop pepper-eggplant trial, Meloidogyne incognita numbers were low through the first season following ASD. By the second eggplant crop, solarization (no amendments or water) averaged > 200 nematodes/100 cm3 soil compared to 10/100 cm3 in ASD treatments with amendments and irrigated with 5-10 cm of water. While integrated nematode control tactics are preferred, stand-alone approaches are also being investigated including vegetable grafting, and biocontrol with Pasteuria penetrans. Grafting for root-knot nematode control in tomato has been successful; however melon grafting did not improve root-knot nematode control, although yield of some rootstock scion combinations improved compared to non-grafted plants. Greenhouse and microplot trials on P. penetrans are ongoing but preliminary results indicate good potential for field application. IDENTIFICATION AND DIVERSITY OF THE ROOT-KNOT NEMATODES AFFECTING YAM (DIOSCOREA SPP.), IN NIGERIA. Kolombia1,2, Y.A., G. Karssen2,3, N. Viaene2,4, P Lava Kumar1, D. Coyne5 and W. Bert2. 1International Institute of Tropical Agriculture (IITA), PMB 5320, Oyo Road, Ibadan, Nigeria; 2Nematology Research Unit, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Gent Belgium; 3National Plant Protection Organization, 6706 EA Wageningen, The Netherlands; 4Institute for Agricultural and Fisheries Research (ILVO), 9820 Merelbeke, Belgium; 5IITATanzania, Plot 25, Mikocheni, P. O. Box 34441, Dar es Salaam, Tanzania. Root-knot nematodes, Meloidogyne spp. represent an important threat to yam (Dioscorea spp.) production in West Africa. With the aim to characterize the species affecting yam plants, for control and resistance screening purposes, surveys were conducted in the main yam producing areas. Infected, galled tubers were collected from farmers and markets in 12 states in Nigeria. To trap Meloidogyne species, infected yam peels were inoculated in a ring surrounding the root system of susceptible tomato cv ‘‘Ibadan local’’ and celosia plants. Young egg-laying females were extracted from roots of infected tomato and or celosia plants and used for enzymatic (Esterase: EST and Malate dehydrogenase: MDH) analysis with PhastSystem. Out of 46 populations examined, results revealed that yam tubers were mostly infested by Meloidogyneincognita (68%), followed by Meloidogyne javanica (16%), Meloidogyne enterolobii (2%) and Meloidogyne arenaria (5%). In most of the samples, tubers were infected by a single species. However, in 9% of the analysed samples, mixed populations of M. enterolobii and M. incognita (7 %) and M. enterolobii, M. incognita and M. javanica (2 %) were recorded. The distribution of the Meloidogyne spp. in the main yam producing areas is discussed. This constitutes the first report of M. arenaria and M. enterolobii on yam in West Africa. IDENTIFICATION AND MOLECULAR ANALYSIS OF MYLONCHULUS PAITENSIS YEATES, 1992 BASED ON 18S RDNA. Koohkan, M. and E. Shokoohi. Plant Protection Department, Agricultural Faculty, Shahid Bahonar University of Kerman, Kerman, Iran. The genus Mylonchulus is one of the most widely distributed genera among the order Mononchida. Mylonchulus paitensis was originally described from New Caledonia and then reported from Pakistan and Iran. During a survey of grasses in the Kerman province, Iran, a species of Mylonchulus was extracted using the adapted Baermann tray method and identified according to morphological characters as M. paitensis. Molecular study of 18S rDNA region of M. paitensis and comparison with available sequences deposited in GenBank, demonstrated that the studied population resembles Mylonchulus sigmaturus (AB361446, AB361447 from Japan and AY284755 from the Netherlands) and Mylonchulus arenicolus (AF036596 from UK) with 98% similarity. Phylogenetic analysis using the neighbor-joining method revealed that this population forms a monophyletic group with M. sigmaturus, Mylonchulus brachyuris, Mylonchulus oceanicus, Mylonchulus hawaiiensis, Mylonchulus rotundicaudatus, Mylonchulus arenicolus and Mylonchulus mulveyi. The maximum genetic distance (using Maximum Composite Likelihood method) was 0.084 between Iranian M. paitensis and M. mulveyi (AB361448; AB361449). This species has 17 and 20 nucleotide difference with Japanese and the Netherland population of the same species, respectively. In addition it differs from M. arenicolus by 18 nucleotides. The content of G+C in the 18s rDNA sequence of this isolate is 42.3%. This is the first study of 18S rDNA of M. paitensis in Iran. In conclusion, morphological and molecular study indicates that the Iranian population is M. paitensis. Furthermore, exploring more genes (e.g. mtDNA and 28S rDNA) are necessary for better understanding the relationship of the species of this group of nematodes. LONG-TERM EFFECTS OF EIGHT TREATMENTS TO CONTROL PLANT-PARASITIC NEMATODES AND VERTICILLIUM DAHLIA. Korthals, G.W., T.C. Thoden, W. van den Berg, L.P.G. Molendijk and J.H.M. Visser. Applied Plant Research, Wageningen University and Research Centre, Edelhertweg 1, 8219 PH Lelystad, The Netherlands. There is an urgent need to develop sustainable methods for management of soil pathogens, such as Pratylenchus penetrans and Verticillium dahliae. Ultimately this should be investigated with long-term measurements of biological and chemical parameters and their final impact on crop yield. The present study focusses on eight soil treatments (compost, chitin, marigold, grass-clover, biofumigation, anearobic soil disinfestation, a physical control method and a combination of marigold, compost and chitin) and two reference treatments (a chemical control with 300 L/ha Metam sodium and an untreated control). The effects of the10 treatments were studied for 6 years. The present study did demonstrate that in comparison to chemical control, additions of chitin, anaerobic soil disinfestation and marigold are already excellent alternatives for the

Abstracts 191 control of plant-parasitic nematodes and V. dahliae. Grass-clover, biofumigation, Cultivit and compost are not effective alternatives for chemical control yet and further development is needed. All treatments caused a yield increase in comparison with the control. The biggest increases of more than 60 % were found for the treatments with chitin. Furthermore it was demonstrated that these yield increases were probably less influenced by changes in chemical soil properties, but the consequence of changes in the soil biota, in this case especially the effective control of P. penetrans and V. dahliae. Furthermore it has been demonstrated that most of these soil treatments could be implemented in an arable crop rotation and probably adapted for many areas of the world. INVESTIGATIONS OF AN EMERGING PATHOGEN, BELONOLAIMUS SP., INFECTING PEANUT IN FLORIDA, USA. Kutsuwa1, K., D.W. Dickson1, J.A. Brito2, A. Jeyaprakash2 and A. Drew3. 1Entomology and Nematology Department, University of Florida, Bldg. 970, Natural Area Dr., P.O. Box 110620, Gainesville, Florida 32611, USA; 2Division of Plant Industry, Florida Department of Agriculture and Consumer Services, P.O. Box 147100, Gainesville, Florida 32614, USA; 3MultiCounty Extension Specialist, University of Florida, Bronson, Florida 32621, USA. Sting nematodes (Belonolaimus spp.) are important ectoparasitic nematodes, highly pathogenic on a wide range of plants in the sandy soils of the southeastern United States. Although this nematode is commonly found in Florida as a soilborne pathogen on turfgrasses and numerous agronomic and horticultural crops, it has not been reported infecting peanut. In the summers of 2012 and 2013, a sting nematode was found infecting three different peanut cultivars being grown on two separate peanut farms in Levy County, Florida. Belonolaimus spp. nematodes extracted from soil averaged 44/100 cm3 on peanut ‘TifGuard’, 39/100 cm3 on ‘Bailey’ in 2012 and 2013; and 28/100 cm3 on ‘Georgia 06’ on another farm in 2013. The damage was seen as large irregular patches at both farms. The foliage of infected plants were severely stunted and showed symptoms of nutrient deficiency. The root systems were severely abbreviated and there were numerous small, round brown lesions observed on pegs and pods of infected plants. Harvest taken from infected areas showed a yield reduction of 64% compared with non-sting nematode infested sites. Phylogenetic analysis of the D2-D3 expansion fragments of 28S rRNA and ITS1RNA genes from the nematodes infecting peanut were found to be 99% identical to Belonolaimus longicaudatus as reported by previous investigators. The sequences were deposited to GenBank (Accession No. KF963097-KF963100). Additionally, preliminary studies using the isolates from peanut showed similar morphology to that reported for B. longicaudatus. To our knowledge this is the first report of sting nematode infecting peanut grown under field conditions in Florida. STANDARDIZING DIAGNOSTIC QPCR ASSAYS FOR IDENTIFICATION AND DETECTION OF NEMATODES. Landeweert, R. and W. Mulder. Clear Detections, Binnenhaven 5, 6709 PD Wageningen, The Netherlands. A molecular detection test enables accurate fast screening of nematodes and is well suited for high throughput analyses. The accuracy of Molecular methods can be influenced by many factors. Various factors need to be taken into consideration upon chosing a molecular test as an identification tool, including most optimal gene, primer sequence and (q) PCR chemicals Therefore, the transformation of a promising primer pair into a robust diagnostic (q)PCR assay needs a lot of care and should be well documented. To promote consistency between nematode diagnostic laboratories, standardization of molecular methods is a prerequisite, just like proper method validation. Full understanding of the variables influencing the final results – ranging from proper nematode DNA extraction to proper functioning of the (q)PCR test and data interpretation – is therefore very critical. A number of examples will be presented during this congress on factors that influence (q)PCR test performance, which will be of help when deciding upon the development of one’s own test (‘homebrew’), either published or not, or the purchase of a ready-made diagnostic assay. For standardization purposes is will be of great value if laboratories either provide transparency on all experimental details, or choose to work with pre-defined and well-validated (q)PCR tests. A NEW IN-FURROW NEMATICIDE FOR ROTYLENCHULUS RENIFORMIS AND MELOIDOGYNE INCOGNITA NEMATODE MANAGEMENT IN COTTON. Lawrence, K.S., C. Land and R. Sikkens. Department of Entomology and Plant Pathology, Auburn University, Auburn, Alabama 36832 USA. Experimental nematicide in-furrow combinations were evaluated for Rotylenchulus reniformis and Meloidogyne incognita nematode management for cotton production in Alabama. Field tests were placed in RCBD with five replications and were irrigated. The experimental nematicide was applied at planting in furrow spray application and some treatments added a foliar spray application of the experimental or Vydate C-LV applied at the 6 to 8 leaf stage. Initial population densities were 3553 vermiform life stages per 150 cm3 of soil for R. reniformis and 160 J2 per 150 cm3 of soil for M. incognita. Plant stand or plant survival was similar between all nematicide treatments and the untreated control in both nematode fields and no phytotoxicity was observed with the experimental nematicides. All the experimental nematicide treatments applied at 0.53 l/ ha or 1.06 l/ha reduced (P < 0.05) R. reniformis and M. incognita numbers compared to the untreated control in both tests. The average reduction for the 1.06 l/ha application was 85% and 74% for R. reniformis and M. incognita, respectively. Seed cotton yield were increased (P < 0.10) by the .06 l/ha application. The 1.06 l/ha rate increase seed cotton by 737 and 655 kg/ ha for R. reniformis and M. incognita, respectively. Rotylenchulus reniformis and M. incognita 40DAP population’s densities

192 Journal of Nematology, Volume 46, No. 2, June 2014 (P < 0.05) negatively affected seed cotton yield. Regression equations of yield = -0.0503x + 3147.9; R2 = 0.4515 for R. reniformis and yield = -0.0743x + 1787.2; R2 = 0.6236 for M. incognita. MACROECOLOGY OF FREE-LIVING INTERTIDAL NEMATODES ALONG THE COAST OF CHILE AND THE ANTARCTIC PENINSULA. Lee, M.R. Centro i;mar, Universidad de Los Lagos, Camino a Chinquihue km6, Puerto Montt, Chile. Standard macroecological hypotheses were explored using the free-living intertidal nematode fauna from sandy beaches along the coast of Chile and the Antarctic peninsula. Species richness, abundance, range-size (Rapoport’s rule) and body size (Bergman size clines) were all analysed over the latitudinal gradient. Replicate quantitative samples were collected in the intertidal of 111 beaches between Arica (18.412 8S, 70.326 8W) on the border with Peru down to Margarite Bay (67.818 8S, 67.201 8W) in Antarctica. Meiofauna were extracted and the nematode species present identified and enumerated. The trends were assessed for their association with the principal environmental variables, surface sea water temperature, coastal primary productivity and coastline complexity. Species richness in exposed beaches declined with increasing latitude, as did abundance. Range size increased with increasing latitude. Body size neither declined nor increased with increasing latitude at the phylum level. At the level of individual species changes in body size were inconsistent, with some species increasing, and others decreasing or displaying no trend with increasing latitude. The latitudinal trend in species richness was most strongly associated with temperature, with one possible hypothesis being that the fauna has equatorial origins and that southward dispersal is limited by temperature. Antarctic nematode fauna was less diverse and abundant than the nematode fauna in southern Chile, and contrary to predictions nematodes in Antarctica were not larger than those from more northerly latitudes. These results are relevant as they are based on samples and not data extracted from the literature. THE CHOLECYSTOKININ SIGNALING PATHWAY AS A TARGET FOR THE DEVELOPMENT OF NOVEL NEMATICIDES. Lemoine1 L., T. Janssen1, N. Suetens1, W. Grant2, B. Landuyt1, L. Schoofs1 and L. Peeters1. 1KU Leuven (University of Leuven), Oude Markt 13, 3000 Leuven, Belgium; 2La Trobe University, Melbourne VIC 3086, Australia. Cholecystokinin (CCK) neuropeptides and their corresponding receptors are known to play a role in the management of energy supplies and feeding behavior. Both the receptor and its ligands are structural and functional very well conserved among different species, including nematodes. We conducted a differential feeding micro-array experiment and found that this signaling system affects a lot of genes involved in sugar- and fat metabolism and responses to starvation. The expression of the receptor and its ligands are upregulated under starvation and during the dauer stage, when food consumption is stopped. We also noticed that worms with a defect CCK system display abnormalities in food-consumption, locomotion and egg laying. Localization constructs revealed expression of the receptor in the intestine. We also identified an NLP-12-like neuropeptide precursor (the nematode orthologue of CCK) in Strongyloides ratti and in Parastrongyloides trichosuri, gastrointestinal parasites of rats and Australian possums, respectively. P. trichosuri NLP-12 peptides are able to activate the C. elegans CKR-2 receptor in the nanomolar range, as does C. elegans NLP-12. In C. elegans NLP-12’s inhibitory effects on locomotion and egg laying, two important targets for nematicides, were rescued with P. trichosuri genomic nlp-12. We could conclude from these experiments that the CKR-2 receptor is an interesting target for intervention in the development of nematicides. Therefore, we determined the core receptor-activating peptide in C. elegans, which is conserved among freeliving and parasitic nematodes. As a result we use this as a starting point in the design of non-peptide mimetics and have begun testing analogues. PLURONIC GEL, A USEFUL MEDIUM TO STUDY ENTOMOPATHOGENIC NEMATODE HOST HABITAT FINDING BEHAVIOR IN RESPONSE TO ENVIRONMENTAL CUES FROM THE PLANT. Li1, C., Y. Wang2, F. Pan1 and C. Wang1. 1Key Laboratory of Mollisols Agroecology, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin 150081, China; 2Center for Vector Biology, Rutgers University, New Brunswick, New Jersey, 08903, USA. Pluronic F-127 is a non-toxic and thermoreversible transparent copolymer, which allows free nematode movement inside the gel. Pluronic gel has been proved to be a useful medium to study plant parasitic nematode host finding behaviour. For the first time, we used pluronic gel to investigate how plant roots affect localisation and host habitat seeking of entomopathogenic nematodes (EPN). One isolate of EPN, Heterorhabditis bacteriophora HBN (Hb-HBN) from north-eastern China was found to be an effective bio-control agent against Chinese chive maggot (Bradysiaodoriphaga sp.). Among the nematodes tested, Hb-HBN had the strongest attraction to chive roots, stronger attraction to garlic and green onion roots, and weak attraction to tomato and soybean roots 4hr post-exposure (P < 0.05). The result may explain why HB-HBN was the most effective nematode against the maggot. Further tests have shown that the nematodes were more attracted to wounded roots than nonwounded roots, younger roots than older roots, which indicated that the nematodes prefer the habitat where their hosts are more likely present. Moreover, we found that nematode age, species, and strains significantly affected their attraction to chive root. Further studies about whether chive roots emit compounds to recruit EPN to protect their root system from herbivores

Abstracts 193 are being conducted in pluronic gel. These studies will provide further understanding of EPN infection mechanisms and an important parameter to predict the success of field application. TOWARDS INTEGRATED MANAGEMENT OF CEREAL CYST NEMATODE: AN EXAMPLE FROM RESEARCH IN JIANGSU PROVINCE, CHINA. Li, H. and X. Wang. Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, People’s Republic of China. The cereal cyst nematode (CCN) was first reported on wheat in Jiangsu Province, China in May, 2009. Jiangsu Province is the fifth largest producer of wheat in China (approx. 2.3 Mha). Knowledge of the biology of CCN and the development of control methods are essential for integrated management of this nematode. During a survey in the province, CCN was detected from 152 towns in 47 counties. The wide distribution and high population numbers had notable adverse impact on wheat production. All CCN populations were identified as Heterodera avenae and finished only a single life cycle on winter wheat during a growing season. The hatch peak and therefore the largest infection to roots were annually at the end of February. Five granular nematicides (GR) were screened in field trials for effectiveness in CCN control and 0.5% avermectin GR was successful in both inhibiting nematode reproduction and as well as improving wheat growth. A dosage of 30 kg/ hm2 was recommended to be applied in heavily infested fields during the regreening period. Seed-coating chemicals were also screened and Gannong seed-coating III exhibited high control efficacy. The pathotypes of H. avenae Jiangsu populations were determined as belonging to the Ha1 group. Planting resistant cultivars is usually the most economical and practical measure for CCN control and three wheat cultivars were evaluated as highly resistant to H. avenae. The Huamai No.1 cultivar can successfully be planted in heavily infested areas. INTERACTIONS BETWEEN COTTON AND RENIFORM NEMATODE. Li1, W., P. Agudelo1, and C. Wells2. 1School of Agricultural, Forest, and Environmental Sciences, Clemson University, Clemson, South Carolina, USA, 29634; 2Department of Biological Sciences, Clemson University, Clemson, South Carolina, USA, 29634. Reniform nematode (Rotylenchulus reniformis) is an important problem for cotton production in the United States, but knowledge of cotton resistance to reniform nematode is very limited. The objectives of this project were: to generate a histological and gene expression time course for the cotton root-reniform nematode interaction; to identify and validate key candidate genes involved in cotton response to reniform nematode infection; and to identify metabolic and regulatory pathways associated with syncytium formation. Resin sections of root tissue from susceptible and resistant cotton were made after inoculating with reniform nematode, using a split-root system, and observations were made at 3, 6, 9, and 12 days after inoculation with the nematode. Trinity de novo transcriptome assembly was performed using root RNA from multiple infection stages, and transcripts were functionally annotated with Blast2GO. Candidate genes that were strongly up- and down-regulated during syncytium development were identified. Genes for detailed characterization were selected based on differential expression and our understanding of nematode-host root biology. Target proteins were localized in the root cells with fluorescent microscopy. The information gathered in this project is critical to the breeding and engineering of nematoderesistant cotton plants. SOYBEAN CYST NEMATODE RESISTANCE IN PI 567516C SOYBEAN: A PROMISING NEW SOURCE OF RESISTANCE. Lian1, L., F. Wang2, R.L. Denny1, J.H. Orf3, N.D. Young1 and S. Chen1,4. 1Department of Plant Pathology, University of Minnesota, St. Paul, Minnesota 55108, USA; 2Life, Agriculture and Forestry College, Qiqihar University, Qiqihar, Heilongjiang 161006, China; 3Department of Agronomy and Plant Genetics, University of Minnesota, St. Paul, Minnesota 55108, USA; 4Southern Research and Outreach Center, University of Minnesota, Waseca, Minnesota 56093, USA. Soybean cyst nematode (Heterodera glycines) is the most serious yield-limiting pathogen on soybean (Glycine max), which is most effectively managed by host resistance. However, most commercial soybean cyst-resistant cultivars in the USA are developed from PI88788 and Peking, and frequent use of the limited number of resistance sources has shifted virulence phenotypes of H. glycines populations (HG Types) to new types that can overcome originally resistant cultivars. A recently identified soybean line PI 567516C is a promising alternative source of soybean cyst nematode-resistance. PI 567516C is highly resistant to HG Type 2.5.7 (race 1) which is virulent to PI 88788. Interestingly, PI 567516C is highly resistant to some populations of HG Type 1.3.6 (race 14) which is virulent to Peking, but not other populations of race 14. Genetic studies revealed that the major QTLs conferring H. Glycines-resistance in PI567516C are distinct from the quantative trait loci (QTL) in PI 88788, Peking, and many other -resistance sources. Two QTLs located on chromosomes 10 (LG O) and 8 (LG A2) in PI 567516C were identified to be resistant to H. glycines. The QTL on chromosome 10 was the major one with 32.0% additive heritability and 68.7% of the genetic variance of resistance to HG Type 2.5.7, and the closest single nucleotide popymorphism (SNP) ()marker was BARC-008021-00209. The QTL on chromosome 8 displayed 6.8% additive heritability and 14.8% of the genetic variance of resistance to HG Type 2.5.7 and the closest SNP marker was BARC-028207-05794. The two SNP markers can be used to select soybean cyst nematode-resistance from PI 567516C in soybean breeding through marker-assisted selection.

194 Journal of Nematology, Volume 46, No. 2, June 2014 PLANT NEMATOLOGY, EDUCATION AND TRAINING IN CHINA. Liao1, J. and H. Li2. 1Laboratory of Plant Nematology/Guangdong Province Key Laboratory of Microbial Signals and Disease Control, South China Agricultural University, Guangzhou 510642, People’s Republic of China; 2Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, People’s Republic of China. Due to the great losses caused by some parasitic nematodes on many crops in China, more and more researchers are involved in this area. Presently, there are about two hundred researchers located in different provinces in China. They focus on nematodes of agricultural importance, especially on species identification, basic biology, molecular aspects and disease management. In recent years, the Chinese government funded research on root-knot nematode, cyst nematode and pine wood nematode because of their serious damages on crops and forest. The interaction between nematodes and host plants and novel management techniques will be the ‘‘hot’’ research topics which attract Chinese nematologists. A good system for Nematology education and training has been developed through the efforts of Chinese nematologists. These include Bachelor, Master and PhD degree programmes. Incorporation of Nematology curriculums from USA and Europa at local universities is also a possibility. THE GENOME SEQUENCE AND LIFE-STAGE SPECIFIC TRANSCRIPTOMES OF POTATO CYST NEMATODE. Lilley1, C.J., J.A. Cotton2, V. Blok3, S. Eves-van den Akker1,3, L.M. Jones1, A.J. Reid2, P. Thorpe1,3, M. Berriman2, J.T. Jones3 and P.E. Urwin1. 1Centre for Plant Sciences, University of Leeds, Leeds LS2 9JT, UK; 2Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK; 3James Hutton Institute, Invergowrie, Dundee DD2 5DA, UK. Potato cyst nematodes are major pathogens of potato crops in temperate regions, making them some of the most economically important plant parasitic nematodes. Research to develop novel approaches for control of these and other cyst nematodes will be significantly enhanced by a greater understanding of the molecular basis of the parasitic interaction and the key nematode genes required for this. A complete draft genome sequence of the white potato cyst nematode Globodera pallida has been assembled, together with transcriptomic data from most of the nematode life cycle, particularly focusing on the life cycle stages involved in root invasion and establishment of the biotrophic feeding site. Despite the relatively close phylogenetic relationship with root-knot nematodes, there is a very different gene family content between the two groups and in particular extensive differences in the repertoire of effectors, including an enormous expansion of the SPRY domain protein family in G. pallida, which includes the SPRYSEC family of effectors. This highlights the distinct biology of cyst nematodes compared to the root-knot nematodes that were, until now, the only sedentary plant parasitic nematodes for which genome information was available. The repertoires of genes likely to be important in understanding the unique biology of cyst nematodes and those that represent potential chemical targets and other targets for control have been analysed. A recently assembled draft genome for the closely related species Globodera rostochiensis will allow valuable comparative studies. MOLECULAR CHARACTERIZATION OF RESISTANCE RESPONSES OF COFFEA CANEPHORA ‘CLONE 14’ UPON INFECTION WITH MELOIDOGYNE PARANAENSIS. Lima1,2, E.A., F.A. Carneiro2, T.S. Costa2, E.C.S. Reˆgo2, A. Jorge Ju´nior2, C. Furlanetto1, P. Marraccini2,3, R.M.D.G. Carneiro2 and A.C. Andrade2. 1Dep. Fitopatologia, Universidade de Brası´lia, 70910-900 Brası´lia, DF, Brazil; 2Embrapa Recursos Gene´ticos e Biotecnologia, 70770-917 Brası´lia, DF, Brazil; 3CIRAD UMR AGAP, 34398, Montpellier, France. Coffee is one of the major commodities in the world and an important source of income for producing countries. However, biotic and abiotic stresses are great limiting factors to coffee yield. In Brazil, root-knot nematodes cause considerable yield reduction and the use of resistant plants is the most promising method to control Meloidogyne spp. The aim of this work was to characterize the molecular mechanism underlining the previously identified resistance to M. paranaensis in C. canephora ‘Clone 14’ by means of RNAseq experiments. Differential expression using RNA extracted from roots of plants from clones 14 and clone 22 of C. canephora, previously identified as resistant and susceptible to M. paranaensis, respectively, were grown in sand and inoculated. Root samples were collected at different time points post inoculation as well as roots from an uninfected plant. The RNA was treated with DNAse and subsequently, a portion of the sample was lyophilized for RNAseq experiments and another portion kept for validation by qPCR experiments. Results of the identified candidate genes with differential expression among resistant (Clone 14) and susceptible (Clone 22) genotypes will be presented and discussed. AGGRESSIVENESS OF MELOIDOGYNE JAVANICA POPULATIONS ON COMMERCIAL POTATO CULTIVARS. Lima-Medina1, I., J.T. Schafer2, C.B. Gomes1, M. Vizzoto1, A.C. Krolow1, R.M.D.G. Carneiro3 and V. Correa3. 1 Embrapa Clima Temperado, Cx Postal 403, Pelotas/RS, Brazil; 2Graduated student in Plant Pathology, PPGFS/Universidade Federal de Pelotas, Campus Universitario s/n C. P. 354, Pelotas/RS, Brazil; 3Embrapa Recursos Gene´ticos e Biotecnologia, C.P. 02372, 70849-979, Brası´lia DF, Brazil. The genus Meloidogyne is widely found in the different potato production regions of Brazil. However Meloidogyne javanica is the most frequent species that causes damage in potato crop. In order to study the aggressiveness of M. javanica, four populations (P1, P3 and P4: Est J3; P2: Est J2a) from southern Brazil was evaluated in two commercial potato cultivars (BRS Clara and Agata) in greenhouse conditions. Individual plants of the two cultivars, kept in pots with sterilized soil, were

Abstracts 195 inoculated with 5,000 eggs and second stage juveniles of M. javanica using six replicates. Fifty-five days after inoculation, each plant was evaluated for number of galls/root, galls number/1.76 cm2/tuber, nematode reproduction factor (RF). Subsequently, the sensorial quality and the respective total phenolic compound levels in the tubers were also determined. P2 and P4 were the most aggressive M. javanica populations exhibiting the higher values of all studied nematode variables interfering in the expression of symptoms on both potatoes cultivars. Additionally, all the populations affected the flavour of cooked potatoes in both of the tested cultivars and higher levels of phenolic compounds were observed in potatoes infected with M. javanica P2 population. REACTION OF POTATO CULTIVARS TO MELOIDOGYNE HAPLA AND M. MOROCCIENSIS. Lima-Medina1, I., J.T. Schafer2 and C.B. Gomes1. 1Embrapa Clima Temperado, Cx Postal 403, Pelotas/RS, Brazil; 2PPGFS/Universidade Federal de Pelotas, Campus Universita´rio s/n C. P. 354, Pelotas/RS, Brazil. In Brazil, different species of root-knot nematodes, affect potatoes. However, there are few studies of genetic resistance to Meloidogyne species. The objective of this study was to evaluate the reaction of nine commercial potato cultivars to Meloidogyne hapla and Meloidogyne morocciensis in greenhouse conditions. Potato plants kept in pots with sterilized soil were inoculated with 5,000 eggs and second stage juveniles of M. hapla or M. morocciensis/plant using six replicates/ genotype. ‘Santa Cruz’ tomato plants received the same inoculum level and were used as control. Fifty-five days after inoculation, each plant was evaluated for the number of galls, eggs and juveniles./root system. Subsequently, the reproduction factors (RF) of the two Meloidogyne species were determined in the different genotypes. Among the tested cultivars, BRS Ana, Asterix, BRSIPR Bel, Cota, Cristina, BRS Clara, Catucha, and Eliza were susceptible to both Meloidogyne species. Besides being susceptible to M. morocciensis, ‘Agata’ was also moderately susceptible to M. hapla. FREE LIVING NEMATODES AS INDICATORS OF THE BIOLOGICAL STATUS OF AUSTRALIAN CEREAL SOILS. Linsell1, K., A. Stirling2, D. Hartley3, Herdina1, A Cheshire4, J. Nobbs1, A. McKay1, G. Stirling2 and K. Ophel Keller1. 1South Australian Research & Development Institute (SARDI), GPO Box 397, Adelaide, 5001, South Australia, Australia; 2Biological Crop Protection Pty. Ltd., 3601 Moggill Road, Moggill, 4070, Queensland, Australia; 3 CSIRO Ecosystems Science, GPO Box 1700, Canberra, 2601, ACT, Australia; 4Science to Manage Uncertainty, 24 Winding Way, Belair, South Australia, 5052, Australia. The impact of management practices on the biological status of cereal-growing soils was investigated across a range of Australian soil types and climates through nematode community analysis, across multiple years. A multivariate statistical analysis approach identified the two key drivers influencing changes within free-living nematode communities to be soil type which is linked to regional rainfalls, particularly 1-3 months prior to crop sowing and the application of certain nutrients, particularly N, P, S and Cu. Significant shifts in nematode population structures were also characterised by tillage regimes when analysed by soil type. Stubble management and prior plantings did not influence community changes except where canola and legumes were included in the cereal rotations. A Bray–Curtis measure of similarity characterised the contribution of each species/genera driving the changes between each management/environmental treatment and seventeen free-living species were identified as good indicators. Since manual nematode community analysis is laborious and requires specialised taxonomic skills, molecular technologies were developed to allow routine indicator identification and quantification in soil, which can be delivered as part of diagnostic service for soil-borne pathogens. Nine DNA tests were developed incorporating eleven of the free-living nematode indicators and are predicted to detect more than 80% of the species present in Australian cereal cropped soils. There was a very strong correlation between free-living nematode community structures obtained from the manual count and DNA tests. Therefore, we concluded that DNA tests are a sensitive, quick and robust tool for assessing free-living nematode communities, and provide a useful indication of a soil’s biological status. DAMAGE FUNCTIONS OF MELOIDOGYNE JAVANICA ON ZUCCHINI SQUASH AND RELATIVE LEAF CHLOROPHYLL CONTENT. Lo´pez-Go´mez1, M., F.J. Sorribas2, M. Talavera3 and S. Verdejo-Lucas1,4. 1IRTA. Crta de Cabirls Km 2. 08348 Cabrils, Barcelona, Spain; 2Universitat Polite`cnica de Catalunya. 08860 Castelldefels, Barcelona, Spain; 3IFAPA. Camino de Purchil s/n. Granada. Spain; 4IFAPA. Camino de San Nicola´s, 1. 04745 La Mojonera, Almerı´a. Spain. Yield losses in cucurbits have been reported in many horticultural regions. Zucchini-squash is an important crop in southern Spain accounting for one third of the production in the country. For sustainable management of root-knot nematodes in susceptible crops, it is essential to develop accurate information on population densities that cause yield losses and quantify them into plant damage functions. This study was conducted to determine maximum multiplication rate and equilibrium density of Meloidogyne javanica on zucchini squash cv Amalthee in response to increasing initial population densities and to develop damage function models. Seedlings were inoculated with 0, 0.5, 1, 2, 4, 8, 16, 32, 64, 128, 256, 512, and 1024 eggs of Meloidogyne per cm3 of soil in repeated experiments in a greenhouse. The maximum multiplication rate was 511, and the equilibrium density 8135 eggs + J2 cm-3 soil. The relationship between Pi and relative plant top dry weight fitted the Seinhorst damage function model (R2=0.53; P=0.0002). Values for minimum relative yield, tolerance limit, and constant z

196 Journal of Nematology, Volume 46, No. 2, June 2014 were 0.77, 3.4 J2 cm-3 soil, and 0.95, respectively. Relative leaf chlorophyll content measured 40, 50, 60 and 70 days postinoculation (dpi) showed a significant reduction (P 0.05). However, sunn hemp consistently enhanced beneficial nematodes number in all experiments (P < 0.05). Effects of pigeon pea on beneficial nematodes were inconsistent. Interns had recommended that farmers should add at least 0.5% sunn hemp in their fields to enhance beneficial nematodes, and could choose pigeon pea as an alternate cover crop. Results were presented by interns at Regional or National conferences. It is suggested to include research focused internship projects in undergraduate student’scourse curriculums. THE GENUS HELICOTYLENCHUS IN THE GRASSLAND BIOME OF SOUTH AFRICA. Marais1, M., A. Swart1, A.S. Schoeman2 and L.R. Tiedt3. 1National Collection of Nematodes, Biosystematics Programme, ARC-Plant Protection Research Institute, Private Bag X134, Queenswood 0121, South Africa; 2Sports Turf Solutions, Pretoria, South Africa; 3 Laboratory of Electron Microscopy, North West University, Potchefstroom, 2520, South Africa. The grassland biome is one of the most threatened biomes because it is here that the bulk of South Africa’s maize, beef, and milk production are anchored. Moreover, urbanisation and unbridled mining are also placing immense pressure on this biome. For example 77 % of the Gauteng Province is classified as grassland, but only 5 % of this area is protected. Datasets of the distribution of Helicotylenchus species contained in the South African Plant-Parasitic Nematode Survey (SAPPNS) database were used to test the theory that grassland disturbance leads to decline in biodiversity of nematode fauna. In this paper the diversity of Helicotylenchus species found in pristine grassland, agricultural ecosystems, and urban areas were taken as indicators of ecological disturbance. Sports fields were chosen as representatives of urban ecosystems. The genus Helicotylenchus is widespread in South Africa, with 25 of the 33 South African Helicotylenchus species reported from the grassland biome. The two most common species were Helicotylenchus digonicus and Helicotylenchus dihystera, and both were found in 13 of the 15 different vegetation types that comprise the grassland biome. According to the SAPPNS datasets, there was a decline in biodiversity as the habitat changed from pristine grassland to crop plants to urban areas. Twenty three species were reported from uncultivated areas, 14 species from crop plants, and 10 species from the grasses grown on sports fields. Helicotylenchus is ideally suited for a study on the impact of cultivation on nematode diversity. EVIDENCE OF INCREASING NEMATODE NUMBERS AND THE POTENTIAL OF BIOFRIENDLY PRODUCTS IN THEIR MANAGEMENT. Martin, T.J.G. and C.C. Fleming. Agri-Food and Biosciences Institute, Newforge Lane, Belfast, Northern Ireland. Nematodes are one of the most important pests worldwide causing crop losses of between 9% and 15%. Considering that the projected increase in food consumption by 2050 is assessed at 75%, it becomes imperative that monitoring takes place to have a current up to date knowledge of the pest status for any region. Current trends throughout Northern Europe would indicate that there are nematode populations establishing in regions where previously they had not been detected. These facts combined with the decision to decrease the amount of inorganic pesticides leads to the need for new and novel nematode population management techniques which are more acceptable and environmentally friendly. It will be demonstrated that within the last eight years genera and species of nematodes, not previously detected in Irish soils, have been found and have become established. These are causing major damage in amenity grassland and on some occasions in agricultural crops. The potential of some bio-friendly products are investigated for their ability to manage nematode populations or for their capacity to protect the plant against attack. TIME INTERVAL BETWEEN SUNN HEMP COVER CROPPING AND CASH CROPPING FOR NEMATODE AND CROP MANAGEMENT. Martiney, C.L. and S.P. Marahatta. Science and Math Division, Kaua’i Community College, University of Hawai’I, Hawaii, USA. Sunn hemp (Crotalaria juncea) is a tropical legume that when used as a cover crop, has allelopathic properties that suppresses plant-parasitic nematodes, enhances beneficial nematodes, and improves soil health. However, the allelopathic effects of sunn hemp could also reduce cash crops’ germination, if sunn hemp is planted immediately after cover cropping. To find out an optimum interval of days between sunn hemp cover cropping and cash cropping, a field experiment and a shade house experiment was conducted in 2013. In the field experiment, sunn hemp and pigeon pea (Cajanus cajan) cover crops were separately grown for one month, tilled and incorporated into the soil. During the cover cropping period, a separate plot with no cover crop fallow was maintained as a control.Corn, Zea mays, was seeded immediately after incorporating cover crops into the soil. In the shade house experiment, fresh sunn hemp was incorporated with soil samples at 1.0% (w/w) (sunn hemp+) or non-incorporated (sunn hemp-), kept in a 10-cm diam plastic pot, and left for zero week (W-0), one week (W-1) and two weeks (W-2). At W-0, W-1 and W-2, corn seeds were seeded. In the field experiment, compared to pigeon pea and control, sunn hemp reduced corn germination (P < 0.05). In the shade house experiment, compared to sunn hemp(-), sunn hemp(+) reduced corn germination on W-0 (P < 0.05), but not on W-1 and W-2 (P < 0.05). When farmers use sunn hemp as a cover crop, it is best to wait one week before seeding a cash crop in order to optimize seed germination.

200 Journal of Nematology, Volume 46, No. 2, June 2014 POCHONIA CHLAMYDOSPORIA AND PLANT AQUEOUS EXTRACTS, A COMBINED APPROACH FOR MANAGEMENT OF MELOIDOGYNE INCOGNITA IN BEANS. Martı´nez1, K.V., R.M. Belmont1 and R.H. Manzanilla Lo´pez2. 1Centro de Desarrollo de Productos Bio´ticos Instituto Polite´cnico Nacional. Apartado postal 24. Yautepec Morelos Me´xico 62731; 2Department of AgroEcology, Rothamsted Research. Harpenden Herts, AL5 2JK, UK. Pochonia chlamydosporia is a facultative parasite of Meloidogyne spp. eggs, and is increasingly being used in combination with other crop management strategies, such as the nematicidal properties of extracts from several plant families. In the present work, the combination of both management strategies was tested in beans against M. incognita. Bean plants grown under greenhouse conditions in a soil infested with M. incognita were added with powder from three milled plants (6 g/ 600 g of soil) belonging to three plant species with nematicidal properties (Chenopodium album, Raphanus raphanistrum, Thymus vulgaris) alone or in combination with P. chlamydosporia. One treatment also included the fungus and the three plant species together. All treatments were compared with the nematicide carbofuran. There was a significant reduction in root galling with carbofuran, the combination of the fungus with the three plant species applied together, and the combination of the fungus with Thymus vulgaris. There was no difference in green bean production between treatments. It was concluded that it is possible to combine both management strategies against M. incognita but further studies are required. BIOCIDAL EFFICACY OF ENTOMOPATHOGENIC NEMATODES AGAINST CATTLE TICKS. Maru1, A.K., S. Kachhawaha2, A.U. Siddiqui3 and S.K. Sharma3. 1Department of Entomology, B.A.C., Bihar Agricultural University, Sabour, Bhagalpur-813210, India; 2Krishi Vigyan Kendra, CAZRI, Pali-Marwar -306401, India; 3Department of Nematology, R.C.A., MPUAT, Udaipur-313001, India. Entomopathogenic nematodes have been successfully used as biological control agents of insects of economically important crops. In the present study, the biocidal efficacy of two different strains of entomopathogenic nematodes, Steinernema carpocapsae STSLU and S. carpocapsae STUDR against two different cattle ticks, Rhipicephalus microplus and Hyalomma savignyi was evaluated on the basis of percentage mortality under laboratory conditions. The adult female cattle ticks were inoculated with infective juveniles (IJs) of both the strains of S. carpocapsae at different inoculum levels. All the treatments were replicated four times at 208 C in a B.O.D. incubator. The percentage mortality of the cattle ticks was determined every 24 hours up to 120 hours from the time of inoculation. Rhipicephalus microplus was more susceptible to both strains than H. savignyi.Steinernema carpocapsae STSLU was more efficient than S. carpocapsae STUDR and cause 100 and 97.5 % mortality of R. microplus and H. savignyi, respectively at a concentration of 250 IJs/Petri dish after 120 hours. Both the tested strains showed promise for the control of R. microplus and H. savignyi, and will be evaluated further in field conditions. THE LEVAMISOLE-SENSITIVE NICOTINIC ACETYLCHOLINE RECEPTOR OF GLOBODERA PALLIDA. Marvin1, J., A. Crisford2, L. Jones1, C. Lilley1, V. O’Connor2, L. Holden-Dye2 and P. Urwin1. 1Centre for Plant Sciences, University of Leeds, Leeds, LS2 9JT, UK; 2Centre for Biological Sciences, University of Southampton, Southampton, SO17 1BJ, UK. The potato cyst nematode Globodera pallida costs the UK potato industry over £50 million per annum and novel effective control of the pathogen is essential. The ability to locate and migrate to host-roots in the soil is a common requirement among many plant-parasitic nematodes and may be a key target for control. A target for control may be the mechanism of muscle contraction by the neurotransmitter acetylcholine. The paralytic drug levamisole is known to act via acetycholine receptors. Globodera pallida is more resistant to levamisole than Caenorhabditis elegans. In C. elegans the levamisole-sensitive acetylcholine receptors of body wall muscle are largely comprised of five subunits; UNC-38, UNC-63, UNC-29, LEV-1 and LEV-8. Orthologues of unc-38; unc-63 and unc-29 have been identified and cloned from G. pallida. Orthologues for lev1 and lev-8 have not been identified in G. pallida. The predicted amino acid sequence of Gp-unc-38 lacks important determinants for binding of acetylcholine and other agonists like levamisole. Caenorhabditis elegans mutants that are functionally null for unc-38 display an uncoordinated phenotype and increased resistance to levamisole. Transgenic expression of Gp-unc-38 in this mutant background rescues normal movement suggesting a functional reconstitution of the levamisole sensitive receptor, but does not restore full sensitivity to levamisole. This raises important questions about the arrangement and pharmacology of this receptor in G. pallida that are currently being investigated. Other members of the order Tylenchida share the same complement of receptor subunits and features of UNC-38, which may provide a unique target for control of plant-parasitic nematodes. TYLENCHULUS SEMIPENETRANS BIOTYPE IN SOUTH AFRICA: PONCIRUS BIOTYPE. Mashela1, P.W., Z.P. Dube1 and K.M. Pofu2. 1School of Agricultural and Environmental Sciences, University of Limpopo, Private Bag X1106, Sovenga 0727, South Africa; 2Agricultural Research Council- Vegetable and Ornamental Plant Institute, Private Bag X293, Pretoria 0001, South Africa. The reproductive potential (Pf/Pi) previously used to established the citrus nematode (Tylenchulus semipenetrans) biotype among differential hosts did not allow for inter- and intra-continental comparability. Therefore, the reproductive potential was used to re-assessed the citrus nematode biotype from T. semipenetrans isolates collected from 18 different citrusproducing district municipalities in South Africa on rough lemon (Citrus jambhiri), trifoliate orange (Poncirus trifoliate) and

Abstracts 201 olive (Olea europaea) under greenhouse conditions. Inoculum from each of the 18 districts constituted a single experiment, with three differential hosts arranged in a randomised complete block design, with 15 replications. Three months after inoculation in all 18 experiments, the reproductive potential (eggs + juveniles) suggested that the T. semipenetrans isolates did not reproduce on olive, but reproduced on the other two hosts. In conclusion, the use of reproductive potential provided conclusive evidence that the T. semipenetrans biotype in South Africa was poncirus and the information was comparable with those from other citrus-producing countries. SOIL ALLELOCHEMICAL RESIDUE EFFECTS FROM NEMARIOC-AL PHYTONEMATICIDE ON BRADYRHIZOBIUM JAPONICUM NODULATION, VIGNA UNGUICULATA AND MELOIDOGYNE SPECIES. Mashela1, P.W., K.M. Pofu2 and Z.P. Dube1. 1University of Limpopo, School of Agricultural and Environmental Sciences, Private Bag X1106, Sovenga 0727, South Africa; 2Agricultural Research Council – Vegetable and Ornamental Plant Institute, Private Bag X293, Pretoria 0001, South Africa. Nemarioc-AL phytonematicide consistently stimulated growth of tomato (Solanum lycopersicum) and suppressed population densities of Meloidogyne species under diverse environments. However, information on its soil allelochemical residual (SAR) effects on growth of successor crops and related population nematode densities is scant. A field trial was therefore conducted to investigate the SAR effects of nemarioc-AL on growth of cowpea (Vigna unguiculata) as a successor crop and the related population nematode densities. The SAR conditions were created in a 5 3 5 factorial tomato experiment, with the first and second factors being concentrations and application intervals, respectively. The concentration 3 application interval contributed 21% to total treatment variation (TTV) in number of galls, with permutations consistently reducing number of galls. The concentration factor contributed 12% and 8% to TTV in dry seed mass and number of nematodes, respectively, increasing (39-83%) dry seed mass and decreasing (88-94%) nematode densities. The application interval contributed 7% to TTV in dry shoot mass and number of nematodes each, increasing (39-69%) dry shoot mass and reducing (45-92%) nematode densities. In conclusion, appropriate permutations of concentration and application interval are necessary to allow for cowpea production as a successor crop to a tomato crop after using Nemarioc-AL phytonematicide. MANAGING PHYTOTOXICITIES IN PHYTONEMATICIDES: THE DOSAGE MODEL. Mashela1, P.W., K.M. Pofu2 and Z. Dube1. 1School of Agricultural and Environmental Sciences, University of Limpopo, Private Bag X1106, Sovenga 0727 South Africa; 2Agricultural Research Council Roodeplaat-VOPI, Private Bag X293, Pretoria, 0001, South Africa. Phytonematicides are inherently allelopathic to plants protected against plant-parasitic nematode damage. The dosage model was developed to manage phytotoxicity in phytonematicides. The objective of this study was to use empiricallyderived information to provide the three-step process of developing the dosage model. First, mean concentration stimulation range (MCSR) was established by subjecting test plants inoculated with nematodes to various concentrations of the phytonematicides for 56 days. Significantly affected plant variables were subjected to the Curve-fitting Allelochemical Response Dosage computer-based model to generate seven biological indices, where Dm and Rh indices were used to compute the nonphytotoxic concentration using the MCSR = Dm + (Rh/2) relation. Second, the application interval (T) for the MCSR value was established using the concept of a ‘‘30-day-week-period’’, where T comprised 0, 1, 2, 3 and 4 application intervals. Significantly affected plant variables were further subjected to lines of the best fit, where quadratic curves (bx2 + bx + c) were optimised using –b/2b2 to generate the appropriate T for the empirically-based MCSR value. Using the proportion of T to the crop cycle (Tcc), the application frequency (Tca) was derived (Tca = T/Tcc) to allow for computation of the dosage (D = MCSR 3 Tca, which is currently being successfully used under diverse environments to manage Meloidogyne species without causing phytotoxicity to tomato plants. NAPDH OXIDASES PRODUCE ROS THAT LIMIT CELL DEATH AND FACILITATE NEMATODE INFECTION IN ARABIDOPSIS THALIANA. Matera1, C., M.S. Hasan1, Z.S. Radakovic1, P. Gutbrod1, E. Rozanska2, M. Sobczak2, M. Angel Torres3, F.M.W. Grundler1 and S. Siddique1. 1INRES, Department of Molecular Phytomedicine, University Bonn, Bonn, Germany; 2Department of Botany, Warsaw University of Life Sciences (SGGW), Warsaw, Poland; 3Centro de Biotecnologı´a y Genomica de Plantas (UPM-INIA), ETSI Agro´nomos, Universidad Politecnica de Madrid, Pozuelo de Alarco´n, Madrid, Spain. Plants produce reactive oxygen species (ROS) in response to infection. These ROS activate defence responses and are mainly generated by plasma membrane-localized NADPH (Nicotinamide adenine dinucleotide phosphate) oxidases. Mutation of NADPH oxidase genes eliminates pathogen-induced ROS production and compromises immune responses. However, the functions of ROS in compatible plant-pathogen interactions remain largely unknown. Here, we demonstrated that the roots of Arabidopsis thaliana with mutation in NADPH oxidases failed to produce ROS when infected by the plant parasitic nematode Heterodera schachtii. Inhibition of ROS production by NAPDH oxidase with a small molecule Diphenlyl iodonium limited cell death spread and thus facilitated nematode-infection. Moreover, we discovered that the role of the ROS in antagonizing cell death during nematode infection is independent of salicylic acid. Our study provides understanding of how a pathogenic organism with destructive invasion behaviour is able to switch to a biotrophic life-style that requires viable

202 Journal of Nematology, Volume 46, No. 2, June 2014 host cells for feeding. The molecular mechanisms and signalling pathways involved in ROS-mediated restriction of cell death will be discussed. STUDY OF BRAZILIAN SPECIES OF MELOIDOGYNE: ENZYMATIC AND MOLECULAR CHARACTERIZATIONS. Mattos1, V.S., J.M.S. Monteiro1, J.E. Cares1, V.R. Correa1, 2, M.R.A. Almeida2, J.P. Borges3, R.M.D.G. Carneiro2. 1Dep. Fitopatologia, Universidade de Brası´lia, 70910-900 Brası´lia, Distrito Federal, Brazil; 2Embrapa Recursos Gene´ticos e Biotecnologia, 70849-979 Brası´lia, Distrito Federal, Brazil; 3Embrapa Hortalicxas, 70359-970 Brası´lia, Distrito Federal, Brazil. Root-knot nematodes (Meloidogyne spp.) are amongst the world’s major crop pathogens, due to their wide distribution, extensive host ranges, and ability to cause considerable economic losses. Efficient control of these pathogens depends upon correct species identification. Recently, five Brazilian species of Meloidogyne were described (Meloidogyne petuniae, Meloidogyne pisi, Meloidogyne phaseolus, Meloidogyne brasiliensis and Meloidogyne polycephannulata) without detailed molecular characterization. Therefore, the aim of this study was to characterise the type isolates using enzymatic profiles and SCAR markers. Esterase profiling and SCAR analyses were done according to described protocols. Meloidogyne petuniae Est Pe2 (Rm: 0.95, 1.08) and M. pisi Est Pi5 (Rm: 0.91,0.95,1.12,1.25,1.33) showed new species-specific esterase phenotypes. Meloidogyne phaseolus, M. brasiliensis and M. polycephannulata had the same species-specific esterase phenotypes as Meloidogyne morocciensis (Est A3, Rm: 1.1,1.2,1.3), Meloidogyne ethiopica (Est E3, Rm: 0.9,1.05,1.20) and Meloidogyne incognita (Est I2, Rm:1.0, 1.1), respectively. PCR with species-specific SCAR primers confirmed the results of esterase phenotypes: M. phaseolus and M. morocciensis, both showed a single specific fragment of 420 base pairs (bp), as well as in M. brasiliensis and M. ethiopica (350 bp) and M. polycephannulata and M. incognita (399 bp). Preliminary, morphological studies revealed similarities in characters of females, males and second stage juveniles among these species. Additional studies, including morphology, sequencing of 18S rRNA, internal transcribed spacer rRNA (ITS), D2-D3 fragment of 28S rRNA and phylogenic analyses have been carried out in order to clarify the taxonomic status of these Brazilian root-knot nematode species. EFFECTIVENESS OF ECOSYSTEM CONDITION AND FUNCTION INDICES USED IN SOIL NEMATOLOGY. Matveeva, E., A. Sushchuk and D. Kalinkina. Institute of Biology, Karelian Research Centre, Russian Academy of Sciences, 11 Pushkinskaya St., 185910 Petrozavodsk, Russia. Monitoring of soil nematodes in the Republic of Karelia has provided extensive data on the taxonomic diversity, abundance and community structure of nematodes in coenoses differing in geographical location, type of vegetation and scope of disturbance. The large data pool allowed us to check the effectiveness of the ecological indices derived from nematode fauna analysis (structure SI, enrichment EI, and channel CI indices) for assessing soil ecosystem conditions. It is shown that the indices reflect both changes in the soil or plant cover of natural coenoses, and the consequences of environmental disturbance caused by human activities. E.g., differences depending on the geographical location were found in CI and SI indices. In meadows CI significantly decreased from southwards (from 56 to 23). Northern coenoses exposed to extreme climatic conditions or anthropogenic transformation had a high SI index owing to a large proportion of omnivores in the nematode community structure. In natural coenoses EI and CI indices were associated with the type of vegetation. A combination of the indices enables clear differentiation between nematode communities of meadow and forest habitats. The nematode communities of meadows had higher EI and lower CI values. Low SI and high EI values were detected in disturbed habitats (urban areas, industrial zones). Agrocoenoses yielded similar values, indicating habitat instability and simplification of the soil food web. Thus, ecological indices are effective tools for assessment of the state and functioning of soil ecosystems. PRIMING OF POTATO PLANTS BY TEMPERATURE FOR ENHANCEMENT OF RESISTANCE TO COLD STRESS AND NEMATODE INVASION IN THE NORTH. Matveeva1, E., V. Lavrova1, E. Sherudilo1, M. Seppa¨nen2 and P. Palonen2. 1Institute of Biology, Karelian Research Centre of Russian Academy of Sciences, Pushkinskaya St. 11, 185910 Petrozavodsk, Karelia, Russia; 2University of Helsinki, P.O. Box 27, FIN-00014, Helsinki, Finland. Priming of plants to abiotic and biotic stresses is an important component in plant survival and normal functioning in the North, where low temperatures with sharp fluctuations during day-cycle are experienced and consequences of environment disturbances, including pest invasion, are strongly expressed. Research on priming of plants by low temperature was carried out on a parasite system ‘‘potato - potato cyst nematode Globodera rostochiensis (Wollenweber)Behrens, Ro1’’. Plants of potato genotypes derived from wild species Solanum commersonii, possessing different freezing tolerances and cultivated potato Solanum tuberosum (resistant and susceptible to nematode cultivars) were subjected to a temperature drop from 23 to 58C for 2 h at the end of the night. Afterwards half of plants were infested by potato cyst nematode, (10-20 cysts per plant). It was established that nematode infestation promoted plant growth and development and led to a slight decrease in content of photosynthetic pigments and Fv/Fm ratio in potato genotypes in comparison with non-infested plants. Plant growth and development of susceptible cultivars were inhibited and photosynthetic pigment content was slightly increased. Morphological and physiological traits of resistant cultivars were not affected under G. rostochiensis infestation. Temperature

Abstracts 203 priming diminished differences between infested and non-infested plants and enhanced plant resistance to potato cyst nematode (final nematode population was decreased on 30%). In addition temperature drop enhanced cold resistance in both potato genotypes and cultivars compared with control. NEMATODE COMMUNITIES IN MODERN SOYBEAN CROPPING SYSTEMS IN SOUTH AFRICA. Mbatyoti1, A., D. Fourie1, A. Swart2 and A.H. Mc Donald1. 1North-West University, Unit of Environmental Sciences and Management, Private Bag X6001, Potchefstroom, 2520, South Africa, [emailprotected]; 2National Collection of Nematodes, Biosystematics Division, Agricultural Research Council - Plant Protection Research Institute, Private Bag X134, Queenswood, 0121, South Africa. Local soybean (Glycine max (L.) Merr.) crops host a variety of plant-parasitic nematode species, with Meloidogyne incognita and Meloidogyne javanica being the predominant nematode pests. Information on nematode-soybean associations exists for conventional soybean crops but not for Roundup Ready soybean cultivars, which constitute more than 65% of modern plantings. Nematode surveys were thus conducted during 2012 and 2013 by sampling soil and roots at 11 localities where conventional and Roundup Ready soybean crops were grown in close proximity. Grass in natural areas adjacent to soybean fields was also sampled concurrently. Meloidogyne spp. was generally the predominant nematode pest associated with conventional and Roundup Ready soybean roots. Their population levels ranged from ca. 23000/50g roots of Roundup Ready to ca.175000/50g roots of conventional soybean cultivars. Natural grass hosted up to ca. 1800 Meloidogyne eggs and J2/50g roots. Other plant-parasitic nematodes that were recorded from root and soil samples from soybean and natural vegetation were Pratylenchus spp Helicotylenchus spp., Rotylenchus spp., Scutellonema spp., Criconemoides spp., Criconema spp. and Tylenchorhynchus spp. In terms of non-parasitic nematodes a variety of fungivores, bacterivores, omnivores and predators were recorded in the soil samples from conventional and Roundup Ready soybean, as well as natural fields. OCCURRENCE OF ROOT KNOT NEMATODES IN AFRICAN LEAFY VEGETABLES PRODUCTION SYSTEMS OF WESTERN KENYA. Mbogoh1, J.M., E. Omami1, L. Ngode1, J. Ochuodho1, P. Njira2 and W. Sunda1. 1University of Eldoret, P.O. Box 1125-30100 Eldoret, Kenya; 2Moi University, P.O. Box 1035-30100, Eldoret, Kenya. The current market supply of African leafy vegetables does not meet domestic demand due to production constrains attributed to pests especially root-knot nematodes. A survey was conducted in Busia, Kakamega and Bungoma counties to assess the occurrence of root-knot nematodes in leafy vegetable production systems. Structured questionnaires were administered to 120 selected respondents. Soil and roots were sampled for the presence of Meloidogyne spp. and the data analysed using SPSS. The number of galls/root system differed significantly at P10 spores/nematode). Fourteen P. penetrans clonal lines resulting from the effort were screened for purity by Sanger sequencing analysis of 16s rDNA and house-keeping genes. NEMATOLOGY TRAINING IN THE UNITED STATES OF AMERICA AND WHAT THE FUTURE HOLDS. Melakeberhan, H. Agricultural Nematology Laboratory, Department of Horticulture, Michigan State University, East Lansing, MI 48824, USA. Over the last 50 years or so, nematology training in the United States of America has changed in many ways. The significance of nematodes on food security (as pests and as biocontrol agents) and on understanding basic biology and ecosystem functioning that impact global climate change continue to increase, but the numbers of university and

Abstracts 205 non-university nematology positions have been declining. While some of the decline in positions may reflect changes with time, a lot more needs to be done to change the trend by emphasizing the breadth and depth of what nematology has to offer to society at-large. This will require considering lessons from the past, anticipating global change far into the future and building a pathway for nematology to prosper in the changing times. Against this background, I will discuss the following points in this presentation: i) impact of the chemical era on training and disciplinary development; ii) growth of ecology, entomophilic and molecular nematology; iii) the need for integrating basic, applied, and traditional nematology, iv) adjusting sub-disciplinary domination and views for the greater good; v) taking academic success into the stock market; and vi) understanding local to national level policies impacting training and research funding. INCREASING STUDENT PARTICIPATION TO ENSURE THE FUTURE OF NEMATOLOGY IN RAISING THE STANDARD OF LIFE IN DEVELOPING COUNTRIES STRUGGLING WITH ECOSYSTEM DEGRADATIONS. Melakeberhan, H. Agricultural Nematology Laboratory, Department of Horticulture, Michigan State University, East Lansing, MI 48824, USA. In an increasing intertwined world, balancing food security, adaptation to climate change-driven problems, and improving standard of life and overall planetary ecosystem health present many challenges for all and special opportunities for nematology. Everybody wants to live in good conditions, but people in many developing countries have to get by every day by doing what they could to survive and to feed their families, even if it results in ecosystem destruction. Where ever ecosystem destruction may happen, its impact is global. Most of the resources and the know-how to solve the problems may be centered in the North, but the solutions have to be performed mostly in the South. As E. O. Wilson, towering global figure, reminds us ‘‘. . .the most diversity of organisms and the destruction of life exist in developing countries... and those nematodes representing four out of five multi-cellular animals on the planet and so sensitive to soil ecosystem changes offer so many opportunities for, among other things, understanding environmental changes..’’ (10/22/2007, Global View Series Lecture). None of these challenges can be addressed in sustainable ways without paying attention to soil degradation, which is a longterm problem, and without building expertise and capacity on the ground. These situations present unique opportunities for nematology to build a stronger North-South bridge that will lead to addressing many pressing global issues in tenable ways. Training students and developing two-way collaborative research and outreach programs are some of the mechanisms to develop solutions to the long-term problems. These issues and the role of the internet, biggest equalizer and enabler, will be discussed. QUANTIFYING BIOLOGICAL BASIS OF SOIL HEALTH DEGRADATION IN SELECTED SUB-SAHARAN AFRICA SOIL GROUPS. Melakeberhan1, H., T. Schmidt3, Z.T.Z. Maung1, T. Teal2, S. Yildiz1, W. Kimenju4, C. Kwoseh5 and V. Saka6. 1Agricultural Nematology Laboratory, Department of Horticulture; 2Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824, USA; 3Departments of Microbiology and Immunology, and Ecology and Evolutionary Biology, University of Michigan, Ann Arbor, MI 48109, USA; 4University of Nairobi, P.O Box 30197, G.P.O, Nairobi, Kenya; 5Kwame Nkrumah University of Science and Technology, PMB, UPO, Kumasi, Ghana; 6Lilongwe University of Agriculture and Natural Resources, Lilongwe, P.O. Box 219, Malawi. While the impact of terrestrial ecosystem degradation (TED) on sub-Saharan Africa (SSA) food security and loss of biodiversity is well known, the solutions primarily center on fixing soil physiochemical deficiencies through pH and fertilizer management. When there are structural deficits of organic carbon (%C) and nitrogen (%N), both dependent on biological processes to maintain healthy levels, there is a need to integrate soil biology into soil degradation management strategies. This requires quantifying the biological community structure and function that drives the soil food web, the nutrient cycling platform. As part of building a data base for developing scalable models across soil groups (orders) (the plates up on which different cultures stump their ecosystem change footprints), we investigated nematode and microbial communities in Ferralsol, Lithosol, and Nitosol soil groups under subsistence agriculture in Ghana, Kenya and Malawi. Analysis of soil samples from disturbed (agricultural) and undisturbed (natural vegetation) landscapes in two regions about 30 km (Ghana) to 300 km (Kenya and Malawi) apart shows: i) lower biodiversity in disturbed than in natural landscapes across soil groups and countries; ii) abundance of trophic groups varied by landscape and soil group while frequency of occurrence varied by soil group; iii) similar nutrient mineralization pathways and depleted soil food web across landscapes, suggesting naturally fragile soils; and iv) microbial and nematode communities distinctly separating by soil groups and countries, suggesting difference in biological structures. Implications of the results on understanding TED and integrating soil biology into SSA’s soil degradation management strategies will be discussed. MOLECULAR AND MORPHOLOGICAL CHARACTERISATION OF MELOIDOGYNE HAPLA POPULATIONS (NEMATODA: HETERODERIDAE) FROM ROSE GREENHOUSES IN ETHIOPIA. Meressa1,3, B., H. Heuer2, H. Dehne3 and J. Hallmann1. 1Julius Ku¨hn-Institut, Federal Research Center for Cultivated Plants, Institute for Epidemiology and Pathogen Diagnostics, Toppheideweg 88, D-48161, Mu¨nster, Germany; 2Julius Ku¨hn-Institut Federal Research Center for Cultivated Plants, Institute for Epidemiology and Pathogen Diagnostics, Messeweg 11-12 D-38104, Braunschweig,

206 Journal of Nematology, Volume 46, No. 2, June 2014 Germany; 3Institute for Crop Science and Resource Conservation (INRES), Department of Phytomedicine, University of Bonn, Nußallee 9, D-53115 Bonn, Germany. Meloidogyne hapla, considered mainly as a temperate nematode species, was found for the first time parasitizing rose plants in greenhouses in Ethiopia. A survey was conducted in greenhouses randomly selected from 12 farms distributed in six districts, during 2011 and 2012. Single species nematode cultures were established for each farm by propagating single egg masses on tomato cv. Moneymaker. Identification and characterization was based on molecular sequences of the 28S D2-D3 expansion segments within the ribosomal DNA and the region located between cytochrome oxidase unit II (COII) and the 16S rRNA gene of the mitochondria (mtDNA). In addition, LM and SEM images together with morphometric measurements of the female, male and J2 were compared with M. hapla descriptions. Most morphological characters of the female, male and J2 were similar to other descriptions, with exceptions in some morphometric values. The perennial pattern conforms to characteristics of the species in most aspects, but the vulva slit was shorter than the Hawaiian population. Morphometric differences were observed in both females and J2. Female spear length and J2 body length were greater than the population from East Africa and hyaline tail length was higher than that of the Hawaiian population. Phylogenetic relationships of the Ethiopian M. hapla with other related Meloidogyne species on the basis of both mtDNA and D2-D3 expansion segment sequence analysis revealed highly supported clades containing our isolates and other published isolates from different countries. The Ethiopian isolates shared 83-100% similarity values of D2-D3 sequence with isolates from China, Germany, Moldova and the Netherlands. A NEW METHOD FOR STUDYING POPULATION GENETICS OF CYST NEMATODES BASED ON GENOTYPING BY SEQUENCING. Mimee1, B., P.Y. Ve´ronneau1, M.O. Duceppe1, M. Jean2, F. Belzile2 and G. Be´lair1. 1Agriculture and Agri-Food Canada, Horticulture Research and Development Centre, 430 boul. Gouin, St-Jean-sur-Richelieu, Quebec, J3B 3E6, Canada; 2Universite´ Laval, Faculte´ des Sciences de l’Agriculture et de l’Alimentation, De´partement de Phytologie, 2425 rue de l’Agriculture, Que´bec, Quebec, G1V 0A6, Canada. Cyst nematodes are of great economic importance and infect the roots of several major crops, including cereals, rice, soybean and potatoes. They are responsible for billions of dollars of losses each year and a number of species are regulated pests. Plant resistance is the most effective management tool, but the existence of virulent populations can reduce its efficacy. Collecting information on population genetics is thus crucial for cyst nematodes control. However, current technologies using Single Sequence Repeats (SSRs) are time-consuming and expensive because they require numerous individual larvae and PCR runs. Also, the difficulty to obtain lots of markers limits their use for trait mapping (e.g. pathotypes or avirulence genes identification). Single nucleotide polymorphisms (SNP) are abundant and widespread genetic markers across the genome that has proven to be effective for analyses with nematodes. Recent advances in next-generation sequencing and bioinformatics now allow the development of powerful methods for population genetics studies. Here, we have taken advantage of the reproduction mode of cyst nematodes, resulting in a high genetic diversity within a single cyst, to develop a simple population genetics analysis pipeline based on genotyping by sequencing. The genome-wide allele frequencies of 26 populations of golden nematodes, from eight countries and representing the five known pathotypes, have been compared. This method allowed a clear separation of the pathotypes and a fine analysis of the genetic links among global populations. In addition of being powerful, this tool has proven to be very time and cost effective and could be used for other cyst nematode species. NEMATODE DISTRIBUTION ON HORTICULTURAL CROPS IN VARAMIN, IRAN. Mohammad Deimi1, A.M. and A. Karami2. 1Young Researchers and Elite Club, Takestan Branch, Islamic Azad University, Takestan, Iran; 2Former student, Faculty of Agriculture, Islamic Azad University, Takestan, Iran. Horticulture and fruit production represents 20% of the gross domestic product in Varamin, Iran. We present the distribution of plant-parasitic nematodes in horticultural crops from 25 localities in Varamin. Six soil sub-samples/1000 m2 and 12 sub-samples/ha at 30 cm depth were analysed. In tomato, pepper, eggplant, and lettuce 14 plants/ha were taken. In the samples analysed, a predominance of Meloidogyne incognita and Meloidogyne javanica (28%) were found in horticultural crops produced under cover. Pratylenchus neglectus was found in 78% and M. incognita and M. javanica in 17% of the samples, respectively. Meloidogyne javanica, M. incognita, Ditylenchus dipsaci, P. neglectus and Pratylenchus penetrans were found in lettuce fields. The genus Longidorus was also observed. COMPATIBILITY AND VIABILITY OF STEINERNEMA AND HETERORHABDITIS SPECIES AT DIFFERENT CONCENTRATIONS OF PESTICIDES UNDER LABORATORY CONDITIONS. Mohankumar1, A., P. Sundararaj1 and S.L. Hafez2.1Department of Zoology, Bharathiar University, Coimbatore, India; 2University of Idaho, Idaho, USA. Experiments were conducted under laboratory conditions (26 ± 2 8C) to investigate the compatibility of three chemical insecticides viz., Dimethoate 30% EC (65% a.i.), Dichlorovos 76% EC (92% a.i), Monocrotophos 36% SL (68% a.i) with the infective juveniles (IJs) of newly isolated entomopathogenic nematodes (Steinernema and Heterorhabditis) from the Western Ghats of South India, India. Five concentrations (0.5, 2, 4, 6 and 8 ml/l) of the respective pesticides were replicated five times

Abstracts 207 and 1000 IJs from each genera were added individually to the solution and viability was assessed microscopically at four hour intervals for 12 hours. Data indicated that there was a significant difference in survival rate of nematodes among the concentrations and also between the genera. Steinernema spp. was more tolerant to insecticides compared to Heterorhabditis spp. Two insecticides, dichlorovos and monocrotophos caused 100% mortality at the highest concentration of 6 and 8 ml/L. In general both the genera are compatible with all three insecticides at the lower concentration of 0.5, 2 and 4 ml/L. SANTIZERS AS NEMATODE CONTROL AGENTS - COULD IT WORK? Mohlala, R., M.S. Daneel, W.P. Steyn and G. Tefu. Agricultural Research Council - Institute for Tropical and Subtropical Crops, Private Bag X11208, Nelspruit 1200, South Africa. Nematodes cause serious problems on vegetable crops resulting in up to 20% yield losses if not properly treated. Due to the phasing out of class 1-nematicides such as aldicarb and methyl bromide, the agricultural industry is in urgent need of alternative chemicals, one such avenue is sanitizers. Many of these products can kill micro-organisms very effectively in water and on hard surfaces however, very little research has been done with sanitizing agents in soil environments. After successfully testing the efficacy of Agrigold (copper compound) and Sporekill (QAC), both sanitizers, in preventing egg hatching in the laboratory, greenhouse trials were conducted to test their efficacy for nematode control and growth enhancement on tomato. A complete randomized design was used and tomato plants were infested with 2000 Meloidogyne spp. eggs per 2 l plant bag. In the QAC trial, product A (wetting agent) was included after claims of nematicidal activity. Results showed that plant growth enhancement was significantly higher for Agrigold and product A compared to the untreated control. In the Agrigold trial Meloidogyne sp. numbers per g of roots were 265 – 323 in 1/500 Agrigold compared to 500 for the untreated control treatment. Product A and Sporekill provided similar results. Although growth was enhanced with all the products tested nematode numbers often were higher due to a better root system. It is clear that these products enhance plant growth but do not seem to be very effective in nematode control. HOW TO CONTROL MELOIDOGYNE CHITWOODI. Molendijk, L.P.G., J.H.M. Visser and G.W. Korthals. Applied Plant Research, Wageningen UR, Edelhertweg 1, 8200 AK Lelystad, the Netherlands. Since the 1980s Meloidogyne chitwoodi is an important topic in arable farming in the Netherlands. Its quarantine status and the quality damage on important cash crops as potato, carrots, black salsify and gladiolus cause a strong demand for implementing control strategies which can prevent yield losses and spread of this species as much as possible. In cooperation with farmer organisations and scientists a nematode control strategy was developed and implemented to control M. chitwoodi with a minimum use of nematicides. In this presentation some essential elements of the nematode control strategy will be illustrated. The focus will be on the design of strategic crop rotations with non-hosts or resistant varieties based on knowledge of the field conditions. Results on the risk of introducing M. chitwoodi in different soil types with infested seed potato tubers will be presented. Finally the inventory on the occurrence of different populations of M. chitwoodi in the Netherlands will be discussed. As a result of the cooperative activities of farmers, extension services, routine sampling agencies and nematologists the awareness on M. chitwoodi increased considerably during the last decade. We are optimistic that the successful integrated approach of controlling potato cyst nematodes can be repeated for Meloidogyne species. CONSERVATION AND AUGMENTATION OF ENTOMOPATHOGENIC NEMATODES ON CITRUS FOR CONTROL OF FALSE CODLING MOTH. Moore1,2, S.D., A. Manrakhan3, M. Gilbert4, W. Kirkman1, J.-H. Daneel3, J. de Waal5 and R.-U. Ehlers6. 1Citrus Research International, PO Box 20285, Humewood 6013, South Africa; 2Department Zoology & Entomology, Rhodes University, Grahamstown, South Africa; 3Citrus Research International, PO Box 28, Nelspruit 1200, South Africa; 4Citrus Research International, PO Box 2201, Matieland 7602, South Africa; 5Dow Agrosciences, PO Box 355, Paarl 7620, South Africa; 6e-nema, Klausdorfer Str. 28-36 24223 Schwentinental, Germany. False codling moth (FCM), Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Tortricidae) is an important pest of citrus and other crops in sub-Saharan Africa. Control measures have traditionally focussed on the above-ground life stages (eggs, neonate larvae and adults) but have ignored the soil-borne stages (prepupa, pupa and eclosing adults). Several years of thorough and compelling laboratory trials with entomopathogenic nematodes (EPNs) indicated extreme susceptibility of FCM to EPNs. Conservation of naturally occurring Heterorhabditis zealandica Poinar (Rhabditida: Heterorhabditidae) through non-usage of a nematicide (cadusafos) resulted in dramatically lower fruit infestation by FCM. However, a couple of earlier surveys indicated that naturally occurring EPNs are found in a very small minority of agricultural soils. Consequently, large scale field trials targeting the soil-dwelling lifestages were conducted with relatively low doses of 10 and 20 infective juveniles (IJs)/cm2. A single application of H. bacteriophora Poinar to a citrus orchard floor reduced FCM infestation of fruit by up to 81%. However, results were generally variable. As a result, an approach using regular (monthly) applications at 10 and even 5 IJs/cm2 was investigated, with improved results. PARACTINOLAIMUS MORUS N. SP. (DORILAYMIDA: ACTINOLAIMIDAE), A NEW NEMATODE FROM VENEZUELA. Morales-Montero, P. and E. San-Blas. Instituto Venezolano de Investigaciones Cientı´ficas, Centro de Estudios Bota´nicos y Agroforestales, Laboratorio de Proteccio´n Vegetal, Av. 8 entre Calles 79 y 80, Maracaibo, edo. Zulia, Venezuela, CP 4001.

208 Journal of Nematology, Volume 46, No. 2, June 2014 Paractinolaimus morus n. sp., found associated with guava (Psidium guajava) plants were fixed in formaldehyde (5%) and mounted in glycerin for morphometrical and morphological characterization. Paractinolaimus morus n. sp. resembles Paractinolaimus vigor in general form and in ratios a, b and c. The new species differs in size and by the morula shape of the pars dilatata which is diagnostic of P. morus. Females are characterized by 1342 - 1955 mm body length, four large onchia and numerous denticles in the buccal cavity, odontostyle 20 – 26 mm long, reproductive tract amphidelphic and reflexed; both branches of equal length and vulva opening with longitudinal and transversal slits forming a cross. Tail conoid and ventrally arcuated. Males not common, comprising less than 0.5% of the population. General morphology except for reproductive system same as that of females, 8 to 9 ventromedian supplements in series and tail short and rounded. This is the first report of Paractinolaimus in Venezuela. FIRST REPORT OF DITYLENCHUS GALLAEFORMANS (TYLENCHIDA: ANGUINIDAE) INDUCING GALLS ON CLIDEMIA FENDLERI (MELASTOMATACEAE) FROM VENEZUELA. Morales-Montero1, P., S. Flores2, S. Subbotin3,4 and E. San-Blas1. 1Instituto Venezolano de Investigaciones Cientı´ficas, Centro de Estudios Bota´nicos y Agroforestales, Laboratorio de Proteccio´n Vegetal, Av. 8 entre Calles 79 y 80, Maracaibo, edo. Zulia, Venezuela, CP 4001; 2 Instituto Venezolano de Investigaciones Cientı´ficas, Centro de Ecologı´a, Laboratorio de Ecologı´a de Suelos, Km 11, Carretera Panamericana, Altos de Pipe, edo. Miranda, Venezuela, CP 1010A; 3Plant Pest Diagnostics Center, California Department of Food and Agriculture, 3294 Meadowview Road, Sacramento, CA 95832-1448, USA; 4Center of Parasitology of A.N. Severtsov Institute of Ecology and Evolution of the Russian Academy of Sciences, Leninskii Prospect 33, Moscow, 117071, Russia. Several Ditylenchus species have been reported from Central and South America, but only one species of this genus, D. dipsaci parasitizing onion and garlic, has been found in Venezuela. In 2009, a neotropical nematode Ditylenchus gallaeformans attacking several species of invasive melastomatacean weeds was found in Brazil and Costa and later the description of this species was published. The nematode induces formation of gall-like structures on the foliage and inflorescences. The host range included nine Melastomataceae species (Miconia ibaguensis, M. albicans, M. calvescens, M. coralline, M. latecrenata, M. mendoncae, Clidemia capitellata, C. hirta, and Leandra lacunosa). Ditylenchus gallaeformans was found in a tropical rainforest in the central part of Cordillera de la Costa from Venezuela as a parasite of Clidemia fendleri (Melastomataceae) causing deformations and galls on inflorescences and abaxial surface of the leaves. Morphological, morphometrical and molecular characterization of this population is provided. The PCR-D2-D3-28S-RFLP diagnostic profile for D. gallaeformans generated by six restriction enzymes is presented. Clidemia fendleri being an endemic plant of Venezuela is a new host of D. gallaeformans. This finding is a first record of this nematode in Venezuela. PINE WILT DISEASE, AND THE PINEWOOD NEMATODE: A WORLDWIDE ISSUE, A NEMATOLOGICAL CHALLENGE. Mota1,2, M. 1NemaLab/ ICAAM - Instituto de Cieˆncias Agra´rias e Ambientais Mediterraˆnicas, Universidade de E´vora, Nu´cleo da Mitra, Ap. 94, 7002-554 E´vora, Portugal; 2INIAV/Unidade Estrate´gica de Investigacxa˜o e Servicxos de Sistemas Agra´rios e Florestais e Sanidade Vegetal, Av. da Repu´blica, Quinta do Marqueˆs, 2784-159 Oeiras, Portugal. [emailprotected]. Bursaphelenchus xylophilus, the pinewood nematode (PWN), and causal agent of pine wilt disease (PWD), was detected for the first time in 1999, in Portugal, Europe. In 2008, the PWN was detected in new forest areas in the centre of the country despite efforts developed by the national forestry and quarantine authorities to control the nematode and its insect vector (Monochamus galloprovincialis). The nematode has also recently been reported to be present in Spain and on Madeira Island. Circulation of non-treated wood and wood products may explain the spread of the nematode. Control strategies have been focused on the vector by using chemical traps, by cutting down symptomatic trees, heat-treatment of lumber, and monitoring of main roads and ports through which lumber and wood products are transported, by the Portuguese authorities. The nematode constitutes a threat to the rest of Europe, if proper measures are not taken by European governments. The same applies to other world regions which offer appropriate bio-ecological conditions such as susceptible pine species, a Monochamus sp. and ideal temperatures for disease development. The issue constitutes a major challenge to Nematology in regards to nematode (and insect) bio-ecology, pathogenicity, use of molecular biology in diagnostics and detection, histopathology, etc. Many gaps in the knowledge of this complex biological system persist. The involvement of bacteria, associated with the PWN in causing pine wilt, has been claimed. New quick detection methods and understanding of the nematode population dynamics are being developed. Nematode genomics may provide some insight to better understand the pathogenic effects caused inside the plant. Pathogenicity testing of susceptible pine species is imperative. A review of recent progress is hereby presented. BIONEMATICIDE EFFECT OF SOME PLANT EXTRACTS ON MELOIDOGYNE INCOGNITA CONTROL ON TOMATO. Mousa, E.M., M.E. Mahdy and R.A. Bakr. Agricultural Botany Department, Faculty of Agriculture, Menoufia University, Egypt.

Abstracts 209 Experiments were carried out to evaluate the nematicidal effect of some plant extracts against root-knot nematodes Meloidogyne incognita in tomato. Three different ornamental plants were used i.e. Oleander (Nerium oleander), Marjoram (Origanum marjorana) and Datura (Datura stramonium). Nematode parameters were evaluated under laboratory and greenhouse conditions. Results obtained showed that plant extracts caused a great decrease of both egg hatching and larvae mortality under laboratory conditions. Tomato seedlings exposed to the treatment of plant extracts at the time of transplanting showed high reduction of both number of females and egg masses after two months of nematode inoculation. Number of galls was significantly reduced compared to the control. Concentrations of phenoloxidase and peroxidase enzymes in the infected plants were markedly increased by all treatments compared to untreated plants. Plant growth parameters were also enhanced compared to the control. VOLATILE ORGANIC COMPOUNDS IN ROOT-KNOT NEMATODE (MELOIDOGYNE SPP.)–HOST INTERACTIONS. Murungi1, L.K., D. Coyne2 and B. Torto1. 1Behavioral and Chemical Ecology Department, International Centre of Insect Physiology and Ecology, PO Box 30772-00100 Nairobi, Kenya; 2International Institute of Tropical Agriculture, P.O. Box 30772-00100, Nairobi, Kenya. Crop losses due to root-knot nematodes (Meloidogyne spp.) in African vegetable smallholder farms range between 30 and 100%. Coupled with their high levels of incidence, they constitute a major constraint to vegetable crop production. Although olfaction is known to play a key role in the host seeking process in entomopathogenic nematodes, little is known about the mechanism of attraction between Meloidogyne spp. and their hosts. The ability of the infective stage of Meloidogyne spp. nematodes to detect host-specific odors likely enables them to locate and infect suitable hosts. We hypothesize that roots of perpetual spinach emit such volatile chemical attractants. This poster highlights our exploratory research to understand and identify olfactory cues mediating host plant-nematode interaction. Assays conducted to understand the Meloidogyne spp. nematode-perpetual spinach interaction and volatile organic compounds involved are also described. The implications of our current findings in relation to Meloidogyne spp. nematode management are discussed. MANAGEMENT OF ROOT-KNOT NEMATODES-BACTERIAL WILT COMPLEX USING RESISTANT TOMATO VARIETIES IN COASTAL KENYA. Muriuki1, L.K., G.M. Kariuki1, Z.M. Kinyua2, R.K. Gathu and D.L. Coyne3. 1 Department of Agricultural Science and Technology, Kenyatta University, P.O. Box 43844-00100 Nairobi, Kenya; 2Kenya Agricultural Research Institute (KARI), P.O. Box 14733-00800 Nairobi, Kenya; 3International Institute of Tropical Agriculture (IITA), c/o icipe, Kasarani, P.O. Box 30772-00100, Nairobi, Kenya. Tomato and peppers valuable vegetable crop in Kenya are severely affected by pests and diseases, especially root-knot nematodes (Meloidogyne spp.) and bacterial wilt (Ralstonia solanacearum), which are both associated with high yield losses. The objective of this study was to evaluate tomato and pepper germplasm with varying levels of resistance to root-knot nematodes and bacterial wilt, individually and as a complex. Tomato cv. Cal J, Kilele (Local checks), Hawaii, Okistu, N-UG (AVRDC) and pepper cv. PI201232, PBC384, PP0237-7502 (AVRDC), California wonders and Commandant (Local checks) were screened at Pwani University, Kenya. Three week old seedlings were transplanted and each inoculated with Meloidogyne spp. (500 J2) and R. solanacearum 107 cfu/ml either singly or as combined inocula, and assessed against a noninoculated control, which contained sterile soil only. Level of resistance of each cultivar was rated based on plant growth and disease parameters. Significant differences were observed between treatments and the control. Reduction in growth parameters was highest when the two pathogens were inoculated together compared to when inoculated singly. California wonder was moderately susceptible compared to the other four pepper cultivars, which were moderately resistant to both pathogens. Tomato cv. Kilele and Okistu were moderately resistant and the other three cultivars were susceptible to both root knot nematodes and bacterial wilt. The study therefore shows that all pepper cultivars except California wonder pepper, and tomato cv. Kilele and Okistu can effectively be used in management of root–knot nematodes-bacterial wilt disease complex in coastal Kenya. THE YAM NEMATODE, SCUTELLONEMA BRADYS, A NEW THREAT TO POTATO. Mwamula1, A.O. and N. Viaene1, 2. 1Ghent University, Department of Biology, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium; 2Institute for Agricultural and Fisheries Research (ILVO), Plant, Crop Protection, Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium. The study investigated the possible threats of Scutellonema bradys on potato production by evaluating the host suitability and damage symptoms on five potato varieties grown in Europe and Africa. The probability of nematode entry, spread and establishment in Europe was also investigated through a pest risk analysis (PRA). Inoculation of potato with S. bradys in a pot test caused varying damage levels to the different potato varieties. The nematode infected both roots and tubers, resulting in the scaly surface appearance of tubers, brown root lesions and tuber rot. The number of nematodes recovered from roots, tubers and soil was greatest for the varieties Desiree, Spunta and Nicola (2-fold population increase), while reproduction was low on Diamant and Draga, where little or no increase was observed (reproduction factors 1.1 and 0.8, respectively). The PRA showed that the likelihood of entry of S. bradys from developing countries into the European Union (EU) is high due to

210 Journal of Nematology, Volume 46, No. 2, June 2014 importation of contaminated yam tubers, although reaching a susceptible host is less probable. Establishment of the nematode is possible on a number of hosts including yam, potato and tomato, especially in the most southern parts of the EU. Nematode spread is likely through several pathways: tubers, peels and soil from infested areas or attached to equipmentor by travelers carrying plant parts and/or soil samples. The study demonstrated that S. bradys can damage potato and highlighted the need to reduce the risk of nematode entry and spread in the European Union region, as well as in Africa. ECOLOGICAL ROLE OF BACTERIA ASSOCIATED WITH THE PINE WILT DISEASE SYSTEM. Nascimento1, F.X., C.S.L. Vicente1, P. Barbosa1, M. Espada1, P. Vieira1, K. Hasegawa3 and M. Mota1,2. 1ICAAM - Instituto de Cieˆncias Agra´rias e Ambientais Mediterraˆnicas, Departamento de Biologia, Universidade de E´vora, Nu´cleo da Mitra, Ap. 94, 7002554 E´vora, Portugal; 2INIAV/Unidade Estrate´gica de Investigacxa˜o e Servicxos de Sistemas Agra´rios e Florestais e Sanidade Vegetal, Av. da Repu´blica, Quinta do Marqueˆs, 2784-159 Oeiras, Portugal; 3Chubu University, Department of Environmental Biology, Chubu University 1200 Matsumoto, Kasugai, Aichi 487-8501, Japan. Mutualistic and beneficial relationships between nematodes and bacteria are present in nature, mostly occurring because of nutritional dependence and pathogen protection, and intrinsically related with the environment, the ecological conditions and the life stages of the nematode. Thirty-three years have passed since the first hypothesis suggesting a bacterial role in pine wilt disease that was initially thought to be caused only by the phytopathogenic nematode Bursaphelenchus xylophilus. In the late 1970s, researchers reported that bacteria associated with the pinewood nematode, B. xylophilus, could produce toxins that lead to pine wilt disease in pine seedlings. It was also suggested that a double vector system for pine wilt disease exists, where the bacteria were vectored by B. xylophilus, and B. xylophilus in turn vectored by an insect from the Monochamus spp. The specific involvement of bacteria in such complex disease is still controversial, although recently the number of studies focusing on the importance of bacteria has increased considerably. This review brings new insights on the role of bacteria in PWD. SOURCES OF RESISTANCE TO ROOT-KNOT NEMATODES IN COWPEA GERMPLASM FROM SOUTH-EASTERN AFRICA. Ndeve1, A.D., W.C. Matthews1, R.M. Chiulele2, J.R. P. Santos1, J.D. Ehlers3 and P.A. Roberts1. 1University of California, Department of Nematology, 3401 Watkins Drive, Riverside California 92521, USA; 2Eduardo Mondlane University, Dep. of Crop Production, 3453 Julius Nyerere Ave, Building 1, Maputo 257, Mozambique; 3University of California Riverside, Dep. of Botany and Plant Sciences, Riverside California 92521, USA. Cowpea (Vigna unguiculata L. Walp) is an important source of food in Africa, and an relatively inexpensive source of proteins. However, nutritional and economic benefits of this crop are limited under infestation by root-knot nematodes, Meloidogyne incognita and M. javanica. Rk-genes, sources of resistance, have been identified and introgressed into some commercial cultivars. Broad-based resistance is the most efficient strategy to circumvent root-knot nematode damages on cowpea, and the known resistance is based on a complex set of genes, Rk, Rk2 and Rk3. In a search for additional sources of broad-based resistance to RKNs, fifty-three Southeast African genotypes (land-races and accessions), comprising cowpea major gene pool 2, were screened for resistance to Meloidogyne spp. in the field, greenhouse and growth pouch tests. Data for root galling (GI), eggs per gram root (EgR) and egg masses per root system (EM), were analyzed using SAS statistical program version 9.0, following the Mixed Model Procedure. Results indicated significant (P < 0.05) differences in reaction to Root-knot nematodes. Eleven genotypes were consistently highly resistant to both species based on GI, EgW and EM parameters. The egg masses per root system parameter for M. javanica on some genotypes which was not significantly low. Genotypes FN-2-9-04, Var-3A, Var-11D, Namuesse-D, INIA-5A, Gile-k-local, Vita-7, FAEF-14-Inhaca-E, Muinana-Lawe and Massava-11 showed 50% or more reduction in galling and egg reproduction compared to the susceptible genotype, UCR779. These resistance sources are being compared to known Rk-genes in allelism tests to determine their uniqueness and value in breeding. BIOFUMIGATION WITH BRASSICA JUNCEA, RAPHANUS SATIVUS AND ERUCA SATIVA FOR THE MANAGEMENT OF FIELD POPULATIONS OF GLOBODERA PALLIDA. Ngala, B.M., P.P.J. Haydock, S. Woods and M.A. Back. Crop and Environment Sciences Department, Harper Adams University, Newport, Shropshire, TF10 8NB, United Kingdom. The potato cyst nematode, Globodera pallida is the most economically important nematode pest of potatoes in the UK, inflicting an annual cost of approximately £50 million to UK potato industry. Historically, farmers in the UK have relied on granular nematicides and synthetic fumigants to control G. pallida. Environmental concerns about the use of synthetic chemicals mean their future under reformed EU legislation is uncertain. Therefore, there is urgent need to search for sustainable replacements. Biofumigation, the suppression of soil borne pests and pathogens by biocidal compounds released when brassicaceous green tissues are disrupted, is increasingly being viewed as an effective and potential replacement to these fumigants. The present study examined Brassica juncea, Raphanus sativus and Eruca sativa on G. pallida in three field experiments conducted in Shropshire, UK. Samples of each Brassica species were analysed for their glucosinolates content just before incorporation. Brassica juncea significantly (P = 0.03) reduced G. pallida population while R. sativus prevented

Abstracts 211 further multiplication (Pf/Pi » 1.0) in experiment-1. In experiment-3, both B. juncea and R. sativus either treated with or without metconazole demonstrated significant partial and complete biofumigation on G. pallida encysted eggs. Glucosinolate concentrations varied significantly (P < 0.001) between the different plant parts (roots and shoots) as well as time of cultivation. A close correlation was observed between glucosinolate concentrations and mortality of G. pallida encysted eggs for summer cultivated brassicas. The results have demonstrated that B. juncea and R. sativus can play an important role in PCN management, particularly if they are included in an integrated pest management scheme. DISTINCT PATTERNS OF PLANT GENE EXPRESSION REVEAL THE BASIS FOR AGRONOMIC FLEXIBILITY IN M. HAPLA. Nielsen1, D., V.M. Williamson2, P. DiGennaro3, and D. McK. Bird1, 3. 1Bioinformatics Research Center, NCSU, Raleigh, North Carolina 27695, USA; 2Department of Nematology, UC Davis, Davis, California 95616, USA; 3 Department of Plant Pathology, NCSU, Raleigh, North Carolina 27695, USA. Field isolates of Meloidogyne hapla differ in their pathogenicity on particular crops, presumably as a consequence of genetic variation in the effector repertoire between nematode strains. To explore how genetic diversity in the pathogen influences host gene expression, we developed an extensive tool-kit for M. hapla, including whole genome sequence from 3 inbred, independent wild isolates, and a linkage map based on analysis of F2 progeny from a cross between strains LM and VW9. These M. hapla strains differ in key parasitic attributes, including resistance-breaking, gall size, number of feeding sites, and fecundity. We performed cross-species expression QTL (eQTL) analysis by mapping 2 billion ESTs, revealing nematode loci responsible for the changes in host gene expression. This eQTL approach quantifies the expression of all host and parasite genes, while simultaneously reporting exomic SNPs in M. hapla which we used for genotyping and mapping. Based on analysis of 98 F2 lines from the LM x VW9 cross, we have identified 127 plant genes whose expression is influenced by allelic variation at one or more parasite loci. We found that plant transcription factors were statistically overrepresented, including many that are highly expressed in organs other than roots. This suggests that nematodes are usurping host signalling and that distinct strains activate different pathways. Remarkably, the M. hapla genome has tightly defined loci that influence expression of large numbers of plant genes. Reassuringly, although most of the nematode genes within a QTL are newly discovered, some have been previously implicated in host-parasite signalling. THE EFFECT OF INCREASED TEMPERATURES ON POPULATIONS OF POTATO CYST NEMATODES. Niere, B. Julius Ku¨hn-Institut, Institute for National and International Plant Health, Messeweg 11/12, 38104 Braunschweig, Germany. Temperature has a great influence on the development of nematodes. At higher temperatures, nematode population in general are expected to develop faster or have more generations. This will lead to higher final population densities and consequently will have a negative impact on crop yield. During the next decades projections of climate change for northwestern Europe predict a mean air temperature increase by +28C by 2050 and up to +48C by 2100. In this region, potatoes is an important crop and production in some areas is affected by the potato cyst nematodes. The effect of an increase in mean temperature on the development of different populations of Globodera pallida and Globodera rostochiensis was studied in climate chamber experiments using a standard and an increased temperature setting. The mean temperature in the climate chambers differed on average 38C over the course of the experiments. In these experiments, potato genotype was added as an additional factor. The experiments followed a protocol for resistance testing of potato cultivars. Nematode multiplication rates on the varieties De´sire´e, Hansa, Amanda, Innovator, Aveka, Pallina, Eurobona and Seresta were determined and the relative susceptibility of these varieties towards Ro1, Pa2 and Pa3 populations was assessed under different temperature conditions. Although both potato cyst nematodes species are reported to have different temperature requirements, all populations developed well at higher temperatures. Results will be presented and discussed in the light of expected temperature changes in the future. AN ASSESSMENT OF CURRENT EUROPEAN UNION LEGISLATION ON THE CONTROL OF POTATO CYST NEMATODES. Niere, B. and E. Pfeilstetter. Julius Ku¨hn-Institut, Institute for National and International Plant Health, Messeweg 11/12, 38104 Braunschweig, Germany. Potato cyst nematodes, Globodera pallida and G. rostochiensis, are harmful organisms of potato and quarantine regulations apply in most countries. In the European Union (EU), measures are specified in the ‘‘Plant Health Directive’’ (2000/ 29/EC) and the ‘‘Control Directive’’ (2007/33/EC). Both Directives focus on regulating pathways for the introduction and spread of potato cyst nematodes. The ‘‘Control Directive’’ specifies the measures that need to be taken in order to determine their distribution, to prevent their spread and to control them. New elements such as an annual survey to determine their distribution were introduced. Other measures, like the use of resistant varieties have been specified to address the problem of virulence variation in potato cyst nematodes. Because of the EU restrictions on chemical use, resistant varieties are considered one of the most important control measures. Overall, the new EU legislation has brought a higher level of harmonization in official measures. Additional pathways for the spread of nematodes have been addressed. Now, processors of potatoes need to have officially approved waste disposal procedures for, e.g., residual soils. Since the Control Directive only addresses ‘‘European populations’’ of potato cyst nematodes, the provisions of the Plant Health Directive needed to be

212 Journal of Nematology, Volume 46, No. 2, June 2014 updated to align with the requirements of the Control Directive and to maintain the level of protection against the introduction of virulent populations from South America threatening potato production in the EU. The phytosanitary measures and recent changes will be presented and an assessment given on their relevance in controlling these pests. CONTROL OF PLUM SAWFLIES (HOPLOCAMPA MINUTA AND HOPLOCAMPA FLAVA) BY ENTOMOPATHOGENIC NEMATODES. Njezˇic1, B. and R-U. Ehlers2. 1University of Banja Luka, Faculty of Agriculture, Bulevar vojvode Petra Bojovica 1A, 78000 Banja Luka, Bosnia and Herzegovina; 2e-nema GmbH, Klausdorfer Strasse 28-36, 24223 Schwentinental, Germany. Plum sawflies (Hoplocampa minuta and Hoplocampa flava) are among the most important pests of European plum (Prunus domestica, L.) worldwide. In organic production, no control measures are available. In conventional production synthetic insecticides are available, but application is at petal fall when bees can still be present in an orchard. Since sawflies spend part of their life cycle in the soil the use of entomopathogenic nematodes (EPN) to control soil dwelling stages was considered. Efficacy of three species of entomopathogenic nematodes Steinernema feltiae, Steinernema carpocapsae and Heterorhabditis bacteriophora were tested against plum sawflies larvae and adults. EPN efficacy against larvae was tested in laboratory and field conditions. Laboratory assays were performed against 1, 10, 20 and 45 days old larvae. Mortality of 1 day old larvae was 92-100 %, whereas no mortality was observed for older larvae. In a field trial an area under the plum trees´ canopies was treated with nematodes, just before first larval fall to the soil was anticipated. The following year, before adult emergence from the soil, the treated areas were covered by insect proof nets. White sticky plates were placed under the net to catch emerging adult sawflies. In a separate trial adult plum sawflies were targeted. Trees were completely covered by insect proof nets. Nematodes were applied on the soil surface before adult insect emergence was anticipated. A reduction of 90-98 % in fruit infestations was recorded in treatments with nematodes compared to non-treated control plots. This is the first report of successful control of plum sawflies with EPN. SUSCEPTIBILITY OF BLUEBERRY CULTIVARS TO REPLANT DISEASE ASSOCIATED WITH CRICONEMOIDES ORNATUM. Noe, J.P., G.B. Jagdale, W.T. Holladay and P.M. Brannen. University of Georgia, Athens, Georgia 30602, USA. Blueberry (Vaccinium spp.) replant disease (BRD) is characterized by poor growth, yellowing, stunting, and severely reduced yields in replanted areas, and its incidence is correlated with high numbers of Criconemoides ornatum. We evaluated five cultivars of each of Rabbiteye (Brightwell, Ochlocknee, Powder Blue, Premiere, Vernon) and southern highbush (Emerald, Farthing, Rebel, Star, Legacy) types for their resistance/tolerance to BRD in two fields in Appling and Bacon Counties, Georgia. Six replicate plots of each cultivar (n = 4 plants) were arranged in a randomizedcomplete-block design and planted in 2011. Nematode populations in each plot were assessed in May and October 2013. Plant vigor, plant volume, and percent plant survival were recorded in October 2013. Criconemoides ornatum population densities increased between May and October for all cultivars, but increases were greatest for highbush cultivars. Population densities of M. ornatum were higher and plant growth parameters, including plant survival, were lower on highbush than on rabbiteye cultivars. A combined-site comparison showed that highbush had 5-6 times higher population densities of C. ornatum and 4-17 times lower plant growth parameters than rabbiteye cultivars, suggesting that blueberry replant disease is more severe on southern highbush. Differences were observed in plant growth and survival among the five southern highbush cultivars. Opportunities may exist within germplasm to improve tolerance to blueberry replant disease through cultivar development. CURRENT NEMATICIDE OPTIONS AND THEIR INTEGRATION WITH IPM STRATEGIES. Noling, J.W. University of Florida, IFAS, Citrus Research & Education Center, Lake Alfred, Florida 33850, USA. Chemical management options have undergone considerable change in the past decade due mainly to increasing regulatory scrutiny and labelled use restrictions. To overcome some of these challenges and to better target application, variable-rate application systems for all types of nematicides have been developed to prescriptively apply nematicides in a site-specific, variable-rate manner. New chemical products, both under development and those which have completed government registration, illustrates the new emphasis on safer, reduced toxicity biorational nematicidal products such as seed treatments which reduce human and environment impacts by avoiding direct application to soil. The recent loss of methyl bromide has contributed to the current coformulated use of different soil fumigants to expand the spectrum of soil pest control for many high value fruit and vegetable crops. Fumigants like 1, 3-Dichloropropene and MITC generating products continue to play an essential role for nematode control in a variety of lower value vegetable and agronomic crops. As soil fumigants become more and more regulated and their use restricted, a variety of different approaches to soil disinfestation are being considered. These include methods and timings of chemical application, emission reduction technologies, as well as integration of other pest management techniques, particularly for weeds which serve as alternative hosts to nematodes. Successful adoption of any nematode management options will depend on cost, return, ease of use, and effectiveness of the different strategies.

Abstracts 213 SORPTION OF FLUENSULFONE, A POTENTIAL NEMATICIDE FOR CONTROL OF THE POTATO CYST NEMATODE, GLOBODERA PALLIDA. Norshie, P.M., I. Grove and M.A. Back. Nematology and Entomology Group, Crop and Environment Research Centre, Harper Adams University, Newport, Shropshire, TF10 8NB, UK. Two laboratory batch experiments determined sorption of fluensulfone (a new nematicide of the fluoroalkenyl group) by six arable soils from potato fields in England. The experiments studied the sorption of the technical grade and a 15% granular formulation and were part of a larger study to evaluate fluensulfone efficacy for control of the potato cyst nematode, Globodera pallida. In Experiment 1, sorption as a function of soil type was determined as per Freundlich sorption constant KF obtained from isotherms over four initial concentrations (1.25, 2.5, 5.0 and 7.5 mg L-1). Experiment 2 compared the kinetics and extent of sorption of the technical grade and the granular product in order to determine effects of the formulation on sorption. The results showed that both soil type and form of fluensulfone significantly influenced the extent of sorption. However, sorption was low across the soils. In experiment 1, the KF and sorption normalised to soil organic carbon KFOC, ranged from 0.72 – 1.72 mg Kg-1 soil and 63.9 – 114.6 mL Kg-1 soil respectively. In experiment 2, the formulation did not influence the kinetics of sorption but decreased KD by four times, and this was ascribed to limited availability of fluensulfone in solution for uptake. Both experiments identified soil organic matter as the soil factor to have influenced fluensulfone sorption. It is suggested that sorption may not limit availability of fluensulfone in the soil solution for nematicidal activities and that the nematicide could be used for control of G. pallida in a wide range of soils. INTERACTION BETWEEN THE NEMATICIDE NIMITZ AND HERBICIDES USED IN SUGARCANE PRODUCTION AND PRATYLENCHUS ZEAE. Novaretti1, W.R.T., E. Benetti2, F.M.L. Silva2, R.J.F. Duarte2 and A.M. Reis3. 1ANNA Laborato´rio – Piracicaba, Sa˜o Paulo, Brasil; 2Milenia Agrocieˆncias – Londrina, Parana´, Brasil; 3Usina Diana – Avanhandava, Sa˜o Paulo, Brasil. The occurrence of negative interactions in sugarcane resulting from the joint application of carbamates nematicides and herbicides, which have substituted ureas in its chemical composition, has been demonstrated in several research studies, resulting in typical symptoms of phytotoxicity. The typical symptoms of phytotoxicity were lleaves that became yellowed and dry. The symproms may vary from moderate to intense.. The objective of this experiment was to evaluate the possible negative interaction among Nimitz nematicide and the different herbicides treatments used at sugarcane planting. Theexperimental design was a randomized block with split plots in a field naturally infested by Pratylenchus zeae, in a level considered medium for the crop. The treatments consisted of Tebutiurom 500 SC at 2.4 L / ha; Metribuzim 480 SC at 4.0 L / ha; Hexaron 600 WG (Diuron 468 + Hexazinone 132) at 2.0 kg / ha, Tebutiurom 500 SC at 2.4 L / ha plus Diuron 800 WG at 2.0 kg / ha and hand weeded. The sub-plots treatments were: control without nematicide, Nimitz 2.0 and 4.0 L / ha and Carbofuran 350 SC at 7.0 L / ha. All nematicide treatments significantly reduced the population numbers of P. zeae, with percentage reduction ranging from 36.67 to 98.64%, depending on the treatment and dosage applied. Regarding productivity, the values obtained at harvest did not show negative interactions among the Nimitz product and the herbicides. The control of nematodes leads to production increases of more than twenty tons of sugarcane per hectare. AN AUDIT AND HOST STATUS ASSESSMENTS OF PLANT-PARASITIC NEMATODES AND WEEDS IN SUBSISTENCE AGRICULTURE, WITH REFERENCE TO MELOIDOGYNE SPECIES. Ntidi1, 2, K.N., L. Bronkhorst1, H. Fourie2 and A.H. McDonald2. 1Agricultural Research Council-Grain Crops Institute (ARC-GCI), Private Bag X1251, Potchefstroom 2520, South Africa; 2North-West University, School of Environmental Sciences and Development, Private Bag X6001, Potchefstroom 2520, South Africa. Plant-parasitic nematodes not only infect and damage agricultural crops. A variety of weed species serves as reservoirs for nematode pests and also competes with crop plants for the available soil water and nutrients. A nematode survey was conducted on 67 localities in the nine provinces of South Africa to establish which nematode genera and species are predominant on the economically important weed species in small-scale agricultural areas. Fourteen weed species were, furthermore, screened for their host suitability to Meloidogyne javanica and Meloidogyne incognita race 2, respectively, in two independent greenhouse trials. Each seedling was inoculated with ± 5 000 root-knot nematode eggs and second-stage larvae (J2) 14 days after planting and nematode evaluations were done 56 days later. Twenty-nine plant-parasitic nematode species belonging to 15 genera, as well as two fungus-feeding nematode genera and one species were identified from soil and root samples of weeds collected from 67 localities situated in subsistence production regions of South Africa. Host status assessments indicated that three of the fourteen weed species evaluated, maintained high numbers of both M. javanica and M. incognita eggs and J2/root system and had RF-values >1, indicating susceptibility. FIELD APPLICATION OF ENTOMOPATHOGENIC NEMATODES FOR THE CONTROL OF CODLING MOTH (CYDIA POMONELLA) IN APPLE AND PEAR ORCHARDS. Odendaal, D., M.F. Addison and A.P. Malan. Department of Conservation Ecology and Entomology, Faculty of AgriSciences, University of Stellenbosch, Private Bag X1, Matieland, 7602, South Africa.

214 Journal of Nematology, Volume 46, No. 2, June 2014 Codling moth,Cydia pomonella (L.) (Lepidoptera: Tortricidae), is a key pest of apples and pears throughout South Africa. Concerns over the environmental impact, widespread dispersal of resistant populations of codling moth, and the sustainability of synthetic pesticides, have encouraged the development and use of alternative pest management technologies within an integrated pest management (IPM) strategy. Entomopathogenic nematodes (EPN) are lethal pathogens of insects and have the potential to play an important role in an IPM programme. The impact of inundative semi-field application of commercially available EPN and the effect of environmental conditions on the mortality of diapausing codling moth larvae were investigated in an apple orchard. Codling moth larvae, reared under diapausing conditions, were used to culture infective juveniles in vivo for use in the different trials. Wire-mesh cages filled with apple tree bark and 20 last-instar codling moth larvae were used as the containment method. For each treatment, eight trees, in a randomised design, were used. Cages dipped in different nematode species and concentrations were kept moist, while temperature and moisture levels were recorded for 24 h after application. Mortality by infection with EPN was confirmed by dissection, four days after application. Steinernema yirgalemense, at five different concentrations, were investigated as well as Steinernema feltiae and Heterorhabditis bacteriophora. Steinernema yirgalemense caused the highest level of mortality of codling moth larvae, however, no significant difference was found between the S. yirgalemense concentrations investigated. THE NEW NEMATICIDE NIMITZ (FLUENSULFONE): ITS ADVANTAGES AND LIMITATIONS. Oka, Y. Nematology Unit, Gilat Research Center, Agricultural Research Organization, Mobile Post Negev 85280, Israel. Fluensulfone, a new nematicide of the fluoroalkenyl group, has proven very effective in controlling root-knot nematodes, Meloidogyne spp. Advantages of fluensulfone are; low toxicity to mammals and non-target organisms, irreversible nematicidal activity against second-stage juveniles of Meloidogyne spp. in contrast to currently used nematicides, even motile juveniles that had been treated with fluensulfone and rinsed in water became immobile with time and thirdly, high systemic activity in plants. A single foliar spray of peppers with a fluensulfone solution at 3.0 g L–1 prior to inoculation reduced the galling index by 80% and the number of nematode eggs by 73 to 82% of controls. The reduction in these parameters by fluensulfone was much higher than that obtained with other nematicides at the same concentration. No evidence of enhanced biodegradation or cross-biodegradation of fluensulfone by other compounds in the laboratory. Repeated soil application of fluensulfone did not reduce the nematicidal activity of fenamiphos or cadusafos, and repeated applications of these nematicides did not lower the nematicidal activity of a subsequent application of fluensulfone. Lastly, no evidence of occurrence of fluensulfone-resistant Meloidogyne javanica in the laboratory. Limitations of fluensulfone are; phytotoxicity to some crops and no or very weak systemic activity to nematodes inside plants. Foliar spray or soil drenching to pepper after inoculation with Meloidogyne incognita did not inhibit nematode development inside roots. Further no or very weak activity to some migratory nematodes. Aphelenchoides besseyi, Aphelenchoides fragariae, Bursaphelenchus xylophilus and Ditylenchus dipsaci were highly tolerant at concentrations effective to Meloidogyne species. NEMATODE FAUNA OF A PERMANENTLY SUBMERGED PADDY FIELD. Okada1, H. and S. Niwa2. 1National Institute for Agro-Environmental Sciences, 3-1-3, Kan’nondai, Tsukuba city, Ibaraki 305-8604, Japan; 2Tomakomai Experimental Forest, Field Science Center for Northern Biosphere, Hokkaido University, Takaoka, Tomakomai 053-0035, Japan. Few studies have reported the nematode fauna in paddy fields, although these fields are recognized as biodiversity hotspots in Asia. We have reported seasonal dynamics of nematode communities in common types of paddy fields in Japan, which are temporarily submerged in a rice growing period (TSP). Here, we report the nematode fauna of a paddy field which is permanently submerged throughout a year (PSP). This is an unusual situation for rice cultivation, but we consider it a model habitat to investigate the nematode fauna of freshwater wetlands in Japan. In our previous comparison with an adjacent upland rice field, we found a unique nematode community in TSP. We also found that the major taxa of upland field, Filenchus, Acrobeloides and Rhabditidae occurred even in TSP, and that they are likely to be reproduced in a drained period. In our current study, we hypothesized that these taxa would not occur in PSP, because PSP is submerged throughout a year. We also hypothesized that PSP would have some characteristic taxa which prefer permanently submerged conditions. To examine these hypotheses, we took soil samples in May, Aug. and Nov. in 2007 and 2008 in PSP and adjacent terrestrial habitats (ATH) as a reference site. As we hypothesized, Filenchus, Acrobeloides and Rhabditidae hardly occurred in PSP, although they did occur in ATH. We found, however, other taxa occurred commonly in both PSP and ATH. We also suggest Paraplectonema and Paraphanolaimus are characteristic in PSP to prefer permanent submergence. HOST CHEMICAL COMPONENTS ATTRACT THE PHORETIC NEMATODE CAENORHABDITIS JAPONICA. Okumura1, E., R. Ozawa2, T. Yoshiga3 and Y. Takeuchi1. 1Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake, Sakyo, Kyoto 606-8502, Japan; 2Center for Ecological Research, Kyoto University, Hirano 2-509-3, Otsu, Shiga 520-2113, Japan; 3Department of Applied Biological Sciences, Saga University, Honjo 1, Saga 840-8502, Japan. The bacteriophagous nematode Caenorhabditis japonica forms a phoretic and necromenic association with the subsocial burrower bug, Parastrachia japonensis. Caenorhabditis japonica dauer larvae (DL) are specifically found on the body

Abstracts 215 surface of females. In the previous study, we found that DL are attracted to hexane extracts of the body surface components of P. japonensis in a species-specific manner. In this study we conducted GC-MS analysis of the hexane extracts from P. japonensis females and bioassays to identify the attractants in the extracts. Hexane extracts were prepared by washing P. japonensis body surface with hexane. We detected four P. japonensis-specific chemicals to be used in a chemoattraction assay. Chemoattraction assay was done on a 6-cm NGM plate with 1-cm diam. test and control spots near the edge of the plate. After 9 ml of 10, 1, 0.1, and 0.01 ml/ml diluted chemical in hexane and hexane alone was applied onto test and control spots, respectively, DL were inoculated onto the center of the plate and the number of nematodes on each spot was counted every 10 min for 1 hour. Among the chemicals tested, DL were attracted to octadecane and (E)-2-hexenal, and the most effective concentration was 1 ml/ml. Sauer larvae attracted to the chemical remained on the spot as when they did on the host hexane extracts. This data suggest that C. japonica DL may recognize both octadecane and (E)-2-hexenal derived from body surface of P. japonensis as host-specific chemicals. THE DIVERSITY OF ROOT KNOT NEMATODES AND IMPLICATIONS IN CROP PRODUCTION. Onkendi1, E., A. Mongae2 and L. Moleleki1. 1University of Pretoria, Department of Microbiology and Plant Pathology; 2McCain Foods, Pretoria, South Africa. Root knot nematodes (RKN) or Meloidogyne spp pose a significant threat to crop production globally. Yield losses, rejection of produce and transmission of RKN infected planting material are some of the factors that ultimately lead to major losses to the growers. With the on-going withdrawal of chemical nematicides from the market, it is likely that an increase in yield losses due to RKN can be anticipated. It is therefore imperative that the threat of RKN affecting various crop plants is accurately evaluated. In this study, the diversity of RKN infecting potato plants in different potato growing regions in South Africa was evaluated. To this end, the intergenic region (IGS), 28S D2-D3 expansion segments within the ribosomal DNA (rDNA) together with the region between the cytochrome oxidase subunit II (COII) and the 16SrRNA gene of the mtDNA were targeted for PCR amplification, sequencing and phylogenetic analysis. The three tropical species; M. javanica, M. incognita and M. arenaria were identified as the dominant species, occurring in almost every region sampled. Meloidogyne hapla and M. enterolobii occurred in Mpumalanga and KwaZulu–Natal respectively while M. chitwoodi was isolated from two growers located within the Free State. Implications of the growing numbers of RKN in the rhizosphere as potential synergy between RKN and secondary pathogens in crop production are discussed. A NEW SPECIES OF LAIMAPHELENCHUS (TYLENCHIDA: APHELENCHOIDIDAE) FROM SERBIA. Oro, V. Nematology Laboratory, Institute of Plant Protection and Environment, T. Drajzera 9, 11000 Belgrade, Serbia. Species of the genus Laimaphelenchus are very small nematodes, usually less than 1 mm in length. The use of a scanning electron microscope (SEM) can provide more detailed information about morphology of this group of nematodes as scanning electron microscopy may reveal structures and morphological patterns that cannot be seen with an optical microscope, which may be helpful in the taxonomy of Laimaphelenchus spp. Some morphological characters are so small that their dimensions are measured in nanometers. An undescribed species of Laimaphelenchus was found in Belgrade, Serbia. Nematodes from glycerol were transferred to ethanol, dried at critical point, gold coated and scanning electron micrographs taken with a Jeol JSM-6460 LV SEM. In addition to the characters that define the genus Laimaphelenchus the presence of cephalic papillae and inner labial papillae (sensillae) were observed. The new species has a unique tail bearing four pedunculate tubercles with 10-12 finger-like protrusions. The role of the protrusions is explained. In addition to morphological characterization, the molecular characterization of the species was based on 18S, 28S and COI molecular markers. BROAD HOST RANGE OF THE COCKROACH-GUT PARASITIC NEMATODE LEIDYNEMA APPENDICULATA. Ozawa1, S., C.S.L. Vicente1, 2, P.G. Koehler3 and K. Hasegawa1. 1Department of Environmental Biology, College of Bioscience & Biotechnology, Chubu University, Kasugai, Aichi 487-8501, Japan; 2ICAAM, Departamento de Biologia, Universidade de E´vora, Nu´cleo da Mitra, Ap. 94, 7002-554 E´vora, Portugal; 3Department of Entomology and Nematology, University of Florida, Building 970, Natural Area Dr., Gainesville, Florida 32611, USA. Normally, the relationship between host and parasite is highly specific and its balance has been established over their long evolutionary history. The thelastomatid parasitic nematodes have been reported from many species of cockroaches. Four strains of the smoky brown cockroach, Periplaneta fuliginosa, has been found to be infected with only one nematode species, Leidynema appendiculata, in Japan. However, nothing is known about parasitic nematodes in P. fuliginosa of other countries, or the host specificity of L. appendiculata. We investigated the parasitic nematode in P. fuliginosa UF that has been cultured in the University of Florida for more than 60 years, and found it still infected with only L. appendiculata. Interestingly, the Surinam cockroach, Pycnoscelus surinamensis, distributed in Japan as food for pet reptiles, and the Turkestan cockroach, Blatta lateralis, were also infected with L. appendiculata. Subsequently, L. appendiculata eggs were collected and used to artificially infect various cockroach species. We found the Japanese native cockroach Periplaneta japonica, Blattella nipponica and American cockroach Periplaneta americana were artificially infected with this nematode. Infections of L. appendiculata have been reported in several Blattaria species including lab cultured P. americana in the USA, but we

216 Journal of Nematology, Volume 46, No. 2, June 2014 haven’t seen the infection of this nematode in P. americana in Japan so far. Further studies are needed but now we hypothesize that the original host of this nematode is P. fuliginosa. We also propose that P. fuliginosa are responsible for the distribution and subsequent infection of a variety of hosts by L. appendiculata. MITOCHONDRIAL HAPLOTYPES FOR IDENTIFICATION OF TROPICAL ROOT-KNOT NEMATODE SPECIES AND LINEAGES. Pagan1, C., D. Coyne2, G. Kariuki3, R. M.D.G. Carneiro4 and V.M. Williamson1. 1Dept. of Plant Pathology, Univ. of California, Davis, USA; 2International Institute of Tropical Agriculture (IITA), c/o icipe, Kasarani, P.O. Box 30772-00100, Nairobi, Kenya; 3Dept. of Agricultural Science and Technology, Kenyatta University, Nairobi, Kenya; 4 EMBRAPA - Recursos Gene´ticos e Biotecnologia, C. P. 02372, 70849-979 Brasilia, DF, Brazil. The asexually-reproducing root-knot nematodes (RKN; Meloidogyne spp) are widespread and damaging pests in tropical and sub-tropical regions throughout the world where they are often not identified to species level. The mitochondrial genome due to its uniparental inheritance is a useful tool for comparing and identifying these closely related hybrid species. Previously developed primer pairs that together amplify two mitochondrial DNA sequences that span a non-coding spacer and part of the adjacent large subunit rDNA were used to amplify DNA from well-characterized voucher samples including 15 different RKN species. Based on amplification-product sizes and restriction enzyme digestion patterns, Meloidogyne javanica, M. enterolobii, M. incognita could be distinguished and assigned unique mitochondrial haplotypes. Additional key species, including M. hapla and other sexually reproducing species were also resolved by this procedure. Meloidogyne arenaria isolates did not group as a single haplotype, consistent with other reports of diversity within this species. DNA extracted from a juvenile, single female or egg mass can be used for this assay. Ethanol-preserved females or egg masses can also be used allowing samples to be stored and shipped for analysis. This protocol is a rational strategy for initial characterization of RKN species especially in regions where sampling has been limited such as Sub-Saharan Africa. In addition, the DNA sequence of the amplified fragments, especially of the mitochondrial rDNA, is informative regarding the relationship between species, including those not previously identified, based on maternal lineage. SYNTAXIN 31 FUNCTION IN GLYCINE MAX RESISTANCE TO THE PLANT-PARASITIC NEMATODE HETERODERA GLYCINES. Pant1, S.R., P.D. Matsye1, B.T. McNeece1, K. Sharma1, A. Krishnavajhala2, G.W. Lawrence2, V.P. Klink1. 1Department of Biological Sciences, Mississippi State University, Mississippi State, MS 39762, USA; 2Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, MS 39762, USA. A Glycine max syntaxin 31 homolog (Gm-SYP38) is expressed in Heterodera glycines-induced syncytia undergoing an incompatible interaction. The observed Gm-SYP38 expression is consistent with prior gene expression analyses that identified the alpha soluble NSF attachment protein (Gm-alpha-SNAP) resistance gene because homologs of these genes physically interact and function together in other genetic systems. Syntaxin 31 protein resides on the cis face of the Golgi apparatus. It binds alpha-SNAP-like proteins, but has no known role in resistance. Gm-alpha-SNAP overexpression induces Gm-SYP38 transcription, making it reasonable to test the function of Gm-SYP38. Overexpression of Gm-SYP38 rescues G. max[Williams 82/PI 518671], genetically rhg1-/-, by suppressing H. glycines parasitism. In contrast, Gm-SYP38 RNAi in the rhg1+/+ genotype G. max[Peking/PI 548402] increases susceptibility. Overexpression of both Gm-alpha-SNAP and Gm-SYP38 induces the transcriptional activity of the cytoplasmic receptor-like kinase BOTRYTIS INDUCED KINASE 1 (Gm-BIK1-6) which is a family of defense proteins. BIK1-like genes are known to anchor to membranes through a 5’ MGXXXS/T(R) Nmyristoylation sequence. Gm-BIK1-6 had been identified previously by RNA-seq experiments as expressed in syncytia undergoing an incompatible reaction and its overexpression rescues the resistant phenotype. In contrast, Gm-BIK1-6 RNAi increases parasitism. The analysis demonstrates a role for syntaxin 31-like genes in resistance that until now was not known. IDENTIFICATION OF RESISTANCE TO CEREAL CYST NEMATODE HETERODERA FILIPJEVI IN WINTER WHEAT GENOTYPES THROUGH ASSOCIATION MAPPING. Pariyar1, S.R., A.A. Dababat2, S. Siddique1, G. Erginbas2, A. Morgounov2, J. Leon3 and F.M.W. Grundler1. 1Institute of Crop Science and Resource Conservation, Molecular Phytomedicine, University Bonn, D-53115, Bonn, Germany; 2International Maize and Wheat Improvement Centre (CIMMYT), Ankara, Turkey; 3Institute of Crop Science and Resource Conservation, Plant Breeding, University Bonn, D-53115, Bonn, Germany. The aim of this study was to identify quantitative trait loci)/genes conferring resistance against the cereal cyst nematode Heterodera filipjevi in wheat. Cyst number per plant was determined in 161 winter wheat genotypes in two consecutive years in growth room trials. Susceptibility was very low in 1%, low in 17%, medium in 37%, high in 34% and very high in 11% of the genotypes. Four genotypes (NUDdakota, ECOnomka, KATea and LANtian 12) with very low and low susceptibility, respectively, were selected and nematode infection, development, and reproduction were analyzed in detail. The infection rate compared to highly susceptible cv. Bezostaya 1 was significantly lower and nematode development inside roots was significantly delayed in NUD, ECO, KAT and LAN at 2, 5, 10 and 15 dpi. The number of cysts per root system was also significantly lower in NUD, ECO, and LAN. However, cyst size was significantly smaller only in LAN and ECO. All

Abstracts 217 populations were genotyped with an Illumina 90K SNP iSelect wheat chip at TraitGenetics, Germany, and association between molecular markers and quantitative trait loci was analyzed. Twenty eight loci associated with H. filipjevi development were identified in which 11 of them were associated with low nematode susceptibility, while 17 were associated with very high susceptibility. In addition, six markers were located on chromosome 1A where quantitative trait loci regions associated with Heterodera avenae resistance had been detected in previous studies. Future work will focus on identifying the genes conferring resistance to H. filipjevi and revealing their mode of action. CHROMADOREAN NEMATODE PHYLOGENY REVISITED BASED ON COMPARATIVE ANALYSIS OF COMPLETE MITOCHONDRIAL GENOME SEQUENCES. Park1, J.-K., J. Kim1, S.-H. Lee1, T. Kim1 and S.A. Nadler2. 1 Program in Cell Biology and Genetics and Department of Parasitology, College of Medicine, Chungbuk National University, Cheongju 361-763, South Korea; 2Department of Entomology and Nematology, University of California, Davis, California 95616, USA. Recent phylogenetic hypotheses for chromadorean nematodes have almost exclusively been based on nuclear ribosomal DNA genes, mainly SSU rRNA. However, relationships among major chromadorean lineages based on SSU sequences have been challenged by phylogenetic hypotheses based on complete mitochondrial genomes. We revisited some phylogenetic issues for chromadorean nematodes based on comparative analyses of complete mitochondrial genomes sampled from major representatives of chromarodean infraorders. The resulting mitochondrial genome tree for nucleotide and amino acid sequence data including 12 protein-coding genes depicted the following relationships, only some of which were inconsistent with SSU phylogeny: (1) non-monophyly of nematode clade III; (2) non-monophyly of Panagrolaimonorpha, (3) the nested position of Diplogasteromorpha within the Rhabditomorpha clade, (4) non-monophyly of Tylenchomorpha (independent origins of plant parasitism within chromadorean nematodes), (5) sister relationship between Rhigonematomorpha and Ascaridomorpha, (6) monophyly of Aphelenchoidea. Phylogenetic relationships among the major chromadorean orders revealed from analysis of mtDNA sequence are generally consistent with results of mitochondrial gene order comparison, with the exception of Strongyloides stercoralis Heterorhabditis bacteriophora, and Ascaridia spp. Some relationships remain contradictory between mtDNA and nuclear rDNA trees. These conflicting relationships require reappraisal using both combined analyses of different data sets, and application of new data from additional nuclear genes. It is anticipated that additional taxon sampling for mitochondrial genome sequences will provide a wealth of information on mitochondrial genome evolution and data for developing phylogenetic hypotheses for chromadorean nematodes. RAPID DETECTION OF THE CEREAL CYST NEMATODE (HETERODERA AVENAE) USING LOOP-MEDIATED ISOTHERMAL AMPLICATION. Peng1, D., X. Xu1,2, H. Peng1 , W. He1 and D. Jiang. 1State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, People’s Republic of China; 2State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, People’s Republic of China. Cereal cyst nematode (Heterodera avenae) is the most important plant-parasitic nematode on cereal crops in the wheat producing areas of China. In this study, a rapid and highly sensitive assay and diagnostic tool for the identification of H.a avenae was developed using loop-mediated isothermal amplification (LAMP). The LAMP assay targeted RAPD fragments of H. avenae. Three LAMP primers were designed and specificity of the LAMP assay was confirmed using different nematode species including Heterodera filipjevi, Heterodera goettingiana, Heterodera elachista, Heterodera glycine, Heterodera latipons, Meloidogyne javanica and Meloidogyne arenaria. The detection threshhold of the LAMP assay was as low as 10-2 and 10-4 of single juvenile and cyst DNA, and the detection sensitivity of the LAMP method for H. avenae DNA is 100 times higher than normal PCR-based detection methods. The LAMP amplifications could be observed visually after the addition of SYBR Green I and the lateral flow dipstick.. LAMP assay could be utilised for the detection of nematodes in wheat roots infected by H. avenae. The LAMP assay developed in this study is highly effective, easy to perform and readily adaptable for the early identification and monitoring of H. aenae on diseased seedlings in the field. INTEGRATED MANAGEMENT OF CEREAL CYST NEMATODE HETERODERA AVENAE - A CASE STUDY EFFECT OF SEED-COATINGS TO CONTROL CEREAL CYST NEMATODE ON WHEAT IN CHINA. Peng1, D., R. Hao1,2, W. Huang 1, C. Liu1,2, H. Li2 and H. Li 3. 1State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, People’s Republic of China; 2Key laboratory of Monitoring and Management of Disease and Insects, Ministry of Agriculture, Nanjing Agricultural University, Nanjing 210095, People’s Republic of China; 3Grassland Science Institute of Gansu Agricultural University, Lanzhou 730070, People’s Republic of China. Cereal cyst nematodes (CCN, Heterodera avenae and H. filipjevi) are the most economically important plant-parasitic nematodes on cereal crops in China, occurring in 16 provinces. Estimated yield losses were 18%-35% in Henan, 11-18% in Hebei, and 15%-28% in Qinghai province. One generation of H. avenae is completed per growing season. Heterodera avenae is the dominant species in the wheat production areas. Control of CCN on wheat by means of six seed-coatings

218 Journal of Nematology, Volume 46, No. 2, June 2014 (Gannong seed coating I, Gannong seed coating II Gannong seed coating III, abamectin seed coating AV1, abamectin seed coating AV2 and 5.76% emamectin benzoate), were evaluated in this study. Wheat seeds were coated with the different treatments before sowing and the numbers of cysts in soil and wheat yield were evaluated after harvesting. The results showed that the numbers of cysts dropped significantly after seed-coating. The highest reduction in cyst numbers were 56.0%, 53% and 47% in the treatment by Gannong seed coating III (1:35), Gannong seed coating I (1:50) and Gannong seed coating II (1:35), respectively. The wheat yield increased by 37.6%, 19.4%, 17.9% and 17.7%, respectively for Gannon seed coating III (1:35), Gannon seed coating I (1:50), Gannon seed coating II (1:35) and Gannon seed coating I (1:35). The selfpatented Gannong seed coating III not only has the better efficacy for the control of CCN, but also has characteristics of environmental safety, lower toxicity, labor and cost saving, which is suitable for widely application in practical disease control. TRANSCRIPTIONAL ANALYSIS OF CEREAL CYST NEMATODE HETERODERA AVENAE AT THREE DIFFERENCE STAGES THOUGH RNA SEQUENCING. Peng, H., G. Wang, J. Cui, W. Huang, W. He, D. Peng. State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China. The cereal cyst nematode, Heterodera avenae is a major nematode pest on cereal crops worldwide and causes significant economic yield losses in many countries; however the molecular characterization of its plant-parasitism has been limited. In this study, the transcriptomes were sequenced from three difference stages of H. avenae including pre-parasitical J2, J4 and white female by Illumila Hiq 2000. Over 47348000, 45186000 and 45260000 reads were generated from pre-parasitical J2, J4 and white female separately. After assembling the reads, 66,962, 43,953 and 62,697 contigs with an average length of 1546, 1029 and 1068 bp were selected for further analyses, respectively. Homology searches revealed that 48.8% - 62.2% of unigenes of H. avenae were similar to known genes. 5671, 6244 and 6961 unigenes were announced separately using Gene Ontology; 1824, 2029 and 2275 unigenes were mapped in KEGG biochemical pathways. The main purpose of this study was to identify the effectors of H. avenae using different approaches. 203 clusters with similarities to effectors from other plant parasitic nematodes were identified within the data. On the other way, 2323 clusters had signal peptides but no transmembrane domain were identified, these genes that might include novel effectors. Finally, the transcriptomes were used to identify possible target genes for RNA interference (RNAi)-based control strategies. A WAX ESTER PROMOTES COLLECTIVE HOST FINDING IN THE DAUER LARVA OF PRISTIONCHUS PACIFICUS. Penkov1, S., A. Ogawa2,4, U. Schmidt3, D. Tate2, V. Zagoriy1, S. Boland1, M. Gruner3, D. Vorkel1, J-M. Verbavatz1, R. J. Sommer2, H-J. Kno¨lker3 and T.V. Kurzchalia1. 1Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany; 2Department for Evolutionary Biology, Max Planck Institute for Developmental Biology, Tu¨bingen, Germany; 3Department of Chemistry, Technische Universita¨t Dresden, Dresden, Germany; 4Laboratory for Developmental Dynamics, RIKEN Quantitative Biology Center, Kobe, Japan. Survival of nematode species depends on how successfully they disperse in their habitat and in finding a new host. Under harsh environmental conditions many free-living nematodes stop development after the second larval (J2) stage and enter into a stage called the dauer larva. For dispersal, dauer larvae of multiple nematodes have evolved a specific behaviour called nictation or waving: worms stand on their tails and wave the body allowing for attachment to larger animal vectors. Nictation of individuals and attachment to insects under laboratory conditions is also described in the nematode Pristionchus pacificus. This behavior is crucial for Pristionchus since, under natural conditions, these nematodes rely on a tight association with scarab beetles, called necromenic. The dauer larvae invade the host, wait for its natural death, and then resume development by feeding on microorganisms that grow on the host carcass. Herein we report that Pristionchus pacificus nematodes synthesize a dauer stage-specific, extremely long-chain polyunsaturated wax ester that we name nematoil. Dauer larvae secrete nematoil to their body surface as an adhesive lipid that promotes the congregation of multiple individuals into large tower-like structures called dauer towers. The formation of the dauer towers is part of a novel host finding strategy, collective nictation, a process in which the whole dauer towers wave, presumably to maximize the chance of attaching to a new host organism. TESTING THE MONOPHYLY OF THE SUPERFAMILY DOLICHODOROIDEA SENSU SIDDIQI, 2000 (TYLENCHIDA): EVIDENCE FROM THREE RIBOSOMAL GENES. Pereira1 , T., L. Caixeta2, M. Mundo-Ocampo3, J. Cares2 and J. Baldwin1. 1Department of Nematology, University of California, Riverside, California 92521, USA; 2Department of Phytopathology, University of Brasilia, DF 70743-060, Brazil; 3CIIDIR-IPN, Unidad Sinaloa, Mexico. The superfamily Dolichodoroidea sensu Siddiqi, 2000 is comprised of four families, 30 genera, and about 450 species. Within Dolichodoroidea, the family Telotylenchidae is the most representative with four subfamilies, 20 genera, and having Tylenchorhynchus as the most specious and widely distributed genus. The extant classification proposed by Siddiqi (2000) is exclusively morphological-based and current molecular phylogenies are based upon a few representatives. Specimens were collected worldwide including Brazil, Argentina, Mexico, USA, Vietnam, and Iran. Morphological identification is based on

Abstracts 219 LM and SEM. The monophyly of the superfamily Dolichodoroidea is tested based on three ribosomal genes (18S, ITS, and 28S). Preliminary results showed the non-monophyly of the superfamily Dolichodoroidea regardless of the gene used. For instance, the genus Tylenchorhynchus showed a strongly supported clade with 16 different sequences. However, some Tylenchorhynchus sequences also grouped with other genera (e.g. Quinisulcius, Macrotrophurus, and Dolichodorus). The subfamily Merliniinae was monophyletic, however not related with Telotylenchinae. Instead, this group was placed as a sister taxon of Psilenchidae. Phylogenetic trees based on ITS and 28S genes showed some similarities: a large Tylenchorhynchus clade, a sister relationship between Merliniinae and Psilenchidae, the monophyly of Dolichodorus and Belonolaimus, and the polyphyly of Tylenchorhynchus. On the other hand, the 18S gene had no resolution to resolve most of the relationships within Dolichodoroidea. These results showed partial agreement with the classification provided by Siddiqi (2000). Nevertheless, the relationships at lower taxonomic levels need to be further evaluated. Such findings will be further improved by including additional taxa and genes. INTRA- AND INTER-SPECIFIC GENETIC VARIATION IN THE GENUS CEPHALENCHUS: IMPLICATIONS FOR SPECIES DELIMITATION. Pereira1, T., K. Chang1, M. Mundo-Ocampo2 and J. Baldwin1. 1Department of Nematology, University of California, Riverside, CA 92521, USA; 2CIIDIR-IPN, Unidad Sinaloa, Sinaloa, Mexico. Molecular phylogenetics of Tylenchina sensu De Ley and Blaxter, have been heavily biased to economically important groups of plant parasites. Conversely tylenchs such as fungal feeders (i.e. Tylenchidae), not directly associated with crop losses have been under represented in phylogenies. Broader taxon sampling of these groups is crucial to more fully understanding the evolution of plant parasitism in nematodes. Herein, molecular and morphological data from eight Cephalenchus populations from Brazil, USA, Mexico, Canada, and Vietnam were used to evaluate the phylogenetic position of the ¨ rley, 1880. Using a multilocus approach genus, traditionally (i.e. based on morphology) classified within the Tylenchidae O (ribosomal: 18S, ITS, 28S; mitochondrial: COI), levels of genetic variation were compared within and between these populations and discussed with respect to implications for species delimitation. Preliminary results demonstrated diverse levels of intraspecific variation among populations (0.1% to 10.5% for 28S and 2.1% to 15% for ITS). Generally, for both genes, interspecific genetic variation was higher than intraspecific variation; however, one population from Brazil (BRZ-01) showed high levels of intraspecific variation, comparable to those found between geographically divergent populations. Findings suggest that sequence comparison (% similarity) may be inadequate to define species affinities but rather hypothesis of monophyly must be tested to assess the validity of species boundaries as well as to recognise natural groups at higher taxonomic ranks. SELECTION FOR RESISTANCE TO MELOIDOGYNE SPP. IN COFFEA ARABICA ACCESSIONS UNDER GREENHOUSE AND FIELD CONDITIONS. Peres1, A.C.J., A.S. Jorge Junior2, J.K.A. Mattos1, V.R. Correa2,3, S.M.L. Salgado4 and R.M.D.G. Carneiro2. 1Universidade de Brası´lia, Faculdade de Agronomia e Medicina Veterina´ria, 70.910970 Brası´lia, DF, Brazil; 2Embrapa Recursos Gene´ticos e Biotecnologia, C.P. 02372, 70849-970 Brası´lia, DF, Brazil; 3 Universidade de Brası´lia, Dept. de Fitopatologia, 70910-900 Brası´lia, DF, Brazil; 4EPAMIG, 36.570-000 Vicxosa, MG, Brazil. Root-knot nematodes are amongst the most economically important plant-parasitic nematodes infecting coffee in Brazil, particularly, Meloidogyne paranaensis and Meloidogyne incognita. The objective of this study was to identify coffee materials obtained from the Coffee Germplasm Bank (EPAMIG), resistant to M. paranaensis and M. incognita race 1, which have shown promising production under field conditions. Seedlings of 18 different accessions, a resistant (cv. IPR-100) and a susceptible (cv. Mundo Novo) cultivar, were inoculated under greenhouse conditions. Nematode reproduction factor was used to infer coffee responses (resistance/susceptibility) to nematode infection. Five accessions from crossing between Catuaı´ Vermelho x Amphillo MR 2161, one accession from Catuaı´ Vermelho x Amphillo MR 2474, two accessions from Timor Hybrid UFV 408-01 and the resistant control cv. IPR-100 were resistant to M. incognita race 1. Four accessions from crossing Catuaı´ Vermelho x Amphillo MR 2161, one accession from Timor Hybrid UFV 408-01 and the cv. IPR-100 were resistant to M. paranaensis. Field testing with parental genotypes showed that plants originated from Catuaı´ Vermelho x Amphillo MR 2161 or Amphillo x H. Natural MR 36-349 progenies were resistant and highly productive, while plants that originated from progenies Catuaı´ Vermelho x Amphillo MR 2474 , Timor Hybrid UFV 408-01 (treatment 8) and Catuaı´ Vermelho x Amphillo MR 2161 were resistant and with good yield. Overall, materials from treatments 3, 5, 8, 14, 19 and 20 were resistant to both M. paranaensis and M. incognita race 1 under greenhouse/field conditions and showed promising agronomic traits to be used in coffee breeding programs. SURVEY AND REVISION OF NEMATODES INHABITING NORTH AMERICAN MILLIPEDS. Phillips, G. and E.C. Bernard. University of Tennessee, 2505 E. J. Chapman Drive, 370 Plant Biotechnology Building, Knoxville, TN 379964560 USA. Nematodes that parasitize millipeds are commensal kleptoparasites. Thelastomatidae and Rhigonematidae are two common families that inhabit the intestine of North American millipeds. In North America, 16 nematode species have been

220 Journal of Nematology, Volume 46, No. 2, June 2014 recognized from millipede fauna. The primary goal of this research is a comprehensive taxonomic analysis of these nematodes and their specific host-parasite relationships with millipeds. In dissections undertaken so far, 269 millipeds from 6 orders (15 species) have yielded 0–417 nematodes. Thelastomatids are typically encountered in the distal hindgut while rhigonematids are mostly observed in the proximal hindgut at the pyloric region. Polydesmid millipeds usually contain the most nematodes. In these millipeds, the nematodes may be so concentrated in the pyloric area that the intestine appears completely occluded, yet milliped movement and health appear unaffected. Spirobolid millipeds, such as Narceus americanus, yield the largest rhigonematid nematodes, some reaching 10.2 mm in length. Rhigonematids typically are more numerous but thelastomatids are more diverse, with at least ten species found so far. Some millipeds, such as some parajulids and platydesmids, are devoid of nematodes while others are heavily infested. Nematodes found in the milliped order Spirostreptida (Choctella cumminsi and Orthoporus ornatus) clearly show that thelastomatids are more prevalent than rhigonematids. At least two nematode taxa have been collected that are not yet classifiable to family. Milliped width and nematode presence appear to be correlated; millipeds less than 2 mm wide in cross-section do not contain nematodes. NEMATODE INFESTATION, AFLATOXIN CONTAMINATION AND ASPERGILLUS SPP. BIOMASS IN GROUNDNUTS FROM SMALL-HOLDER FARMERS IN KWAZULU-NATAL. Phokane1, S., D. Fourie2, E. Ncube1 and B.C. Flett1,2. 1Agricultural Research Council-Grain Crops Institute, Private Bag X1251, Potchefstroom, 2520, South Africa; 2Unit of Environmental Sciences and Management, North-West University, Private Bag X6001, Potchefstroom, 2520, South Africa. Groundnut (Arachis hypogaea L.) is an important food source due to the high essential fatty acid, protein and energy content of the kernels. Nematodes pests are often detected in groundnuts and often create infection sites for fungal pathogens. The aim of this study was to quantify nematode infestation, aflatoxin contamination and Aspergillus spp. biomass in groundnut kernels from small-holder farmers in northern KwaZulu-Natal, South Africa. Thirty kernel samples were collected after harvest from farmer’s fields at five localities. Nematodes were extracted using the Bearman-tray method, while aflatoxin contamination was analysed using the LC-MS technique. Aflatoxin producing Aspergillus spp. were quantified using realtime PCR. Four plant-parasitic nematode genera were identified, namely: Ditylenchus africanus, Helicotylenchus spp., Meloidogyne spp. and Pratylenchus spp. Hull and kernel samples from Manguzi and Mbazwana had the highest plantparasitic nematode infestation levels, while no nematodes were present in either hulls or kernels obtained from Vryheid. Ditylenchus africanus and Meloidogyne spp. had the highest frequency of occurrence in both hull and kernel samples that were collected at Manguzi and Mbazwana. Population levels of the latter genera were 12 and 22 times higher in the hulls compared to those in the kernels, respectively. Pratylenchus spp. were present in hulls from all localities, while Helicotylenchus spp. were present in very low numbers in kernels only from Manguzi. Aflatoxin contamination was not detected in any of the groundnut samples; however, low levels of aflatoxin-producing Aspergillus spp. biomass, ranging from 0.0068 to 11.46ng/ml, were detected in samples from Manguzi and Mbazwana. IMPLEMENTATION OF A HIGH THROUGHPUT REAL-TIME PCR METHOD FOR POTATO CYST NEMATODE DETECTION IN SCOTLAND. Pickup, J., A. Reid, Y. Cole and C. Longley. Science and Advice for Scottish Agriculture, Roddinglaw Road, Edinburgh EH12 9FJ, UK. In the three years since 2010, when the European Directive 2007/33/EC on the control of PCN came into force, implementation of the new statutory measures, particularly the new harmonized soil sampling rate, we have seen a three fold increase in the number of samples processed at SASA. To manage this increase with the same staffing resources, SASA has had to overhaul the traditional diagnostic methods we used previously (i.e. cyst extraction by Fenwick can and diagnosis by visual examination). A new system has been developed for the collection and processing of soil samples, extraction of DNA from float material and detection of PCN by high throughput real-time PCR. The whole process is controlled and monitored by a bespoke software package (Seed Potato Unified Data System or SPUDS). Approximately 18,000 samples are analysed each year, with the incidence of PCN positive samples remaining similar to that determined by visual examination. As the number of samples taken per field has increased, the area of land recorded as infested with PCN has consequently increased by over two-fold. NEMATODES AS A THREAT TO THE FRENCH FRY PROCESSING INDUSTRY. Pieterse, B.J. McCain Foods (SA), P.O. Box 1023, Delmas, 2210, South Africa. McCain Foods (SA) is committed to supply safe and healthy food to customers and strives towards limiting the impact of their operations on the environment. Most of the potatoes needed for processing are supplied by local potato growers. McCain Foods (SA) also grows approximately 400 ha of potatoes. The french fry industry in South Africa is very competitive. Finished products have to be produced and processed at a cost lower than that of imported products. This implies limiting the cost of production on-farm and also keeping cost of processing as low as possible. Pesticides represent one of the major costs of production. Apart from the cost factor McCain Foods (SA) is also aware of environmental impacts and strive towards application of less active ingredients per ha. Efficiency in the factory is determined by quality factors such as dry

Abstracts 221 matter content and tuber defects such as nematode damage. Equally important is the yield obtained in terms of the finished product. Processing nematode invested tubers means that the factory process has to be adjusted to peel more tuber tissue than normal. With severe infection and deeper tissue damage it may be necessary to use mechanical peeling instead of steam peeling and so resulting in even less yield as well as increased cost of production. The processing industry, french fry production in this case, urgently needs improved nematode control solutions to be competitive in the world and in so doing contribute to the economy of the country by supplying safe and cost effective products to the consumer. ENDOPHYTES, TISSUE CULTURE PLANTS, ORGANIC MATTER AND FALLOW: THE BEST STRATEGY TO MANAGE PLANT PARASITIC NEMATODES IN BANANAS. Pocasangre, L.E. Tropical Crops, EARTH, University, Costa Rica. It is well established that after black Sigatoka, plant parasitic nematodes are the main constrain in banana production in the tropic and subtropics. The most important nematodes associated with banana are: Radopholus similils, Helicotylenchus multicinctus and Meloidogyne incognita. Currently the conventional way to manage plant parasitic nematodes in bananas is with two or three applications of nematicides, which can cost between 300-450 USD/ hectare/ year. However even with this treatment the majority of banana plantation still have plant parasitic nematodes population of over 10000 plant parasitic nematodes/100 g roots.. On the other hand, there is a increased pressure from consumers and supermarkets to eliminate or reduce the application of nematicides due to the contamination of groundwater, rivers, oceans and most important the risk of health problems for field workers. Wholefood supermarkets are buying sustainable banana fruit produced in EARTH University. In the new ratin policy or standard, nematicides like vydate and herbicides are forbidden. Field trials conducted in the commercial banana plantation of EARTH University had shown that the use of tissue vulture plants (TCP) protected with endophytes as well as adding 6 tonnes of compost base on rachis reduced the population of plant parasitic nematodes as well as improved the vigor of the plantation. We also demonstrated three years of nearly eradicate R. similis and populations of less than 1000 nematodes /100g of roots were recorded. Research to establish protocols for the use of tissue culture plants, endophytes, compost and fallow as control methods for plant parasitic nematodes in banana production is needed. POTENTIAL USES OF TRADITIONAL MEDICINAL PLANTS AS ALTERNATIVE CROPS IN SOIL WITH HIGH LEVELS OF MELOIDOGYNE SPECIES. Pofu1, K.M. and P.W. Mashela2. 1Agricultural Research Council, Vegetable and Ornamental Plant Institute, Private Bag X293, Pretoria, 0001, South Africa; 2School of Agricultural and Environmental Sciences, University of Limpopo, Private Bag X1106, Sovenga 0727, South Africa. Momordica balsamina (Cucurbitaceae) has various traditional medicinal uses in South Africa. Preliminary field observations when M. balsamina plants were inter-planted with tomato (Solanum lycopersicum) plants for repelling whiteflies, suggested that the former were not infected by Meloidogyne species, while the latter were heavily infected. Two separate experiments were therefore, conducted to determine the nematode-resistant potential of M. balsamina to Meloidogyne incognita and Meloidogyne javanica. Seven treatments, viz. 0, 250, 650, 1 050, 1 450, 1 850 and 2 250 eggs and second-stage juveniles of each nematode species were arranged in a randomized complete block design, with 12 replications. At 56 days after inoculation, the reproductive factor values at all levels of inoculation for each species were less than one, while nematode infection had no effect on yield components of M. balsamina. In plant-parasitic nematology, when RF values are less than unity and nematode infection has no effect on growth of the test plant, the nematode-plant relation is resistant. In conclusion, M. balsamina was resistant to M. incognita and M. javanica and could therefore be used as an alternative crop for managing population densities in smallholder farming systems. MANAGING MELOIDOGYNE SPECIES IN WATERMELON PRODUCTION USING INDIGENOUS NEMATODERESISTANT ROOTSTOCKS IN SOUTH AFRICA. Pofu1, K.M. and P.W. Mashela2. 1Agricultural Research Council, Vegetable and Ornamental Plant Institute, Private Bag X293, Pretoria, 0001, South Africa; 2School of Agricultural and Environmental Sciences, University of Limpopo, Private Bag X1106, Sovenga 0727, South Africa. An inter-generic grafting procedure which eliminated incompatibility challenges between Citrullus and Cucumis genera was developed. Two separate micro-plot experiments, one on Citrullus cv. ‘Congo’ onto Cucumis inter-grafts and the other on Citrullus cv. ‘Charleston Gray’ on Cucumis inter-grafts were initiated to determine the performance of the inter-grafts with respect to: (1) Cucumis nematode-resistant rootstocks retaining their nematode-resistance status to Meloidogyne incognita and (2) Citrullus cultivars as scions retaining their growth potential. The three treatments, viz. (i) un-grafted watermelon, (ii) watermelon-on-Cucumis africanus and (iii) watermelon-on-Cucumis myriocarpus, were each inoculated with 1 000 eggs and second-stage juveniles of M. incognita. Relative to untreated controls, grafting cv. ‘Congo’ on C. africanus and C. myriocarpus reduced RF values by 92-93%, while for cv. ‘Charleston Gray’ the reduction was by 94-96%. Similarly, root galls were reduced by 94-96% and 90-98% for cv. ‘Congo’ and cv. ‘Charleston Gray’, respectively. Generally, treatments had no effect on performance of watermelon inter-grafts. In conclusion, Cucumis seedling rootstocks retained their non-host status capabilities to M. incognita, while scions of the two Citrullus cultivars retained their growth potential when grafted on Cucumis species.

222 Journal of Nematology, Volume 46, No. 2, June 2014 USE OF CLIMEXÒ BIOCLIMATIC MODELS TO IDENTIFY MELOIDOGYNE ARTIELLIA AND DITYLENCHUS GIGAS RISK PRONE AGRO-ECOLOGICAL REGIONS IN THE WORLD. Poole1, M.C. and S. B. Sharma2. 1Department of Agriculture and Food Western Australia, 3 Baron-Hay Court, South Perth WA 6151, Australia; 2Murdoch University 90 South Street Murdoch WA 6150, Australia. The British root knot nematode, Meloidogyne artiellia and stem nematode, Ditylenchus gigas are economically important nematode species. Host range for M. artiellia includes cereals, legumes and crucifers and is known to cause severe losses to chickpea. Compared with Meloidogyne incognita and Meloidogyne javanica, M. artiellia currently has limited distribution, confined mainly to the Mediterranean and European regions. Compared with Ditylenchus dipsaci, D. gigas has a limited host range and geographical distribution. In this study bio-climatic models were developed using CLIMEXÒ bioclimatic software to identify climatic regions suitable for M. artiellia and D. gigas to establish. The bioclimatic models for M. artiellia and D. gigas indicated that suitable areas existed in several countries in Africa, Asia, Europe, North America, Oceania and South America. The bio-climatic models suggest that both nematode species have the potential to adversely impact host crop production in many agro-ecological regions where they have not yet established. At least eight countries in Africa, ten countries in Asia and four countries in South America have climatic conditions suitable for M. artiellia. Similarly, at least two countries in North America, six in South America, seven in Africa, ten in Asia, eleven in Europe and two in Oceania have climatic conditions suitable for D. gigas. The introduction of M. artiellia and D. gigas to these new geographical regions is highly likely in absence of effective biosecurity measures for pathways that have the potential to introduce nematode species. The paper highlights the need for identifying nematode pest free regions and maintaining their freedoms by implementing appropriate nematode surveillance strategies for early detection of nematode incursions and preparedness to implement eradication responses. TWO NEW STEINERNEMA SPECIES FROM THE ICORNUTUM GROUP FROM ZULIA STATE (VENEZUELA). Portillo, E., P. Morales-Montero and E. San-Blas. Instituto Venezolano de Investigaciones Cientı´ficas, Centro de Estudios Bota´nicos y Agroforestales, Laboratorio de Proteccio´n Vegetal, Av. 8 entre Calles 79 y 80, Maracaibo, edo. Zulia, Venezuela, CP 4001. During a survey of entomopathogenic nematodes in Zulia state (northwestern Venezuela), two populations of the genus Steinernema were isolated from soil samples. Steinernema sp. (cachirı´ strain) was isolated from cultivated pastures whereas Steinernema sp. (LPV474 strain) was isolated from a transitional fallow land. The presence of two horn-like papillae in the lip region, indicate that both species belong to the ‘bicornutum-group’ of Steinernema spp. For taxonomic studies, 10 Galleria mellonella were exposed to infective juveniles, first and second generation males and females were collected. For morphological and morphometric studies, 20 specimens from different stages of the nematodes were heat killed at 60 8C for 2 minutes; fixed in TAF and mounted in glycerine. The two populations were compared with the other nematodes from the ‘‘bicornutum group’’. Steinernema sp. (cachirı´ strain) differs from Steinernema riobrave, Steinernema yirgalemense and Stienernema pakistanensis in spicule and gubernaculum length and in number and shape of genital papillae. The species also differs from S.riobrave in the presence of vela on spicule and from S. pakistanensis in the absence of mucrons. Steinernema sp. (LPV474 strain) can be distinguished from Steinernema bicornutum by the absence of mucrons, spicule length and shape, tail length and gubernaculum length. These two species represents the first reports of Steinernema from Venezuela. DEVELOPING EFFECTIVE CONTROL STRATEGIES TO CONTROL THE CITRUS NEMATODE, TYLENCHULUS SEMIPENITRANS, IN SOUTHERN AFRICAN CITRUS ORCHARDS. Pretorius, M.C. and C. Kotze. Citrus Research International, PO Box 28, Nelspruit, 1200, South Africa. Tylenchulus semipenetrans infects citrus worldwide and is the most abundant and frequent plant-parasitic nematode in citrus orchards with yield losses of up to10% recorded. Multiple commercial applications of these compounds proved to be highly effective in reducing nematode populations in orchards. The use of several of these toxic compounds have, however, globally become under pressure. The search for alternative control methods to effectively control citrus nematodes is a priority in the Southern African citrus industry. Pre-plant fumigation of replant soils with methyl bromide during the 1980’s was highly effective in reducing nematode numbers in citrus orchards and also increased yields and fruit size. Available commercial fumigants and post-plant treatments were done on a replant orchard at Crocodile Valley Citrus Co. with the treatments being 50% methyl iodide and 50% chloropicrin methyl bromide; metam-natrium; 1.3 dichloropropene, 1.3 dichloropropene/chloropicrin; Furfural; Nematode egg stimulant + nematicide and Cadusafos. Initial results indicated that the female nematode counts in the fumigated treatments were significantly lower, decreasing between 81 and 98% compared to the untreated control. Tree height and stem diameter of the fumigated treatments (276 cm) increased significantly compared to the unfumigated treatments (254 cm) and untreated control.Two non-toxic, post-plant products were also evaluated with the Bayer product (BCS AR 83685 SC 500), reducing the female population counts by a minimum of 70% after only one application. Certain dosages of the Makhteshim product (Nimitz (MCW-2) also effectively reduced the female population numbers by 72%. The need to seek cost effective alternatives for the South African citrus industry is imperative for future research.

Abstracts 223 HOST STATUS OF COMMERCIAL WHEAT AND SUNFLOWER CULTIVARS TO MELOIDOGYNE SPP. Pretorius1, M., S. Steenkamp2, H. Fourie1 and A.H. Mc Donald1. 1North-West University, Unit of Environmental Sciences and Management, Private Bag X6001, Potchefstroom, 2520 South Africa; 2Agricultural Research Council-Grain Crops Institute, Private Bag X1252, Potchefstroom, 2520 South Africa. Meloidogyne spp. cause extensive damage to a number of crops because of their well-adapted feeding mechanism, their high reproductive potential and wide host ranges. Although producers still rely heavily on chemical control to manage these nematode pests, reliable but toxic nematicides has been taken off the market. The future of the remaining nematicides is also uncertain. Host plant resistance is an alternative management tool that can assist in keeping root-knot nematodes below damage threshold levels. Twenty commercially-available sunflower and 29 wheat cultivars were tested in greenhouse trials against Meloidogyne incognita and Meloidogyne javanica, respectively, to determine their host status to these two root-knot nematode species. A susceptible tomato cv. Floradade was included in both trials as a susceptible standard. Parameters used to select for resistance included reproduction factors, egg-laying female indices and number of eggs and J2 per root system. None of the sunflower cultivars tested showed resistance to M. javanica. All the wheat cultivars tested exhibited resistance to M. incognita. The latter screenings will be repeated during the next growing season to confirm resistance of these wheat cultivars to M. incognita. EVALUATION OF A BIOLOGICAL SOIL CULTURE AS A POTENTIAL ORGANIC AGENT FOR MANAGING ROOT-KNOT NEMATODES. Pretorius, M.S.A., W.J. van Aardt, A.H. Mc Donald, C. Jansen and H. Fourie. Unit for Environmental Sciences and Management, North-West University, Private Bag X6001, Potchefstroom, 2520. Extensive research efforts are underway to identify novel agents for the management of Meloidogyne spp. that parasitize and damage a range of crops. The effect of various filtered and non-filtered concentrations of a soil-derived organic and biological compound, Soil Bio-Muti (SBM), was investigated on the motility, respiration and reproduction of Meloidogyne incognita in separate experiments. All SBM concentrations both filtered and non-filtered, reduced J2 motility significantly (p#0.05) from 3h to 24h after exposure compared to the tap water control. Significant interactions existed for ELF indices, egg and J2 numbers/root system in terms of the treatments, filtered and non-filtered entries 30 days after J2 that had been exposed to the SBM concentrations had been inoculated on roots of susceptible tomato seedlings. Furthermore, a progressive decrease in J2 respiration occurred as the SBM concentrations in which J2 where suspended, increased. Results of this study suggest that SBM has inhibiting effects on J2 motility and oxygen consumption as well as reproduction of Meloidogyne incognita. Follow-up studies will be conducted to identify the biological agents and/or their secondary metabolites that adversely affected M. incognita in studies done to date. Field testing is also envisioned since this may shed more light on the mechanism of the causing agents that could render an alternative and environmentally safe product for use in the management of root-knot nematodes. IPM TO CONTROL ROOT KNOT NEMATODES ON TOMATO IN BEIJING OF CHINA. Qiu, J., T. Liu and W. Liu. Institute of Plant & Environment Protection, Beijing Academy of Agriculture and Forestry Sciences, Beijing China 100097. Control of Meloidogyne incognita using an IPM approach was examined in this study.The treatments were Microfume plus metham-sodium, granules made by raised broth of Paecilomyces lilacinus, the concentrate raised broth of Aspergillus niger-Y61 or sesame dregs, alone, or combined with lythidathion granules or abamectin granules. Treatments were applied by bunch irrigation before the transplanting of tomato seedlings. Also, resistant varieties of tomato such as Xian Ke series, X.K 5, X.K 6 and X.K 8, were used to be the contrast, since they have resistance to root knot nematode in protected cultivation. The number of root knots, juveniles and egg masses/100g soil were counted. The best treatment was fumigation followed by P. lilacinus granules or A. niger-Y61 broth or sesame dregs respectively; plus lythidathion granules or abamectin granules. The result showed the root knot numbers were reduced from 61% to 87%; the J2 numbers were reduced from 85.33% to 87.11%; reduced rate of egg masses over 81.47%. Our assay indicated that the most effective control for root knot nematode (M. incognita) on tomato is to use resistant varieties of X.K5. The use of chemical pesticides can be reduced significantly from 10% to 50% by the combination of nematocide lythidathion granules plus P. lilacinus granules or sesame dregs or A. niger-Y61broth. INTENSITY OF DROUGHT STRESS ON SUGARCANE - MELOIDOGYNE INCOGNITA PATHOSYSTEM. Quintela1, M.P., E.M.R. Pedrosa1, M.M. Rolim1 and A. Chaves2. 1Universidade Federal Rural de Pernambuco, Rua Dom Manoel de Medeiros, s/n, Recife, 52171-900, Pernambuco, Brazil; 2Estac xa˜o Experimental de cana-de-acxu´car do Carpina, Universidade Federal Rural de Pernambuco, Carpina 55810-000, Pernambuco, Brazil. This study aimed to evaluate early sugarcane development and activity of catalase and ascorbate peroxidase enzymes under different intensity and lasting of drought stress associated with Meloidogyne incognita parasitism. An experiment was carried out in a greenhouse using micro propagated seedlings of sugarcane RB92579 variety, for 90 days. Experimental design was completely randomized in factorial arrangement 7 (drought stress -56% of pot water capacity (PWC) for 30 days, 56% PWC for 60 days, 56% PWC for 90 days, 22% PWC for 30 days, 22% PWC for 60 days, 22% PWC for 90 days and the

224 Journal of Nematology, Volume 46, No. 2, June 2014 control, 90% PWC) 3 2 (M. incognita density – 0 and 20.000 eggs per pot) with four replications. Increases in time and intensity of drought stress decreased M. incognita reproduction and rate of sugarcane leaf area development, but increased ascorbate peroxidase activity. The longer the drought, the slower was post-stress plant recovery. Catalase activity was not affected by drought or nematode stress. The most stressful conditions provided the highest reductions on plant height, stalk diameter, shoots and root dry biomass, as well on biomass allocation to shoots, but it increased biomass allocation to the root system and the root/shoot ratio. ROOT-KNOT NEMATODE EFFECTORS TARGET HOST CELL NUCLEUS TO ESTABLISH FEEDING CELLS. Quentin, M., L. Zurletto, M. Jaouannet, M. Magliano, M.N. Rosso, E. Danchin, P. Abad and B. Favery. INRAUniversite´ de Nice-Sophia Antipolis-CNRS, UMR 1355-6243 Institut Sophia Agrobiotech: 400 route des Chappes F-06903 Sophia Antipolis France. Root-knot nematodes are obligate endoparasites able to infect almost all cultivated plants worldwide. These nematodes maintain, for weeks, a biotrophic relationship with their hosts by inducing the differentiation of root cells into hypertrophied and multinucleate feeding cells. Nematode effectors produced in the esophageal glands and injected within host cells through the stylet certainly play a role during infection to manipulate plant cell morphogenesis and physiology, and promote nematode establishment. Blocking the activity of parasitism gene products involved in the success of infection would offer specific alternatives to control nematodes in the field. In this regard the nematode effectors are particularly attractive targets. In a search for new nematode effectors, we used transcriptome analysis and comparative genomics to identify Meloidogyne incognita genes encoding proteins potentially secreted in the host tissue during the early steps of infection. We identified candidate parasitism genes that were specifically expressed in early parasitic stages, encoded predicted secreted and nuclear localized proteins. We localized the expression of several candidate parasitism genes in the pharynx, the intestine or specifically in the esophageal glands of parasitic juveniles. Among these new effectors identified, we demonstrated the secretion of MiEFF1 by the nematode in planta and immunolocalised the secreted MiEFF1 in the nuclei of giant-cells. As functional analyses and characterization of their plant targets progress, the roles of these M. incognita nuclear effectors are currently elucidated. Here we present these effectors and the hypothesis for their roles with regards to the unique feeding behaviour of these pests. SPATIAL DISTRIBUTIONS OF NEMATODE TAXA IN ARABLE AND (SEMI-)NATURAL FIELDS. Quist1, C., P. Mooyman1, S. van den Elsen1, D. Brus2, C. Mulder3, A. Termoshuizen4, J. Bakker1 and J. Helder1. 1Laboratory of Nematology, Wageningen University, P.O. Box 8123, 6700 ES, Wageningen, The Netherlands; 2WUR-Alterra, Soil Geography, Droevendaalsesteeg 3, 6708 PB Wageningen, The Netherlands; 3National Institute for Public Health and the Environment (RIVM), A. van Leeuwenhoeklaan 9, 3720 BA Bilthoven, The Netherlands; 4BLGG Research, Binnenhaven 5, 6709 PD Wageningen, The Netherlands. Because of their abundance, their trophic diversity and their relatively straightforward extractability, nematodes have a potential as a proxy for the biological condition of soils and sediments. So far the potential of this group is underexploited for technical reasons; the microscopic analysis of nematode assemblages is labour intensive and requires extensive expertise. With the availability of a relatively large molecular framework, it became possible to design molecular assays that allow for the quantitative analysis of individual taxa against complex DNA backgrounds. The availability of high throughput tools to monitor nematode assemblages makes it possible to investigate the spatial distribution of nematode taxa at field level. Experts are aware that some nematodes occur in patches whereas others are more or less evenly distributed over a field. In order to assess the degree of patchiness of individual nematode taxa, twelve fields (4 from each marine clay, river clay and sandy soils) of a single hectare each were investigated in great detail. Using a sampling grid optimized for geostatistic analysis, 96 composite samples were taken from each of these fields. Each of these samples were analysed with 24 molecular assays (corresponding to 23 nematode taxa and one internal control). Preliminary results, visualized as semi-variograms and surface maps, show a great variation in distribution patterns not only on the basis of nematode taxa but also based on soil type. The data that will be presented are essential for the design of scientifically sound sampling schemes for agricultural and natural soil at hectare scale and above. POST-PLANT NEMATODE CONTROL IN QUEEN PINEAPPLE PLANTINGS IN HLUHLUWE, SOUTH AFRICA THE STORY AFTER ALDICARB. Rabie, E.C., and B.W. Mbatha. ARC-ITSC, Hluhluwe Research Station, P.O. Box 194, Hluhluwe 3960, South Africa. Nematodes can be considered the most damaging pest of Queen pineapples cultivated in northern KwaZulu-Natal. Although more than 30 nematode species were found associated with pineapples in South Africa only Meloidogyne javanica and Pratylenchus brachyurus are of economic importance, off which the latter is the most damaging, especially on sandy soils. Control practices prior to planting include fallowing, crop rotation, application of soil amendments and chemical control. Chemical control of nematodes is the primary control method applied in Queen pineapple cultivation in South Africa. Preplant control is essential when a nematode population is already present. The only option for post plant control is the

Abstracts 225 application of systemic nematicides due to the migratory endoparasitic character of P. brachyurus. The necessity of post plant control will be determined by the length of the crop cycle and the residual effect of the nematicide used for preplant control. Until recently nematode control was mainly done by preplant soil fumigation followed by the application of fenamiphos, oxamyl of aldicarb as post-plant treatment. The combination of fumigation and aldicarb gave the best control with the additional effect of mealybug and redmite control. Aldicarb was withdrawn from the market in 2011. The application of other granular nematicides into the plant funnel/heart was investigated for post-plant nematode control. If applied just before or after rain, this application method is found more effective as well as environmentally safe due to the minimal contact with the environment and lessened possible contact with the operators. PERSPECTIVES ON GLOBAL REGULATION AND STEWARDSHIP OF SOIL FUMIGANTS. Racke, K. and J. Busacca. Dow AgroSciences, Crop Protection R&D, 9330 Zionsville Road, Indianapolis, Indiana 46268 USA. Soil fumigants play a critical role in global agriculture for the effective management of nematodes and other soil-dwelling pests in many high value crops. As regulatory requirements and societal expectations for crop protection chemicals have evolved, manufacturers and growers have had to adapt and innovate to maintain access to an effective toolbox of products. With respect to soil fumigants, regional regulatory re-evaluation programs as well as the global phase-out of methyl bromide under terms of the Montreal Protocol have significantly impacted production practices and created additional opportunities for change. What has been the past experience and what is the future outlook for soil fumigants in such an evolving landscape? This paper will review significant regulatory developments related to soil fumigants, with particular emphasis on global availability of key products and impacts of major re-evaluation programs such as U.S. re-registration and the EU review. Key factors for effective navigation of regulatory change will be examined including new data generation, advanced human health and environmental assessments, practices to manage emissions, and the importance of assessing agronomic benefits and grower practices as part of the re-evaluation process. The overarching importance of effective product stewardship practices for maintenance of fumigant products will also be examined. Examples of regulatory requirements and stewardship practices will be drawn from experiences with the fumigant 1,3-dichloropropene (1,3-D), the active ingredient in TeloneÒ soil fumigants. DEVELOPMENTAL PLASTICITY CORRELATES WITH ACCELERATED MORPHOLOGICAL EVOLUTION IN DIPLOGASTRID NEMATODES. Ragsdale, E.J., V. Susoy and R.J. Sommer. Department of Evolutionary Biology, Max Planck Institute for Developmental Biology, 35 Spemannstreet, 72076 Tu¨bingen, Germany. Developmental plasticity has been proposed as a vehicle for phenotypic diversification in animals and plants. However, few studies test the morphological potential of plastic traits in a phylogenetic framework. Here, we test the evolutionary consequences of developmental plasticity in the nematodes of Diplogastridae, which include species with moveable teeth, a predatory lifestyle, and in many cases stomatal dimorphism. We have studied the evolution of diplogastrid feeding-structures by analyzing changes of form and complexity, particularly in the context of a robust molecular phylogeny. In our analysis, stomatal dimorphism was supported to be an ancestral feature of Diplogastridae. The origin of the plasticity coincided with the appearance of predatory teeth and immediately preceded a radiation of complex feeding-forms. Comparative morphometric analyses show that developmental plasticity is associated with a shift to increased rates of phenotypic evolution. Furthermore, the fixation of a single phenotype has coincided with a further increase in disparity of stomatal shape. Thus, a macroevolutionary ‘‘pulse’’ of plasticity and the subsequent fixation of forms have allowed evolutionary novelties, increased morphological complexity, and rapid morphological exploration following a release from developmental constraints. COST EFFECTIVE SOLID-STATE PRODUCTION OF ENTOMOPATHOGENIC NEMATODES (STEINERNEMATIDAE). Ramakuwela1, T., J. Hatting1, M.D. Laing2 and S. Hazir3. 1ARC-Small Grain Institute, Bethlehem, 9700, South Africa; 2School of Agricultural, Earth and Environmental Sciences, College of Agriculture, Engineering and Science, University of KwaZulu-Natal, Pietermaritzburg, South Africa; 3Department of Biology, Faculty of Arts and Sciences, Adnan Menderes University, 09010 Aydin, Turkey. Production of entomopathogenic nematodes (EPN) for large scale commercial application has been restricted to developed countries because of high capital input and running costs associated with liquid fermentation. In developing countries, production will probably be achieved using in vivo culturing in insects, or by using in vitro solid culture on a small to medium scale. This study focused on optimising solid culture production of an indigenous Steinernema sp. for insect biocontrol purposes. Six low cost production media were evaluated at optimum temperature. Medium 6 produced the highest yield of IJs (781 678±221 IJs) / 5g medium, the highest level of live IJs (>84%) and the lowest number of adults (90% in all experiments. The optimum storage temperature was determined by incubating IJs in 0.1% formalin solution at 5 temperatures over 84 days. Survival was highest and most stable at 158C, ranging from 84-88%. Storage in a sponge formulation at a concentration of 2.5 million IJs improved survival by 6% compared to aqueous storage at 158C. Furthermore, EPN storage in a sponge at 258C, after a period of low temperature (158C) storage for 84 days, did not

226 Journal of Nematology, Volume 46, No. 2, June 2014 have a detrimental impact on IJ survival and infectivity (87 and 95%, respectively). An estimated retail price (R90.61) of this Steinernema sp. was considerably lower than the market price for other Steinernema species, ranging from R271.50 to R458.55. TESTING OF NEW NEMATICIDES FOR USE IN THE SOUTH AFRICAN SUGAR INDUSTRY. Ramouthar, P.V., N. Ntshobeni , U. Pillay and S.D. Berry. South African Sugarcane Research Institute, Private Bag X02, Mt Edgecombe 4300, South Africa. Due to its efficiency, Temik (aldicarb) was the most widely used nematicide in the South African sugarcane industry. News of its withdrawal highlighted the overuse of this chemical within the industry and this prompted the industry not only to look for a replacement for Temik but also to support continuous research into alternative products. This study deals with the testing of a wide range of products for use in the South African sugar industry. The first stage of testing was conducted using pot trials. Fifteen products were tested and the range included chemical and biological products, liquids and granules and registered and unregistered products. Results from the pot trials were encouraging and based on these results, four products were chosen for field testing. These products contained the active ingredients abamectin, cadusafos, fenamiphos and a compound yet to be revealed by the manufacturer at varying rates. Treatment with the test nematicides showed a 25-92% reduction in plant parasitic nematode numbers in the roots of the plant crop trial at six months and a 46-91% reduction in the ratoon trials. A 12-89% reduction in thrips numbers, currently a major pest in the South African sugar industry, was also noted for one of the products. Registration trials for this product are thus currently in progress. Research for more alternatives is still continuing to reduce over reliance on a single chemical. Further exploration of the biologicals is also a key focus with the promotion of environmentally friendly sustainable farming within the industry. STUDY OF NEMATODES IN THE RHIZOSPHERE OF CITRUS ORCHARDS IN SOUTHEAST OF IRAN. Rashidifard1, M., E. Shokoohi1, A. Hoseinipour1 and S. Jamali2. 1Plant Protection Department, College of Agriculture, Shahid Bahonar University of Kerman, Kerman, Iran; 2Plant Protection Department, College of Agriculture, Guilan University, Rasht, Iran. Citrus orchards are located in the south and north of Iran. The citrus industry is an important part of the national economy of Iran. Plant parasitic nematodes cause losses of approximately 10-30% of the world’s citrus orchards .During a survey on the biodiversity of plant parasitic nematodes in citrus orchards of the Shahdad region in Kerman province (southeastern Iran), 50 samples were collected from the rhizosphere of citrus trees during 2011 and 2012. Nematodes were extracted using the modified Baermann funnel method, fixed, transferred to glycerin and mounted on permanent slides. The nematodes were identified using morphological and morphometrical diagnostic characters, using a light microscope fitted with a drawing tube. Thirteen nematode species, belonging to 10 genera including Agamermis spp., Alaimus sp., Aphelenchus avenae, Hemicriconemoides chitwoodi, Diphtherophora spp., Nanidorus minor, Pratylenchus neglectus, P. scribneri, Psilenchus cf. aestuarius, P. hilarus, P. cf. klingleri and Tylenchorhynchus agri were found from this survey. In addition, Tylenchulus semipenetrans was recovered in most of the soil samples studied. ASSESSING HOST-PLANT GENE EXPRESSION DURING INFECTION OF SYNCYTIUM-FORMING NEMATODE ROTYLENCHULUS RENIFORMIS. Redding1, N., P. Agudelo1 and C. Wells2. 1School of Agricultural, Forest, and Environmental Sciences, Clemson University, Clemson, South Carolina, USA 29634; 2Department of Biological Sciences, Clemson University, Clemson, South Carolina, USA 29634. Reniform nematode, Rotelynchulus reniformis, is a semi-endoparasite capable of infecting more than 300 host plant species in tropical, subtropical, and warm temperate regions. Its mode of parasitism is a complex process whereby female nematodes penetrate host roots and introduce effectors that lead to the formation of feeding sites. Physiological changes in host cells cause them to fuse together into a multinucleated mass called a syncytium from which the nematode feeds. Our long-term goal is to identify vulnerable points in the process of syncytia formation that can be disrupted to halt the nematode life cycle and protect soybean plants. The objective of the current study was to identify genes that are differentially expressed during the early stages of syncytium formation in soybean. A split root system allowed us to inoculate half the roots of each plant with R. reniformis while the rest of the root system remained uninfected. Illumina RNA-seq was performed on infected and uninfected root tissue harvested from three replicate plants at three, six, nine and twelve days post-inoculation. Resulting sequence data were assembled with the Cufflinks pipeline, annotated with Blast2GO, and used to compare infected and uninfected soybean transcriptomes with JMP-Genomics. From the resulting list of differentially-expressed genes, we have identified several candidates believed to be important in syncytia formation. These genes will be examined in additional functional analyses and in situ hybridization studies to confirm their role in syncytium development. DISCOVERY OF A NEW BIORATIONAL NEMATICIDE. Rehberger, L., B. Belkind, G. Venburg and R. Eldridge. Valent BioSciences Corporation, 6131 Oakwood Road, Long Grove, Illinois 60047, USA. A novel and highly effective biorational nematicide recently discovered by Valent BioSciences Corporation shows significant activity against plant parasitic nematodes in laboratory, greenhouse and small-plot field tests. VBC90017 is a

Abstracts 227 patented formulation of two biorational nematicidal active ingredients that synergistically controls nematodes and reduces nematode damage. VBC90017 offers potential timing and applications flexibility for the grower, showing good nematode and galling control from both pre-plant and post-plant applications. Results from a range of laboratory, greenhouse and research small-plot experiments are discussed. VOTIVOÒ: INNOVATION DRIVEN, NEMATODE PROTECTION AND PLANT HEALTH BENEFITS. Riggs1, J., W. Andersch2, U. Pluschkel2 and K. Bugg1. 1Bayer CropScience US, Research Triangle Park, NC, USA; 2Bayer CropScience AG, Alfred-Nobel-Straße 50, D-40789 Monheim am Rhein. Bacillus firmus I-1582 is a natural occurring, non-transgenic spore-forming bacterium developed by Bayer CropScience for suppression of nematode damages by seed-treatment (Poncho/VOTiVO) or by soil-application (FlocterÒ, NorticaÒ). VOTiVO formulations are sold as fluid suspensions or wettable powders demonstrated to have a shelf-life of more than 2 years. The spores of the bacteria also remain active on the seeds for an extended period of time, under storage conditions conducive for seed viability. Seed treatment with Poncho/VOTiVO led to a colonization of the root-surfaces immediately after seed germination by the bacteria. The bacteria grow and multiply by using the organic compounds, excreted by the plants. Root exudates are considered to be involved in the orientation of various nematode species towards their host plants. Spores of Bacillus firmus I-1582 are also able to colonize on nematode eggs and perforate the egg-shell, ceasing the development of the juveniles. Seed treatment with VOTiVO demonstrates enhanced vigor and growth of seedlings, resulting in increased root & shoot-masses in comparison to the control plants. Bacillus firmus is known for its ability to form the phytohormone indol-acetic acid (auxins) or to solubilize phosphorous from soil constituents leading to a more vigorous seedling growth. In corn, the performance of VOTiVOÒ has been evaluated in combination with the insecticidal seed treatment Poncho. Field trials demonstrated that Poncho/VOTiVO is capable to reduce plant damages from a wide range of nematodes. In the majority of field trials, Poncho/VOTiVOÒ treated seeds produced higher yields, 5-6 bu/acre (314-377 kg/ha). ROOT-KNOT NEMATODE RESISTANCE GENE STACKING IN BREEDING LINES USING HIGH-THROUGHPUT GENOTYPING. Roberts1, P.A., B.L. Huynh1, T.J. Close2 and W.C. Matthews1. 1Department of Nematology, University of California, Riverside, California 92521, USA; 2Department of Botany & Plant Sciences, University of California, Riverside, California 92521, USA. As an advancement over breeding for nematode resistance based solely on infection assays and field screens to phenotype and select breeding progenies, high-throughput genotyping platforms are becoming available for comprehensive genomewide marker selection schemes. Genomewide marker coverage based on SNP marker polymorphism in biparental markerassisted recurrent selection (MARS) and marker-assisted recurrent backcross (MABC) populations has been applied to breeding for resistance to multiple root-knot nematode (Meloidogyne) species in the grain legume cowpea (Vigna unguiculata). QTL analysis and genetic mapping were used to identify the locations of resistance trait determinants in the cowpea genome. Foreground selection for marker haplotypes in the QTL regions carrying the positive resistance alleles was combined with background selection for elite plant type using evenly spaced genomewide markers. Using this approach, breeding lines with broad-based resistance were developed which carry resistance genes effective against avirulent and virulent isolates of Meloidogyne incognita, Meloidogyne javanica, and Meloidogyne arenaria, and which combine resistance to both root-galling and nematode reproduction (egg production). Co-selection for resistance to Fusarium wilt, which can forms a disease complex with root-knot nematodes, also has been achieved using the SNP-marker technology. A configurable workflow developed as a Breeding Management System by the CGIAR-Generation Challenge Program for applying the molecular-based breeding tools of this system is being used in the advancement of MARS and MABC progenies to develop nematode resistant varieties. CUBA: TWO DECADES WORKING ON INTEGRATED NEMATODE MANAGEMENT IN AGRICULTURAL CROPPING SYSTEMS. Rodrı´guez1, M.G., E. Fernandez2, L. Hidalgo-Dı´az1, R. Cuadra3, J.M. Draguiche4, H. Gandarilla5 and I. Castro-Lizazo6.1National Centre for Plant and Animal Health (CENSA).P.O.Box 10. San Jose´ de las Lajas. Mayabeque Province. Cuba; 2Plant Health Research Institute (INISAV), Calle 110 #514. Playa Municipality Havana, Cuba; 3Reasearch Institute for Tropical Agriculture (INIFAT), Havana, Cuba; 4Villa Clara Province Plant Health Laboratory, Carretera de Maleza Santa Clara Villa Clara, Cuba; 5National Quarantine Laboratory. National Centre for Plant Health, Agriculture Ministry, Cuba; 6Agricultural University of Havana (UNAH), Mayabeque Province, Cuba. The agriculture policy in Cuba has changed since the last decade of the twentieth century, and today co-existing urban/ periurban agriculture and different production systems are present. Plant parasitic nematodes (mainly Meloidogyne spp., Radopholus similis and Pratylenchus ssp.), represents some of the most harmful pests in horticultural, coffee and fruits systems. The quarantine service is responsible for surveillance in international borders and for sampling the most important crops annually. Several tactics has been developed and introduced in these areas, such as crop rotation, trap crops, biodisinfection and biological control agents, accompanied by capacity building of stakeholders. Several strains of Trichoderma

228 Journal of Nematology, Volume 46, No. 2, June 2014 spp., Bacillus thuringiensis and commercial biological products such as KlamiCÒ (Pochonia chlamydosporia var. catenulata) and HebernemÒ (Tsukamurella paurometabola), have showed good results in different conditions and are included in some integrated nematode management systems in farms, organic gardens and governmental agricultural enterprises. Grafting tomatoes, different preventive and hygienic alternatives and biological control agents are using in crops under shelter conditions, and the use of chemical nematicides is only authorized in high infections. In soil biodisinfection, several organic materials have been used, such as native plants, crops debris, and by-products from sugar and rum industries, among others. Trap crops, use of healthy plants, extraction of nematode susceptible weeds, biological control, and management of non susceptible crops are the most common alternatives in organic gardens. In governmental agricultural enterprises of banana, tuber crops and vegetables they use mainly tillage, healthy seeds, resistant varieties, biological control and crop rotation. A summary of research and results will be presented. DEVELOPING A BIOLOGICAL CONTROL AGENT FOR MOLLUSCS IN SOUTH AFRICA. Ross, J.L. and A.P. Malan. Department of Conservation Ecology and Entomology, Faculty of AgriSciences, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. European molluscs (slugs and snails) have become significant pests in South Africa, affecting agronomy, horticulture and viticulture. Current methods for controlling molluscs rely on chemical molluscicides, such as metaldehyde and carbamate compounds. Both these compounds are poisonous to a range of vertebrates, and studies have shown that methiocarb is toxic to a number of beneficial invertebrates, including earthworms and carabid beetles. Therefore it is important to investigate a method of biological control. To date the most successful commercial method for the biological control of molluscs in Europe is the nematode Phasmarhabditis hermaphrodita. This nematode is mass produced by BASF (formally Becker Underwood UK Ltd) and sold under the trade name of NemaslugÒ. The product works by releasing infectious nematode larvae that seek out molluscs and enter through natural openings. Once inside the mollusc, bacteria associated with the nematode are released which stop the mollusc from feeding. Death usually follows 4-21 days after nematode infection. The Agricultural Pest Act 36 of 1947 prohibits the introduction of exotic animals into South Africa. Therefore no method of biological control currently exists for molluscs in South Africa. In this study we present a systematic survey of mollusc-parasitic nematodes in South Africa, and the development of an indigenous isolate for mass production. ANALYSIS OF VIRAL SEQUENCES WITHIN SOYBEAN CYST NEMATODES. Ruark, C.L., T.L. Sit, S.R. Koenning and S.A. Lommel. Department of Plant Pathology, North Carolina State University, Raleigh, North Carolina 27607, USA. Viruses of nematodes have been understudied yet may have an important influence on nematode ecology. Soybean cyst nematode, Heterodera glycines, is a leading cause of soybean pathogenesis and dramatically suppresses soybean yield in North Carolina. Recently, four RNA viruses were reported in three highly inbred lines of H. glycines originating from Illinois. The objective of our research was to determine if North Carolina soybean cyst nematodes were also a reservoir for these viruses. Total RNA was extracted from cysts and second-stage juveniles of four inbred North Carolina soybean cyst nematode lines (OP20, OP25, OP50 and a race 2 greenhouse isolate). Total RNA was also extracted from cysts of recently collected wild soybean cyst nematode populations from North Carolina fields. PCR products of up to 500 base pairs were generated (with viral-specific primers based on published Illinois sequences), cloned and sent for sequence analysis. All four of the inbred H. glycines lines were positive for a soybean cyst nematode-specific nyavirus (ScNV) while three of the four contained a soybean cyst nematode-specific phlebovirus (ScPV). Sequence variation among inbred North Carolina strains was less than the differences with published Illinois sequences. Some field isolates of H. glycines were positive for both ScNV and ScPV. However, in at least one field population, neither of these viruses could be amplified. More sequence variation existed in field populations of H. glycines as compared to inbred lines. Soybean cyst nematode rhabodovirus (ScRV) and soybean cyst nematode tenuivirus (ScTV) detected in Illinois H. glycines populations were not found in any of the assayed North Carolina populations. Better understanding of nematode viruses may lead to a novel avenue of soybean cyst nematode biocontrol. NEMATODE FOOD WEB DIVERSITY AND STRUCTURE. Ruess, L., R. Cutando and K. Wagner. HumboldtUniversita¨t zu Berlin, Institute of Biology, Ecology Group, Philippstraße 13, 10115 Berlin, Germany. Nematode assemblages in soil reflect substrate texture, climate, biogeography, organic inputs and anthropogenic disturbance. Nematodes use a wide range of resources including bacteria, fungi, roots or microfauna, and these diverse biological interactions result in a key role in soil food webs. Shifts in forest management affect food webs distinctly as the proportions of readily decomposable to recalcitrant compounds are altered by tree species identity and diversity. Such resource diversity at the base of the soil food web is highly relevant for population dynamics and trophic complexity, and thus consumer diversity that influences ecosystem functioning. Besides tundra biomes temperate forests are the largest soil C stocks but only recently tree species diversity and identity effects on belowground compartments came into focus. The impact on the nematode microfood web was investigated in: i) a large scale study on increased land use intensity, i.e. from natural to managed forest stands, and ii) a tree diversity experiment with six species representing different litter quality (ash, lime, beech, oak, spruce, pine) and

Abstracts 229 arranged in five diversity levels (1, 2, 3, 5, 6 species mixtures). In this diversity manipulation study, nematode communities showed only weak interactions with tree diversity but distinct relation to the presence of specific species. In the observational study, nematode faunal analysis revealed disturbance by land use as well as regional effects (e.g. local climate, soil type) on food web diversity and structure. These changes, within and across trophic levels, likely shape decomposition processes and in turn energy flow in belowground food webs. POPULATION DYNAMIC OF MELOIDOGYNE FALLAX IN POTATO CROPS AND MULTIPLICATION IN STORED TUBERS. Rulliat, E., E. Treuillier, F. Ollivier, A. Buisson, C. Sarniguet, C. Prigent and L. Folcher. ANSES-Laboratoire de la Sante´ des Ve´ge´taux-Unite´ de Ne´matologie, Domaine de la Motte au Vicomte 35653 Le Rheu- France. In the European Union, Meloidogyne fallax Karssen, 1996 and Meloidogyne chitwoodi Golden, O’Bannon, Santo and Finley, 1980 are registered as quarantine nematodes according to Directive 2000/29/EC. Within the context of monitoring those quarantine pests, an experiment was conducted with the aim of studying (i) the level of contamination of potato tubers by M. fallax on ultra-early harvested crops and (ii) the capacity of M. fallax to develop within stored tubers. In that respect, a follow-up of the populations dynamic was undertaken on tubers from their initiation to the harvest time i.e. 98 days after planting. The nematological analyses combined both morphobiometrical and molecular techniques. As a result, the findings proved that even if the potatoes were harvested very early, this did not prevent the tubers from being infected by M. fallax. Second stage juvenile (J2) infected the developing tubers. These juveniles developed inside the tubers to adult females which can be detected a few weeks before harvest. Even if the level of M fallax infestation was limited at harvest time, the production of a new generation inside the stored tubers can be observed. Interestingly, this result confirms a well reported phenomenon in Meloidogyne biology but constitutes the first report of this fact for M. fallax. EFFICIENCY OF CHEMICAL TREATMENT OF MELON SEEDS ON THE CONTROL OF MELOIDOGYNE JAVANICA. Saliba1, H.C.R., C.T. Borges1, R. Navroski1, I. Lima-Medina2, J.T. Schafer1, V. Meneghello1 and C.B. Gomes. 1 Faculdade de Agronomia, Universidade Federal de Pelotas, Campus Universita´rio s/n C. P. 354, Pelotas/RS. Brazil; 2 Embrapa Clima Temperado, C.P. Postal 403, Pelotas/RS, Brazil. Root-knot nematodes are a problem pest of melon in Brazil. Considering the lack of resistant cultivars in the Brazilian market and the shortage of information about seed chemical treatment to control this pest, the aim of this study was to evaluate the efficiency of seed treatment for Meloidogyne javanica control in melon plants using a mixture of nematicide, fungicide and insecticide. Melon seeds cv. Gaucho Redondo treated with Thiabendazole 485 SC (4,8 ml) and Thiamethoxan at 4,8 ml, 0,66 ml and 1,63 ml/100 kg of seeds, respectively, were sown in polystyrene trays. Twenty days after seeding, each plant was transferred to a pot containing a mixture of sterilized soil and a substrate at a proportion of 3:2. Subsequently, each plant was inoculated with 5,000 eggs and second stage Meloidogyne javanica juveniles.Twenty-five days after the inoculation each plant was evaluated for the number of galls, second stage juveniles and eggs/root systems. Subsequently the nematode reproduction factor (RF) was determined to estimate the efficiency of seed chemical control of M. javanica. The melon seed treatment did not reduced the number of galls on the melon roots; however the use of chemical products reduced the M. javanica RF by 40% compared to the control. SPATIAL DISTRIBUTION OF POTATO CYST NEMATODES IN THE NILGIRI HILLS OF SOUTH INDIA. Saranya1, C., P. Sundararaj1 and S.L. Hafez2. 1Department of Zoology, Bharathiar University, Coimbatore, India; 2University of Idaho, Idaho, USA. Potato cyst nematodes (Globodera pallida and Globodera rostochiensis) are major nematode pests, causing 80 percent yield loss on potato crop in Nilgiri hills of Tamilnadu, India. An intensive survey was conducted in 13 potato growing villages of Nilgiri Hills to find out the distribution and occurrence of potato cyst nematodes. Nematodes were extracted from a total of 117 soil samples collected from potato fields using standard methods. In total, , 3548 mature cysts were recovered, of which 1075 were viable. Potato cyst nematode incidence was highest at Nanjanad village (61.3%) followed by Ithalar village (56.5%).The lowest incidence was observed in Balacola village (24%) followed by Ebbanadu (14.3%) village. Altitude, temperature and potato crop monoculture practices are the common factors that increase the abundance of potato cyst nematodes at the Nanjanad and Ithalar villages. DEVELOPMENT, GENES AND MORPHOLOGY IN THE PHYLUM NEMATODA. Schierenberg, E., C. Kraus, M. Kroiher, P. Schiffer, J. Schulze. Cologne Biocenter, University of Cologne, Germany. Nematodes, which are thought to date back into the Precambrian, exhibit a body plan that has remained surprisingly uniform, probably due to the unique construction principle with a single-chamber hydroskeleton. In contrast, the genetic and genomic variance between different Caenorhabditis species was found to be unexpectedly high. It is development that ties together these two disparate phenomena. To trace the evolutionary changes that took place in the long history of this phylum we started a comprehensive comparative survey of embryogenesis in a variety of phylogenetic branches leading to hatching juveniles with their taxon-specific morphological peculiarities using 4-D microscope recordings. Applying bioinformatic

230 Journal of Nematology, Volume 46, No. 2, June 2014 tools we have begun to analyse genomes and transcriptomes generated in different laboratories including our own and relate these data to the developmental features. Likewise we found major differences on the genome/gene expression level and with respect to cellular behaviour particularly between basal and more derived species but also between closely related representatives. These differences appear to be more fundamental during the first half of embryogenesis than the second half where a nematode is formed from a ball of cells. Our data indicate that the evolution of development within the phylum Nematoda went from indeterminate early cleavage without initial polarity to invariant cell lineages with early establishment of polarity. Such dramatic changes require modifications in the cell-specification processes and the underlying molecular mechanisms. Many genes that are shared between more basal nematodes and non-nematode outgroups are absent in the model system Caenorhanditis elegans and its close relatives. Thus, rather different ways of how to construct a nematode must have been established during evolution without corresponding reflection on the morphological level. THE EFFICACY OF CROP GUARD (A.I. FURFURAL) AS AN IN-SEASON SOIL TREATMENT FOR THE CONTROL OF NEMATODES ON VINES AND STONE FRUIT IN SOUTH AFRICA. Scholtz, R., E. Buntting and A. Steyn. Illovo Sugar Ltd, South Africa, PO Box 31003, Merebank, 4059, Durban, South Africa. With the phasing out of toxic nematicides and the use of more environmental friendly pesticides as in the agricultural sector internationally, the need for safer pesticides has been developed. Crop GuardÒ (a.i. furfural) is a natural product derived from various plant sources. Sugarcane bagasse is an important raw material used in the production of furfural. Illovo Sugar Ltd. is a leading sugar and furfural producer in Africa and has developed and commercialized Crop GuardÒ as an inseason soil treatment for the control of nematodes in soil under South African field conditions. Crop GuardÒ is currently registered on 18 different crops in South Africa and has also been granted a yellow band registration during 2013. Vine trials were executed at Nylstroom (Modimolle) in the Limpopo Province of South Africa. Treatments were made during the spring and autumn root flush periods of the vines. Results indicated that over three consecutive seasons, 50+25+25 l/ha rate resulted in much lower Criconema (Ring nematode) numbers than the control treatment. A peach trial was executed at Robertson in the Cape Province of South Africa. Applications were made during the spring and autumn root flush periods. Resulting data indicated that the 50+25+ 25 l/ha gave the best results in terms of yield and control on Criconema (Ring nematode) and Helicotylenchus sp. (Spiral nematode). THE EFFICACY OF PROTECT (A.I. FURFURAL) AS A PRE-PLANT SOIL TREATMENT FOR THE CONTROL OF NEMATODES ON PINEAPPLES AND STRAWBERRIES UNDER FIELD CONDITIONS IN SOUTH AFRICA. Scholtz1, R., E. Buntting1, A. Steyn1, E. Rabie2 and I. Meintjies3. 1Illovo Sugar Ltd, South Africa, PO Box 31003, Merebank, 4059, Durban, South Africa; 2Agricultural Research Council, Institute for Tropical and Subtropical Crops, Nelspruit, South Africa; 3Agricultural Research Council, Vegetable and Ornamental Plant Institute, Roodeplaat, South Africa. With the phasing out of Methyl Bromide as a general soil sterilant as well as the banning of certain nematicides in the agricultural sector of many first world countries, and shortly in third world countries too, the need for safer pesticides has developed. ProtectÒ (furfural a.i.) is a natural product derived from various plant sources. Sugarcane bagasse is an important raw material used in the production of furfural. Illovo Sugar Ltd. is a leading sugar and furfural producer in Africa and has developed and commercialized ProtectÒ (furfural a.i) as a soil treatment for the control of certain nematodes and fungi in soil under South African field conditions. A pineapple trial was executed at the Hluhluwe Research Station of the Agricultural Research Council- Instititute for Tropical ands Subtropical Crops, KwaZulu-Natal. Results indicated that a pre-plant rate of 400 l/ha ProtectÒ seven days before plant, resulted in significantly lower population numbers for Pratylenchus brachyurus and Meloidogyne spp., than the untreated control. Harvesting at two years after plant showed higher yields than for the untreated control, or similar yields as the standard EDB/Telone treatments. A strawberry trial was executed in sandy soil at Cullinan, Gauteng, South Africa. Results indicated that a pre-plant rate of 400 l/ha ProtectÒ, seven days before plant, resulted in significantly lower numbers for Meloidogyne sp. than the untreated control. Yield results showed significantly higher fruit mass per ha than for the untreated control or similar yields as the standard MBr/Telone treatments. EFFECT OF DEFENCE ELICITORS CHITOSAN, BIONÒ AND METHYL JASMONATE ON ROOT-KNOT NEMATODE M. INCOGNITA. Schouteden1, N., C. Vos2, A. Elsen3, B. Panis4 and D. De Waele1,5. 1Laboratory of Tropical Crop Improvement, Department of Biosystems, KU Leuven, Willem de Croylaan 42 – box 2455, 3001 Leuven, Belgium; 2Centre of Microbial and Plant Genetics, Kasteelpark Arenberg 20 - box 2460, 3001 Leuven, Belgium; 3Bodemkundige Dienst van Belgie¨, Willem de Croylaan 48, 3001 Leuven, Belgium; 4Bioversity International, Willem de Croylaan 42 – box 2455, 3001 Leuven, Belgium; 5School of Environmental Sciences and Develpoment, Noth-West University, Private Bag X6001, 2520 Potchefstroom, South Africa. Root-knot nematodes, Meloidogyne spp., cause substantial damage to most economically important crops. Over the recent years the use of many nematicides has been prohibited due to their harmful impact on the environment and human health. An alternative to control root-knot nematodes is the application of defence elicitors. Defence elicitors are components that induce

Abstracts 231 systemic resistance in the plant and can thus be applied in nematode management. We tested the elicitors chitosan, BionÒ (active component acibenzolar-S-methyl) and methyl jasmonate in different concentrations for its controlling activity towards root-knot nematodes. The tomato plants were grown in a mixture of sand and potting soil and sprayed with elicitors in different concentrations. Two or three days after spraying, the plants were inoculated with freshly hatched second stage juveniles of Meloidogyne incognita Malaysia and 10 days after inoculation (DAI), 21 DAI and 56 DAI plants were harvested, plant parameters assessed and the gall index was determined. In addition, the roots were collected and stained with acid fuchsin to visualize the nematodes microscopically. In another experiment egg masses and amount of eggs were assessed after 35 DAI in two tomato cultivars. Results show a minor influence of the defense elicitors against M.incognita. Implications of our results and future perspectives will be discussed. MOLECULAR STUDY OF AN ISOLATE OF STEINERNEMA FELTIAE FILIPJEV, 1934 FROM IRAN, BASED ON ITS RDNA SEQUENCE POLYMORPHISM. Sediqi1, E., E. Shokoohi1 and J. Karimi2. 1Department of Plant Protection, College of Agriculture, Shahid Bahonar University of Kerman, Kerman, Iran; 2Department of Plant Protection, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran. A survey of entomopathogenic nematodes was conducted in southeastern Iran. During this study, an isolate belonging to the species Steinernema feltiae was recovered in the Lalezar region, Kerman province. The ITS rDNA was amplified and sequenced. Molecular information based on sequence of the ITS rDNA confirmed this isolate as S. feltiae. The Nblast result showed 8 nucleotides differences between our isolate and the same species belong to Israel and Iran respectively. The phylogenetic relationship of the species studied based on Maximum Likelihood placed this isolate of S. feltiae closed to other populations of S. feltiae from Israel (KC571264, KC571263, KC571261) and Iran (JF920962, JF920964). Pairwise genetic distance using Maximum Composite Likelihood showed no genetic variation between different populations of S. feltiae from Iran. BIODIVERSITY OF ROOT-KNOT NEMATODES (MELOIDOGYNE SPP.) FROM MAJOR TOMATO GROWING AREAS OF ETHIOPIA. Seid1,3, A., W.M.L. Wesemael2,3, W. Decraemer3,4, T.M. Mekete5 and C. Fininsa1. 1Haramaya University, College of Agriculture and Environmental Sciences, School of Plant Sciences -Crop Protection Unit, P.O. Box 138, Dire Dawa, Ethiopia; 2Institute for Agricultural and Fisheries Research (ILVO), Plant Sciences Unit-Crop Protection, Burg Van Gansberghelaan 96, B-9820 Merelbeke, Belgium; 3Ghent University, Department of Biology, Nematology Unit, Ledeganckstraat 35, B-9000 Ghent, Belgium; 4Royal Belgian Institute of Natural Sciences, Vautierstraat 29, B-1000 Brussels, Belgium; 5University of Florida, Entomology and Nematology Department, Bldg.970, Natural Area Drive, Gainesville, FL 32611, USA. The objective of the study was to investigate the biodiversity of Meloidogyne spp. in major tomato growing areas of Ethiopia. Samples were collected from major tomato growing areas based on the elaborated agro-ecological zones of Ethiopia. Tomato farms and gardens in 40 different localities mainly from Rift Valley, Upper Awash and Eastern Hararghe were sampled during January to March 2013. A total of 212 randomly chosen fields were visited and 212 composite soil samples from the rhizosphere (0-25cm deep) of tomato and 123 root samples (approximately 200g each) were collected separately. Samples were collected from ten to fifteen plants per field (1/2 ha) in a zig-zag fashion. Sampling sites range from greenhouse tomato producers to small scale farms. Root Gall Index (RGI) and Egg Mass Index (EMI) were determined based on a 0-5 scale. DNA was extracted from second stage Meloidogyne juveniles (J2s) and PCR was performed using speciesspecific primers as well as primers that amplify the Intergenic Spacer Region (IGS) and 28S regions of the rDNA. PCR products were then sequenced bi-directionally. A 100% incidence of Meloidogyne spp. was detected from 47% of the samples.. An EMI and RGI of 5 were recorded from 40% of the samples. The use of species-specific primers indicated that 90% of the samples were infested by both M. incognita and M. javanica. Sequencing of the 28S region of rDNA and speciesspecific primers also confirmed a first report of M. hapla in Ethiopia at one sampling location ’Zeway’ (1640 meter above sealevel). HISTORY AND INDUSTRIAL DEVELOPMENT OF THE ‘‘RAIN OF DEATH’’ (PASTEURIA SPP.). Sekora, N. and T. Hewlett. Syngenta Crop Protection, Pasteuria Bioscience Laboratory, Alachua, FL, USA. Pasteuria species are obligate parasites of many invertebrates, including plant-parasitic nematodes. The potential of Pasteuria spp. as biological control agents has been studied since the first plant-parasitic nematode-suppressive soil was observed in the 1970’s. Quantities of in vivo Pasteuria penetrans spores sufficient for pot and microplot trials (23109) could be produced from ground root tissue containing infected root-knot nematodes (Meloidogyne spp.). This method was used in 1996 to successfully control Meloidogyne arenaria populations in the soil of the Land Pavilion at EPCOT Center, Disney World. However, the production of enough in vivo Pasteuria-laden material to treat more than a few acres was determined to be too expensive to be practical. Many attempts to culture Pasteuria spp. in vitro were met with failure until 2004, when a small startup company, Entomos, was able to grow Pasteuria penetrans in small volumes of artificial medium. Shortly thereafter, the method was refined to produce large quantities of in vitro P. penetrans was developed using fermentation in

232 Journal of Nematology, Volume 46, No. 2, June 2014 500 mL volumes by the same group, now at Pasteuria Bioscience. Syngenta Crop Protection, having acquired Pasteuria Bioscience in 2013, now has the ability to produce 50,000 L of in vitro Pasteuria spp. for use as a commercial nematicide. Production at this scale has yielded enough P. nishizawae to use as a soybean seed treatment against Heterodera glycines on more than three million acres in 2014. Further development will continue on additional Pasteuria species and will target species of Meloidogyne, Pratylenchus, and Heterodera in the near future. INCREASE IN SEVERITY OF FUSARIUM WILTS IN CUCURBITACEOUS PLANTS CAUSED BY MELOIDOGYNE INCOGNITA. Seo, YunHee and Young Ho Kim. Department of Agricultural Biotechnology and Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul 151-921, Korea. Root-knot nematode (Meloidogyne spp.) and fusarium wilt fungi cause severe damage to cucurbitaceous plants including oriental melon in Korea. Although shintozoa (Cucurbit maxima x Cucurbit moschata) resistance to fusarium wilt has been widely used as a stock for grafting oriental melon cultivars, the disease is still prevalent in major oriental melon-growing areas. Thus, experiments were conducted to examine the reasons for the current prevalence of the fusarium wilt in oriental melons. Twenty-nine Fusarium isolates obtained from oriental melon fields were inoculated on shintozoa, cucumber and oriental melon, in which most Fusarium isolates were more virulent to oriental melon and cucumber than to shintozoa. In this test, wilt severity was increased more in plants by the combined inoculation of both Meloidogyne and Fusarium than by the inoculation of fusarium isolates alone. Light microscopy also showed extensive destruction of vascular tissues with poor giant cell formation by the infection of both pathogens, while no or little extensive destruction of vascular tissues occurred by the infection of either pathogens alone. However, the formation of tyloses, which is responsible for the wilt symptom expression, was frequently observed in the oriental melon and cucumber but not in shintozoa infected with either Fusarium proliferatum or both pathogens. All of these results suggest that root-knot nematodes may be mostly responsible in the current increases of fusarium wilt severity in oriental melon cultivation in Korea, although effects of changes in pathogen virulence on the disease severity were not totally ruled out. MOLECULAR ADAPTATION MECHANISMS IN THE ANTARCTIC NEMATODE PANAGROLAIMUS DAVIDI. Seybold1, A., C. Marshall1, D. Wharton2 and P. Dearden1. 1Department of Biochemistry, University of Otago, Dunedin, New Zealand; 2Department of Zoology, University of Otago, Dunedin, New Zealand. The Antarctic nematode Panagrolaimus davidi tolerates desiccation and freezing and is the only animal known to survive intracellular ice formation. Although physiological mechanisms of this extreme adaptation are partly revealed, mechanisms on the molecular level remain largely unknown. Recent studies identified genes up-regulated during desiccation such as late embryogenesis abundant proteins, heat shock proteins, antioxidants and trehalose genes. Furthermore, the presence of iceactive proteins has been suggested by the observation of recrystallization inhibition and hexagonal ice crystal shape. To identify the genes involved in this extreme adaptation we are currently testing high throughput RNAi techniques, such as feeding and soaking. To investigate whether P. davidi is accessible to RNAi via feeding, the embryonic lethal genes Pd-rps-2 and Pd-dhc as well as the blister gene Pd-duox-42 were tested and the expression level was confirmed by quantitative PCR. While Pd-rps-2 treated samples showed a significant increased lethality compared to the control samples, there was no significant difference at the expression level. To investigate weather P. davidi is more accessible to RNAi via soaking, neurostimulants and desiccation supported soaking techniques were tested. A significant downregulation of Pd-rps-2 was obtained if samples were exposed to desiccation prior to soaking. We are currently testing RNAi on candidate genes such as trehalose and late embyogenesis abundant genes in order to identify the function of these genes during exposure to desiccation and freezing. STATUS OF ENTOMOPATHOGENIC NEMATODES AS BIO-CONTROL AGENTS IN PAKISTAN. Shahina, F. and K.A. Tabassum. National Nematological Research Centre, University of Karachi, Karachi-75270, Pakistan. The study of entomopathogenic nematodes (EPNs) in Pakistan have been initiated in the 1990’s. More than 7000 soil samples were collected from different climatic regions of Pakistan. Up till now extensive work has been carried out on the isolation, distribution taxonomy, biology and efficacy of EPNs and as a result three new species viz., Steinernema pakistanense, Steinernema asiaticum and Steinernema maqbooli were described while seven known species Steinernema abbasi, Steinernema siamkayai, Steinernema feltiae, Steinernema carpocapsae, Steinernema litorale, Heterorhabditis indica and Heterorhabditis bacteriophora were first records for Pakistan. The genetic diversity and phylogenetic analysis of all isolates of Steinernema and Heterorhabdiis were investigated on the basis of ITS regions of rDNA, 12S rRNA mitochondrial gene, D2D3 regions and 28S rDNA. One-hundred-and-forty sequences were submitted to Genbank and a genomic library was constructed. Pathogenicity and efficacy trials of indigenous EPN isolates have also been done successfully with positive and promising results of more than 100 insect pests of different crops. Because of the output of these studies the Intellectual Property Organisation of Pakistan granted twelve national patents. There is a dire need to focus further research on these EPN isolates to explore and exploit their potential as an alternative to pesticides in Pakistan, especially in IPM programmes.

Abstracts 233 BREEDING A SUPER NEMATODE FOR ENHANCED INSECT PEST SUPPRESSION. Shapiro-Ilan1, D.I. and B.J. Adams2. 1USDA-ARS, SE Fruit and Tree Nut Research Laboratory, Byron, GA 31008 USA; 2Department of Biology and Evolutionary Ecology Laboratories, Brigham Young University, Provo, UT, USA. Entomopathogenic nematodes in the genera Heterorhabditis and Steinernema are important regulators of natural insect populations, and are used commercially as biological control agents for pest suppression. Successful biocontrol applications depend on the introduced organism having an array of beneficial traits such as virulence, host-finding, environmental tolerance etc. Thus biocontrol potential can be improved by enhancing these traits. Approaches to strain improvement include discovery, selection, hybridization, transgenics or a combination thereof. These methods have been successfully applied to a number of entomopathogenic nematode species. In addition to enhancing traits, trait stability is another factor that is critical for biocontrol success. Beneficial traits can deteriorate during repeated culturing in laboratory or industrial settings. Deterioration of various traits has been reported in entomopathogenic nematodes. The cause of trait change was found to be genetically based (at least in part) and inbreeding depression was implicated as a significant contributing factor. Recently the creation of homozygous inbred lines was found to deter the negative repercussions of trait change during serial culture. Inbred lines can be generated in the laboratory through serial self-fertilization (heterorhabditids) or sibling mating (steinernematids). Additionally, for heterorhabditids only, multiple inbred lines can be automatically generated in liquid culture because the nematodes cannot mate in the liquid media. Generation of multiple versus single inbred lines for commercial development each has advantages and disadvantages. Selected inbred lines and beneficial trait improvement programs offer a substantial advancement in biocontrol potential for entomopathogenic nematodes. SPATIAL AND TEMPORAL VARIATION OF INTERTIDAL NEMATODE COMMUNITIES AFTER THE DEEPWATER HORIZON OIL SPILL. Sharma1, J., P.M. Brannock2, H. Bik3, K.M. Halanych2, and W.K. Thomas3. 1Department of Biology, University of Texas at San Antonio, San Antonio, Texas 78249, USA; 2Department of Biological Sciences, Auburn University, Auburn, Alabama 36849, USA; 3Hubbard Center for Genome Studies, University of New Hampshire, Durham, New Hampshire 03824, USA. The composition of free-living nematode communities at five intertidal sites in northeast Gulf of Mexico was studied at intervals over two years to examine their role in recovery of biotic communities in benthic sediments after contamination by the Deepwater Horizon oil spill. The diversity and composition of the nematode fauna in contaminated sediments was compared to that of pre-spill sediments. Over 6000 individuals from 70 genera representing 20 families were recovered. The generic richness of nematodes in sediments immediately following the spill decreased but gradually increased after 12 months. The pre-spill intertidal samples showed high evenness without dominance of a single species while immediately after the spill the Thoracostomopsidae and Desmodoridae families were dominant. These observations are confirmed by metagenomic analyses of nematode communities that show a significant decline in abundance of nematodes following the oil spill, with a corresponding increase and overwhelming dominance, of fungal taxa previously associated with hydrocarbons. The vertical distribution of the nematode community showed that the lower depths of 3-10cm had greater generic richness and evenness as compared to the upper 0-3cm although there was variation in species composition between the two depths. The dominance of scavenger and predatory nematode taxa following the oil spill suggests that they may utilize the abundant fungi in post-spill sediments. The re-colonization of nematodes on beaches after an oil spill suggests that they are important in recovery of biotic communities in contaminated sediments. RESTORING ECOSYSTEM SERVICES OF URBAN SOILS: USE OF NEMATODE COMMUNITY AS A BIOINDICATOR TO REESTABLISH MISSING LINKS IN SOIL FOOD WEBS. Sharma, K.1 and P.S. Grewal1,2. 1Environmental Science Graduate Program, Ohio State University, OARDC, Wooster, Ohio 44691, USA; 2Entomology and Plant Pathology Department, University of Tennessee, Knoxville, Tennessee, 37996, USA. Utilizing urban vacant land for growing food crops can promote access to healthy food in low income neighborhoods. However, soil health and quality are major concerns for urban agriculture in post-industrial cities. Replacing existing soil with top-soil from a neighboring area is the most common solution applied, during which no attention is given to the health of the soil organisms being transported. Nematodes form an important and diverse component of the soil food web, with presence across multiple trophic levels, offering the most comprehensive insight into the structure and function of food webs. Previous studies showed that due to intensive disturbance, nematode food webs in urban areas had significantly lower structural complexity and lacked higher trophic guilds (predatory and omnivorous species), thus severely limiting the ecosystem services potential of the soil. We tested a new approach to improve the soil food web structure in targeted vacant lots, to restore the full spectrum of ecosystem services provided. We hypothesized that the missing nematode trophic guilds (along with associated soil organisms) in the soil food web can be transplanted/reestablished using small soil cores (9cm dia, 5cm deep) when coupled with specific organic amendments. Field trials of core transplantation into turf grass plots (1.5 m X 1.5 m) resulted in a significant increase in the structure index over a period of 6 weeks. Positive results from this study could eliminate the need for large-scale topsoil replacements in urban gardens and will pave the way for establishment of sustainable and safe urban food production systems.

234 Journal of Nematology, Volume 46, No. 2, June 2014 NEMATODE QUARANTINE AND BIOSECURITY RELATED TRADE AND GLOBAL FOOD SECURITY ISSUES. Sharma1, S.B., C. Magnusson2, J.J. Chitambar3 and U.C. Kodira4. 1Murdoch University, 90 South Street, Murdoch Western Australia 6150, Australia; 2Norwegian Institute for Agricultural and Environmental Research (Bioforsk), Høg˚ s, Norway; 3California Department of Food and Agriculture, 3294 Meadowview Road, Sacramento, skoleveien 7, N-1432 A California 95832-1448, USA; 4USDA-APHIS-PPQ, Plant Health Programs, 4700 River Road, Riverdale, Maryland 20737, USA. Plant-parasitic nematodes cause economic losses of over 14% in developing countries. Many of the known and highlydamaging species of nematodes are relatively localised in distribution. The consequences of globalisation including advances in transport technologies have raised significant biosecurity concerns, and surge in nematode dispersal risk accompanied by climate change is likely to create new opportunities for nematode populations to move and establish in new geographical regions. Nematode quarantine and biosecurity capabilities are generally lacking in the food insecure developing countries. Nematode quarantines and stringent regulatory measures have been successful in prevention and containment of highly damaging nematodes such as the potato cyst nematodes (Globodera rostochiensis and Globodera pallida) and pine wood nematode (Bursaphelenchus xylophilus). Currently, G. rostochiensis is regulated as a quarantine pest by 118 countries, while G. pallida is regulated by 81 countries. There is every probability for these species to gain entry into non-infested regions as populations below 500,000 cysts per hectare cannot be detected reliably. The pine wood nematode is endemic to North America and has spread to Japan, China, Korea, Portugal and Spain. A global approach to nematode quarantine and biosecurity is needed to minimise international and intercontinental invasion of nematode pests. Policy development and implementation of effective pre-border, border and post border biosecurity measures are needed. Safeguarding of food production regions from invasive exotic plant-parasitic nematodes is a must to achieve food security for the present and future generations. Case studies on regulatory control of potato cyst nematodes and pine wilt nematodes will be presented. RELATIONSHIP BETWEEN BELONOLAIMUS LONGICAUDATUS, TRICHODORUS OBTUSUS, AND TURFGRASS ROOTS. Shaver1, B., P. Agudelo1 and B. Martin2. School of Agricultural, Forest, and Environmental Sciences, Clemson University.1114 Long Hall Clemson, SC 29365; 2Pee Dee Research and Education Center 2200 Pocket Rd. Florence, SC 29506. Natural in-field variability of turfgrass root growth and nematode density makes it difficult to determine effects of nematicide or fungicide treatments applied to manage nematodes or fungal infections that result from nematode feeding. Over the past two years we have analyzed over 1000 soil cores (5-cm diameter by 20-cm depth) from trials on hybrid bermudagrass (Cynodon dactylon X C. transvaalensis) infested with Belonolaimus longicaudatus or from ‘Empire’ zoysiagrass (Zoysia japonica) infested with Trichodorus obtusus. Treatments included three different nematicides, a fungicide, and nematicide and fungicide combinations. A 100 cm3 soil sample from each soil core was collected and assayed for nematode density using centrifugal sugar flotation. Roots in each soil core were then washed free of remaining soil, dried for five days at 708C, and incinerated at 5258C for 3 hours in order to determine dry root weights. To more fully understand the relationship between nematode feeding and root growth, we regressed dry root weight with nematode density. In a separate nematicide trial, we compared the variability of root weights to root architecture. Soil cores (5-cm diameter by 20-cm depth) were collected and roots washed free of soil, scanned on a flatbed scanner, and analyzed for root architecture. Afterwards, root weights were evaluated as described above. Data shows that root architecture may be a better indicator of nematode damage than root weights. Results of the regression analysis will be helpful in understanding treatment effects of this and future nematicide trials. ADAPTED TECHNIQUE FOR EXTRACTING NEMATODES FROM CARROTS. Shubane, A. and M. Marais. National Collection of Nematodes, Biosystematics Programme, ARC-Plant Protection Research Institute, Private Bag X134, Queenswood 0121, South Africa. It has always been difficult to correlate the symptoms of carrots (Daucus carota subsp. sativus) infested with Meloidogyne species and the number of nematodes observed. A technique was adapted to successfully extract root-knot nematodes from carrots. The carrots, including all secondary roots, were peeled with a vegetable peeler; the peels and the secondary roots were cut into 2 mm strips, placed in tap water and macerated in a blender at medium speed for 45 seconds. Ten millilitres of 4 % NaOCl was added to the suspension and aerated with an aquarium pump for 72 hours. After aeration the suspension was poured through a tower of 1000 mm -, 150 mm-, 45 mm - en 38 mm sieves, and thoroughly washed with a strong water stream. The macerated carrot pieces collected on the top sieve were discarded and the rest were washed in centrifuge tubes to which 5 m‘ kaolin was added and centrifuged for 7 min at 3500 rpm. The sample was then centrifuged with a sugar solution (450 m‘ in 1 ‘ of water) for 3 min at 3500 rpm. After washing the nematode supernatant through a 38 m sieve to remove all traces of sugar, the sample was ready to determine the population numbers of both plant-parasitic and free-living nematodes. This technique was also successfully used to extract nematodes from beetroot, potatoes, peanut seeds, and peanut hulls.

Abstracts 235 POSITIONING NEMATOLOGY RESEARCH AND DEVELOPMENT THROUGH NIESA. Sibanda1, Z., H. Talwana2, W. Wanjohi3, W. Kimenju4, K.G. Davies5,6, R.H. Manzanilla-Lo´pez5, C. Masawe7 and N. Luambano-Nyoni8. 1 Goldengro (pvt) Limited, P.O. Box MP 1306, Mount Pleasant, Harare Zimbabwe; 2Department of Crop Science, Faculty of Agriculture, Makerere University, P.O. Box 7062, Uganda; 3Agricultural Science and Technology, Kenyatta University, P.O. Box 43844 – 00100; Nairobi, Kenya; 4Faculty of Agriculture, University of Nairobi, P.O. Box 30197-00100, Nairobi, Kenya; 5Department of AgroEcology, Rothamsted Research, Harpenden, Hertfordshire, AL5 2JQ, United Kingdom; 6School of Life and medical Sciences, University of Hertfordshire, Hatfield, Hertfordshire, AL10 9AB, United Kingdom; 7Ministry of Agriculture and Food Security, Tanzania; 8Sugarcane Research Institute, P.O. Box 30031, Kibaha Pwani, Tanzania. Since its inception in 2005, the Nematology Initiative for Eastern and Southern Africa (NIESA) is a project that has been sponsored by the Gatsby Charitable Foundation (UK) to build capacity in the discipline of Nematology, and to develop a network of expertise in eastern and southern Africa, originally with technical support from a UK consortium – CABI Bioscience, Rothamsted Research and the University of Reading. At present, NIESA is composed of a cadre of qualified nematologists from Kenya, Malawi, Tanzania, Uganda and Zimbabwe. The project has also started to move from its capacity building phase to sharing and transferring NIESA nematologists’ expertise to ascertain the extent to which plant-parasitic nematodes act as a constraint to local and regional crop production and to create awareness among farmers and local communities about the importance of nematodes. To achieve this, NIESA, as a group, will continue carrying out scientific research and training of farmers and phytosanitation staff for the practical benefit for local communities, crop health and food security. Networking, joint research fund application, cross-learning and peer support among practising nematologists in Africa can facilitate an active and interactive support to overcome the lack of a critical mass of nematologists in any one country. It can also link the network to information services available through partner scientists, formal research and training collaborations to improve understanding and raise the profile of Nematology within Africa. Further details of the NIESA partners and activities can be found at the web site www.africannematology.org . NINJA: AN AUTOMATED CALCULATION SYSTEM FOR NEMATODE-BASED BIOLOGICAL MONITORING. Sieriebriennikov1, B., H. Ferris2, R.G.M. de Goede3.1Nematology Unit, Ghent University, K. L. Ledeganckstraat 35, 9000 Ghent, Belgium; 2Department of Entomology and Nematology, University of California, Davis, CA 95616, USA; 3 Department of Soil Quality, Wageningen University, P.O. Box 47, NL-6700 AA, Wageningen, The Netherlands. Nematodes are important bioindicators of soil health and ecosystem services. Index systems based on life-cycle properties, feeding habits, biomass and metabolic activity have been developed for analysis and interpretation of nematode assemblages. We have developed an R code, NINJA (Nematode INdicator Joint Analysis), to calculate indices automatically. The code is compiled in html and distributed over the web. It has a user-friendly interface accessible via: http://spark.rstudio.com/ bsierieb/ninja/. The only input requirement is a table with the taxonomic inventory of the nematode assemblage. The output includes tables and graphs which contain values for: Maturity Index, Maturity Index 2-5, Plant-parasitic Index, Channel Index, Enrichment Index, Structure Index, total biomass, and the composite, enrichment, structure, herbivore, fungivore, bacterivore, predator and omnivore metabolic footprints as well as proportions of different feeding types and c-p classes amongst herbivore and free-living forms. For all measures but the percentages, means and standard deviations are calculated per treatment or sampling site and accompanied by corresponding p-values. The tool is supported by a database, stored on the server, which contains life-history characteristics, c-p (coloniser-persister) assignments, feeding habits and body masses of most soil nematodes. The tool permits adjustment of default feeding habits based on the user’s observation and experience. The NINJA tool is free, flexible and user-friendly; it greatly simplifies nematode faunal analysis and protects against calculation errors. Data interpretation is enhanced by graphs and primary statistical treatment. Intended future expansion of the database will include sequence accession numbers that will enable processing data based on molecular identification. UPLAND COTTON GERMPLASM LINES WITH INTROGRESSED RESISTANCE TO THE RENIFORM NEMATODE. Sikkens1, R.B., K.S. Lawrence1, D.W. Schrimsher2, S.R. Moore3 and D.B. Weaver4. 1Entomology & Plant Pathology, Auburn University, Auburn, Alabama, USA; 2Southern Agronomists, AGRI-AFC, Decatur, Alabama, USA; 3 Research & Development Scientist, Syngenta, Monroe, Louisiana, USA; 4Crop, Soil and Environmental Sciences, Auburn University, Auburn, Alabama, USA. Over the past decade a concerted effort was made to develop host plant resistance to reniform nematode (Rotylenchulus reniformis) in upland cotton (Gossypium hirsutum). Since 2007 this effort has resulted in the release of several germplasm lines with introgressed resistance to reniform nematode. Currently four groups of released germplasm lines can be distinguished based on their source of resistance and developmental background: (a) the LONREN group, with reniform nematode resistance derived from Gossypium longicalyx, (b) the BARBREN group with reniform nematode resistance derived from wild accession GB-713 of Gossypium barbadense, (c) the M713 group, also with resistance derived from the GB-713 accession but with a different developmental trajectory, and (d) the MT2468 group, with reportedly moderate levels of R. reniformis resistance derived from the photoperiodic primitive race accession TX2468 of G. hirsutum. During 2013 we evaluated the potential of 13 lines from these four groups, including three lines of the LONREN group, two lines of the

236 Journal of Nematology, Volume 46, No. 2, June 2014 BARBREN group, five lines of the M713 group and three lines of the MT2468 group. Conventional cultivars FM966 and SG747 were included as R. reniformis susceptible controls. Our studies were conducted in the field, on adjacent plots with and without reniform nematode present, on reniform nematode infested outdoor microplots, and in the greenhouse. Results indicate that germplasm lines BAR 41 of the BARBREN group and all five M713 lines combine great R. reniformis resistance with excellent field tolerance. SURVEILLANCE STRATEGIES FOR EARLY DETECTION OF NEMATODE INCURSIONS. Singh1,2,3, S.K., M. Hodda2, N.C. Banks1,2,4 and Rajan5. 1Cooperative Research Centre for Plant Biosecurity, Bruce, ACT, 2617, Australia; 2 CSIRO Ecosystem Sciences, Canberra, ACT, 2601, Australia; 3Graham Centre for Agricultural Innovation (an alliance between Charles Sturt University and the NSW Department of Primary Industries) Wagga Wagga NSW, 2678, Australia; 4 School of Veterinary and Life Sciences, Murdoch University, Murdoch 6150, Australia; 5Crop Science Division, Indian Council of Agricultural Research, New Delhi 110114, India. The spread of plant parasitic nematode species worldwide has increased over the last two decades and as a result the damage they cause to global crop production. Due to shortage of resources, most countries still depend upon traditional quarantine system of on border examination only. New developments in quarantine systems are being implemented with the involvement and active roles being played by the World Trade Organisation and International Plant Protection Convention. Now member countries carry out pest risk analysis, based upon pre export information/ field data to assess and minimise threat. Listing pests already present in a country and especially knowing what pests are not there through surveys; in territories of that country are the most critical and indispensable data requirements. Recent developments on biosecurity measures designed to prevent the entry and establishment of exotic species, are one of the most cost effective and environmentally friendly means of averting the negative impacts. Plant-parasitic nematodes due to their microscopic size, low probability of detection and diverse range of pathways represent a major challenge for the implementation of biosecurity measures worldwide. Surveillance strategies are an essential part of the plant biosecurity continuum and can greatly aid in the early detection of nematode incursions. In this paper, the design of surveillance strategies is illustrated using examples of potato cyst (Globodera spp.) and pine wood (Bursaphelenchus spp.) nematodes. Methods for prioritization of exotic nematode species; spatial mapping of risks; and targeted sampling taking into consideration seasonal cropping patterns and nematode population growth are discussed. GLOBAL BIOSECURITY RISKS FROM THREE PLANT-PARASITIC NEMATODE SPECIES TO MAIZE, RICE AND WHEAT, USING CLIMEX MODELLING. Singh1,2,3,4, S.K., D.J. Kriticos1,3, G.J. Ash2 and M. Hodda1,4. 1CSIRO Ecosystem Sciences, Canberra, ACT, 2601, Australia; 2Graham Centre for Agricultural Innovation (an alliance between Charles Sturt University and the NSW Department of Primary Industries) Wagga Wagga, NSW, 2678, Australia; 3Cooperative Research Centre for National Plant Biosecurity, Bruce, ACT, 2617, Australia; 4CSIRO Biosecurity Flagship, Canberra, ACT, 2601, Australia. Climate suitability is one of the most important factors affecting the chances of a translocated species establishing in a foreign location. CLIMEX models were developed to estimate the eco-climatic suitability and growth potential of three plantparasitic nematodes; Heterodera zeae, Hirschmanniella oryzae and Meloidogyne graminicola, globally. The models were carefully parameterised based on the species phenology and their global distributions extracted from peer-reviewed literature. To simulate the different field conditions under which these species occur, the models included rain-fed and irrigation scenarios. Except for a few occurrences, the known geographical distributions for all three species were in accord with their projected potential distributions and substantiated by available experimental and field observations. Maps of host crops (maize, rice, wheat) and irrigation areas globally were compared with projected growth index maps of H. zeae, H. oryzae and M. graminicola and countries at risk are identified. All three species have the potential to expand their distribution range in countries where they are not present yet but have favourable eco-climatic and growth index values. Under irrigation scenarios, much greater areas globally are conducive for the growth of these three species than under rain-fed conditions. When assessing the establishment potential and biosecurity risks from exotic species, presence of modified conditions such as irrigation need to be taken into consideration. THE EFFECT OF SPIROTETRAMAT ON ROTYLENCHULUS RENIFORMIS INFECTING PINEAPPLE. Sipes, B. University of Hawaii, 3190 Maile Way, Honolulu, Hawaii, 96822, USA. Spirotetramat is a lipid biosynthesis inhibitor used to control sucking pests in a wide range of crops. The compound has been beneficial in the management of plant-parasitic nematodes in several woody perennial crops. The objective of these tests were to determine if spirotetramat controlled damage in pineapple (Ananas comosus) caused by Rotylenchulus reniformis. In a greenhouse experiment, pineapple crowns were dipped in water or a solution of spirotetramat (730 mlMovento/ha or 7.9 ml/ 10 l water) with each plant receiving the equivalent of 16 ml/plant. The crowns were planted in 20-cm diameter pots filled with a 1:1 sterile soil-sand mix and inoculated with 5,000 mixed life stages of R. reniformis. One month later, half the plants dipped in spirotetramat were treated with a foliar application of 7301ml Movento/ha. The pineapples were destructively

Abstracts 237 harvested 9 months later. Spirotetramat treated plants had lower numbers of nematodes compared to the untreated pineapple (48% and 44% lower), although plant growth was similar. A similar experiment was established in a commercial pineapple field. At 9 months after planting, pineapple D-leaf weights were similar among the same treatments. Where no preplant nematicide was applied, the spirotetramat dip and foliar application tended to increase D-leaf weight (35 g vs 33 g). Populations of R. reniformis in the field were similar regardless of the spirotetramat treatments received. Spirotetramat application affected nematode development on pineapple in the greenhouse and may lessen nematode damage in the field. MODELLING OF TEMPERATURE DEPENDENT DEVELOPMENT OF TROPICAL ROOT-KNOT NEMATODE SPECIES. Sˇirca, S., P. Strajnar, M. Knapicˇ, U. Zˇibrat and G. Urek. Agricultural Institute of Slovenia, Hacquetova ulica 17, 1000 Ljubljana, Slovenia. Root-knot nematodes are poikilothermic organisms and their development is generally temperature dependent. In a previous experiment in growth chambers we estimated that Meloidogyne ethiopica required 67, 48 and 36 days to complete the reproduction cycle, at mean daily temperatures of 18.3, 22.7 and 26.38C, respectively. The base temperature (Tb) for M. ethiopica was established in a pot experiment at 148C. At 13.98C, the species was not able to reproduce. This data we then used to build a model for M. ethiopica development using Michaelis-Menten regression models. The extrapolated model suggests a minimum of approximately 40 DAI’s (at 248C to 308C), while a stop in development is indicated at about 13.98C (at DAI = 365). The model proved to be very useful for predicting M. ethiopica population dynamics. Additionally, we evaluated the differences among linear regression models of temperature – development rate relationships of six Meloidogyne species. The relation between temperature and development time appears to be linear only at temperatures several degrees Celsius above the base temperature. In fact, for the six species included in this study, linear models can be considered as applicable only at temperatures above 198C. At lower temperatures, development rate decreases exponentially, not linearly. Our results suggest that linear models fail to accurately indicate minimum required time for postembryonic development. We argue that Michaelis-Menten or other non-linear models should be used for assessing thermal requirements of ontogenetic development. ULTRASTRUCTURE OF SPERM DEVELOPMENT IN THE GENUS DITYLENCHUS. Slos1, D., P. Ensafi1, M. Claeys1, V.V. Yushin2, W. Decraemer1 and W. Bert1. 1Nematology Research Unit, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium; 2A.V. Zhirmunsky Institute of Marine Biology, FEB RAS, Vladivostok, Russia. The ultrastructure of the spermatogenesis in Ditylenchus arachis and Ditylenchus dipsaci has been studied. The general pattern of spermatogenesis is recognized as ‘rhabditid’, a pattern which is rather conserved throughout the order Rhabditida. Spermatozoa of Ditylenchus represent an aberrant type of male sperm because of the absence of an axoneme and acrosome, a characteristic shared with other nematodes. The sperm development includes the formation of complexes of fibrous bodies (FB) and membranous organelles (MO) which appear in the spermatocytes. These complexes start to dissociate in separated MO and FB in the spermatids. Immature spermatozoa are unpolarised cells with a centrally located nucleus surrounded by spherical fibrous bodies and MO located at the periphery. In both species, the spermatheca contains chains of mature spermatozoa consisting of amoeboid bipolar cells subdivided in a pseudopod devoid of organelles and a main cell body. The main cell body consists of a centrally located nucleus lacking a nuclear envelope, many mitochondria and MO. The MO are connected to the plasmalemma and open to the intercellular exterior via a pore. Hence, our data unequivocally show that MO are present in Ditylenchus, contrary to what has been described before also for D. dipsaci, and thus showing the typical rhabditid pattern. Although both species have a similar spermatogenesis, D. dipsaci differs from D. arachis in having mature spermatozoa in the male testis, a character which is, to our knowledge, not observed before. This is a further indication that the well-studied developmental mechanisms in the informative model Caenorhabditis elegans cannot simply be extrapolated to other rhabditids. PEAT AS A SUITABLE GROWTH MEDIA AND CARRIER FOR DACTYLELLA OVIPARASITICA. Smith Becker, J., H. Witte, and J.O. Becker. Department of Nematology, University of California, Riverside, CA 92521. Dactylella oviparasitica is a facultative parasitic ascomycete of several endoparasitic nematodes. Various botanical materials such as ground corn cob, ground walnut shells and a commercial preparation of reed sedge peat (BioAPT) were tested for their ability to serve as carrier media for the introduction of the hyperparasite into soil. Hyphal growth of D. oviparasitica was inhibited by walnut shells and corn cob caused severe plant stunting when added as a soil amendment at 1% w/w. BioAPT peat supported fungal growth without affecting seedling growth. Grown aseptically on BioAPT peat, the fungus remained viable for at least two years when stored at room temperature or at 4C. In greenhouse trials pasteurized sandy loam was amended with various concentrations of D. oviparasitica peat formulation and seeded with Swiss chard. After seedling emergence, the pots were infested with 750 J2/100 cm3 of Heterodera schachtii. Fungal peat amendment at 1% (DW/DW) reduced the number of recovered nematode cysts 5 weeks after infestation by up to 70% compared to the infested control with autoclaved amendment. As little as 0.05% D. oviparasitica peat amendment reduced the number of cysts by up to 50%. These results suggest that BioAPT peat may serve as a useful media and carrier for the introduction of D. oviparasitica into soil for population suppression of the sugar beet cyst nematode.

238 Journal of Nematology, Volume 46, No. 2, June 2014 NEMATOLOGY EDUCATION IN EUROPE: CLIPPING OR SPREADING WINGS. Smol1, N. and W. Decraemer1,2. 1 Ghent University, Ledeganckstraat 35, 9000 Ghent, Belgium; 2Royal Belgian Institute of Natural Sciences, Vautierstraat 29, 1000 Brussels, Belgium. The aim of university level education in Nematology at Ghent University is to train specialists equipped to take up job positions in all subjects related to nematodes, as well as in broader areas. The programme therefore has a multidisciplinary approach and for this cooperation with other universities and institutes within Europe and beyond is important. However, cooperation and mobility in and out of Europe involves a cascade of administrative and logistic problems. An overview will be given of the different options for collaborative education in Europe at MSc and PhD level, dealing with important topics such as grants, enrolment fees, curriculum and visa issues. The masters course has been in existence at Ghent University for 21 years; the intake of students and the jobs taken up by alumni will be presented and discussed. In addition, the following points will be discussed: How effectively has the availability of this MSc in Nematology been disseminated? Is there a need for Nematology training? Who is willing to cooperate to provide this training? How do we attract students to this field? How do we attract industrial partners? Who can, and is willing to, sponsor education? NEMATOLOGICAL RESEARCH AND DEVELOPMENT IN PAKISTAN: HISTORICAL PERSPECTIVE AND FUTURE PROSPECTS. Soomro, M.H. and F. Shahina. National Nematological Research Centre, University of Karachi, Karachi-75270, Pakistan. Nematodes have been affecting man since times immemorial as parasites of human, animals and agricultural pests. Plant parasitic nematodes are considered hidden enemies of crops which represent unique challenges to agricultural research. Nematological research in Pakistan has commenced in 1974 and up to now extensive surveys have been conducted throughout Pakistan which resulted in the identification of 760 nematode species and 2 new genera of which 229 were new to science. The main institution involved in Nematological research is the National Nematological Research Centre (NNRC) at the University of Karachi, which has been contributing significantly on the biodiversity and biosystematics of nematode fauna especially the description and identification of new genera/species. During recent years, significant progress has been made in the morphological, biochemical, molecular biology, genomics, biocontrol and mass production of beneficial nematodes. A genomic library has been developed at NNRC on the basis of ITS, 28S/D2D3, 18S rDNA regions and 12S rDNA genes of the indigenous root-knot nematodes, entomopathogenic nematodes and their symbiotic bacterial species/ stains, which adds to the worldwide database. Research on biological agents based on the ecological knowledge provides sustainable alternative control methods and replaces highly toxic chemical insecticides. The research efforts of our group and optimization of nematode production methods at NNRC have provided an impetus and advancement in the biopesticides industry in Pakistan. The team feels that the research endeavours of the last 12 years have enabled us to embark upon commercialization of the native strains of EPNs from Pakistan. However, we have yet to learn from colleagues and professionals from around the globe and are searching for international collaboration. CARBOHYDRATES IN THE GLIA-SECRETED MATRIX OF CAENORHABDITIS ELEGANS SENSE ORGANS. Spiegel1,2, Y., Y. Lu2 and S. Shaham2. 1Department of Entomology and Nematology, Agricultural Research Organization, Volcani Center, P.O. Box 6, Bet Dagan 50250, Israel; 2Laboratory of Developmental Genetics, The Rockefeller University, 1230 York Avenue, New York, New York 10065, USA. Amphids are the largest, most important sensory organ of nematodes. These bilateral organs consist of 12 sensory neurons and two glial cells, referred to as sheath and socket cells. At the tip of the nematode head these two come together to form a tube – the amphid channel, which contains a material, the matrix, of which some components are secreted by the sheath glia. These 12 neurons have been studied mainly by attraction assays and genetic tools. There is, however, little knowledge about the matrix composition and origin. By labeling wild-type and various mutants (cilia and glia-ablated) of the nematode Caenorhabditis elegans with fluorescent and gold-conjugated lectins (carbohydrate-binding proteins), we were able to gain preliminary information about the nature and localization of carbohydrates in the amphids and phasmids glia matrix. Nacetyl-glucosamine (GlucNac) residues are located, in wild type species, in different glia cells around the sensory cilia extended from the neuron dendrites within the amphid channel, and also protracted anteriorly and posteriorly to the amphid channels. Mannose residues are located mainly anteriorly and within the glia cells within the amphid channel and in the inner/ outer labial and cephalic neurons in the tip of the nematode ‘nose’. Phasmids reveal the presence of both carbohydrate moieties. In several mutants (e.g. che-2 or osm-6), carbohydrate were presented only in the anterior part of the glia cells, and slim label observed in the phasmid. Amphid neuron dye filling defected species (fig-1 mutant) did not exhibit neither GlucNac, nor mannose residues in both amphids and phasmids. DISPLACEMENT OF MELOIDOGYNE INCOGNITA BY ROTYLENCHULUS RENIFORMIS DURING A 10 YEAR COTTON MONOCULTURE AND IMPLICATIONS FOR SITE-SPECIFIC MANAGEMENT. Spurlock1, T.N., T.L. Kirkpatrick2, C.S. Rothrock3 and S.W. Monfort4. 1University of Arkansas Division of Agriculture, Southeast Research and Extension Center, Monticello, AR, USA; 2University of Arkansas Division of Agriculture, Southwest Research and

Abstracts 239 Extension Center, Hope, AR, USA; 3University of Arkansas Division of Agriculture, Fayetteville, AR; 4Clemson University, Edisto Research and Extension Center, Blackville, SC, USA. Meloidogyne incognita and Rotylenchulus reniformis are economically important nematode species in cotton production. Evidence from 2001-2003, in a texturally diverse cotton field near Portland, Arkansas, USA, suggested that populations of M. incognita had the greatest effect on yield in soils with greater sand content. Additionally, exhaustive nematode assays indicated R. reniformis was increasing in population and distribution in the field. Since that work, the field has continued to be planted in cotton until being planted in soybean in 2013. From the original 512 plots, nematode population densities were measured in the fall immediately after the crop was harvested in 2011 and 2013. Over the 10 year period, populations of M. incognita declined sharply while R. reniformis increased. Univariate Moran’s I indicated significantly aggregated distributions for both pathogens in all years except 2011 and trend surface models indicated Range values decreased for M. incognita but increased for R. reniformis prior to 2013. Bivariate exploratory spatial analysis and spatial regression indicated a significantly dispersed distribution and inverse relationship in 3 of 4 years and 2 of 4 years respectively confirming competition and displacement of M. incognita by R. reniformis. Neither R. reniformis nor M. incognita populations were directly related to yield loss across soil textures. The highest yield impact of nematode populations occurred in the sandier areas of the field indicating site specific management in these textural zones should have the greatest economic return. NEW REPORTS AND MOLECULAR DIAGNOSTICS OF ROOT-KNOT NEMATODES FROM GOLF COURSE GREENS IN THE WESTERN UNITED STATES. Skantar1, A.M., C. Nischwitz2, Z.A. Handoo1, S. Subbotin3, M.N. Hult1, M.E. Schmitt4 and M.A. McClure4. 1USDA-ARS Nematology Laboratory, Beltsville, Maryland, 20705; 2Dept. of Biology, Utah State University, Logan, Utah, 84322; 3California Department of Food and Agriculture, Sacramento, California 95832; 4School of Plant Sciences, University of Arizona, Tucson, Arizona 85721. Several species of root-knot nematodes (Meloidogyne spp.) are known to occur on turfgrass. A survey of 238 golf courses in the Western United States foundroot-knot nematodes in 60% of putting greens sampled. Morphology and phylogenetic trees inferred from multiple DNA markers were used to identify specimens from 110 golf courses. Meloidogyne naasi was the most common species, distributed from coastal Southern California to Washington; Meloidogyne graminis and Meloidogyne marylandi were found mostly in the warmer regions of the Southwest. PCR-RFLP of a mitochondrial DNA region distinguished these two species, providing a simple method for future diagnosis of these nematodes from turf. Specimens from two golf courses in Washington were identified as Meloidogyne minor based on analysis of the 28S, ITS1&2, and IGS2 ribosomal markers and the nuclear protein-coding gene Hsp90 sequences. Sequence-characterized amplified region (SCAR) primers previously reported as specific for Meloidogyne fallax were found to cross-react with M. minor. A population from California was determined to be M. fallax based on juvenile morphology and analysis of the ribosomal markers and Hsp90. The phylogenetic relationships of these populations and known root-knot nematode species reconstructed using Hsp90 gene sequences were congruent with those obtained from ribosomal RNA genes. Resolution of M. fallax and Meloidogyne chitwoodi using Hsp90 was equivalent to species separation obtained with 28S rDNA alignments. The strengths and weaknesses of ribosomal and Hsp90 markers, and the use of SCAR PCR as diagnostic tools will also be discussed. MOLECULAR IDENTIFICATION OF PLANT-PARASITIC NEMATODES OF QUARANTINE CONCERN AT THE UNITED STATES DEPARTMENT OF AGRICULTURE (USDA). Skantar, A.M., Z.A. Handoo, L.K. Carta and D.J. Chitwood. USDA-ARS Nematology Laboratory, Beltsville, Maryland, 20705. One mission of the USDA Agricultural Research Service Nematology Laboratory in Beltsville, Maryland is to provide nematode identifications that are urgently required by regulatory agencies, federal and state authorities, or other ARS scientists for research, regulatory actions, and control purposes. In addition to traditional morphology-based taxonomic approaches, molecular methods are often required to confirm diagnoses or to provide conclusive identification in those instances when species morphology is ambiguous or when adult specimens are unavailable for microscopic examination. Current approaches, challenges, and new developments in nematode molecular identification will be discussed within the context of some recent diagnostic scenarios involving Globodera, Meloidogyne, and Anguina spp. AVICTAÒ COMPLETE CORN – A PREVENTATIVE SOLUTION AGAINST THE HIDDEN PEST. Slaats3, B., J. Dreyer1 and C. Watrin2. 1Syngenta Crop Protection AG, Thornhill Office Park Block 14, Halfway House 1685, Johannesburg, South Africa; 2Syngenta Crop Protection AG, 410 Swing Road, Greensboro, North Carolina 27409, USA; 3 Syngenta Crop Protection AG, Schaffhauserstr. 101, 4332 Stein AG, Switzerland. Maize (Zea mays L.) nematode damage is difficult to identify and is often misdiagnosed by the grower. Due to a growing world population and limited production areas, a demand for higher yields in existing production areas has arisen coupled with increased intensification which is very favourable for nematode multiplication. AvictaÒ Complete Corn is a Syngenta SeedCare solution offering triple protection from the day the seed is planted against nematodes, insects and major soil and seed borne diseases under all environmental conditions, as a convenient ‘‘in the bag’’ solution. This first-of-its-kind seed treatment is immediate and effective ensuring maize seedlings are protected at emergence and early growth stages. With

240 Journal of Nematology, Volume 46, No. 2, June 2014 limited maize nematode control options on the market, Avicta Complete Corn offers protection in the convenience of a seeddelivered treatment – helping to increase plant health and yield potential. Syngenta SeedCare initiated an extensive testing of the yield benefit of Avicta Complete Corn in South Africa. The nematicidal component of AvictaÒ Complete Corn is abamectin which is highly active against a broad range of plant-parasitic nematodes. In one study across 12 trials, Avicta Complete Corn outyielded the standard fungicide/insecticide seed treatment in 80% of the trials delivering a yield increase of 5% on average. In another study across 22 trials under high nematode pressure AvictaÒ Complete Corn led to higher yields than the soil-applied nematicides Terbuphos or Carbofuran in 60% of the time. Overall applying AvictaÒ Complete Corn provides reliable protection and allows growers to maximize yield and economic return. PHYLOGENETIC RELATIONSHIPS OF RHIGONEMATOMORPHA– NEMATODES FROM THE HIND GUT OF DIPLOPODS. Spiridonov, S.E. and S.V. Malysheva. A.N. Severtsov Institute of Ecology and Evolution, Russian Academy of Sciences, Leninskii pr., 33, Moscow,119071, Russia. The posterior part of the diplopod intestine is inhabited with the nematodes of the superfamilies Thelastomatoidea, (Oxyuridomorha), Rhigonematoidea (Rhigonematomorpha) and Ransomnematodiea (Rhigonematomorpha). Thelastomatoidea (pinworms) are similar to the thelastomatids of other arthropods, whereas Rhigonematoidea and Ransomnematoidea are morphologically different from pinworms. The representatives of two families for each of these superfamilies were studied under light and scanning electron microscope. The partial sequences of 18S and 28S rDNA were also obtained. The phylogenetic analysis of each separate ribosomal sequence did not resolve the phylogeny of the nematodes of diplopods with the representatives of separate families forming independent clades. The monophyly of superfamilies was not evident. When partial 18S and 28S rDNA sequences were concatenated forming a 2297 bp long alignment, the phylogeny of each superfamily was securely proven using Bayesian Inference (posterior probabilities 99% and 100% for Rhigonematoidea and Ransomnematodiea, correspondingly). However, the monophyly of Rhigonematomorpha was not supported, with Rhigonematoidea, Ransomnematoidea and Ascaridoidea forming three independent clades with Oxyuridomorpha linking to the basal node of this tripartite evolutionary line. Besides the molecular apomorphies, morphological ones were found for each separate superfamily clade. The following features are considered as secure markers of taxonomic independence of these groups: structure of excretory-secretory system, presence of somatic sensillae, structure of the egg-shell. Phylogenetic analysis based on more representative and characteristic sequences is needed to resolve the phylogeny of these nematodes. EVOLUTION OF NOVELTY: FROM PREDATORY FEEDING TO NOVEL CHEMISTRY, NEW GENES AND EPIGENETICS. Sommer, R.J. Max-Planck Institute for Developmental Biology, Tu¨bingen, Germany. Developmental plasticity has been suggested to facilitate the evolution of novelty and phenotypic diversity, but molecular mechanisms underlying this relationship are little understood. The nematodePristionchus pacificus is a model system for integrative evolutionary biology, combining laboratory studies with field work in ecology and population genetics. Pristionchus pacificusshows phenotypic plasticity in its feeding structures and is able to feed on fungi and other nematodes. Pristionchus executes one of two alternative mouth-forms (Eurystomatous vs. Stenostomatous) after an irreversible developmental decision that requires small molecule pheromones consisting of a blend of ascarosides and paratosides. Interestingly, dauer formation and mouth-form regulation require novel and specific compounds for their regulation. Downstream of small molecule pheromones and a conserved endocrine signaling module involving the nuclear hormone receptor DAF-12, we identified a novel regulator of plasticity eud-1. eud-1 acts in a developmental switch; eud-1 mutations eliminate one mouth form, whereas over-expression of eud-1 fixes it. EUD-1 is a sulfatase that acts dosage-dependently, is necessary and sufficient to control the sexual dimorphism of feeding forms, and has a conserved function in Pristionchus evolution. eud1 represents a novel gene that is much younger than the feeding dimorphism and has been added to a complex genetic network at a terminal position. It is epigenetically regulated in part by genes that are not conserved. These studies show that novel structures are determined by an unpredictable mix of conserved (co-opted) and novel determinants and that candidate gene approaches are of limited use for studying novel biological phenomena. INTERACTIONS OF DROUGHT STRESS, MELOIDOGYNE INCOGNITA AND ARBUSCULAR MYCORRHIZAL FUNGI ON SUGARCANE. Souza1, C.C.M., E.M.R. Pedrosa1, and U.M.T. Cavalcante2. 1Universidade Federal Rural de Pernambuco, Rua Dom Manoel de Medeiros, s/n, Recife, 52171-900, Pernambuco, Brazil; 2Universidade Federal de Pernambuco, Av. Prof. Moraes Rego, 1235 - Cidade Universita´ria, Recife/PE, 50670-901, Brazil. The objective of this study was evaluating effects of the interaction of drought stress [50 and 100% of pot water capacity (PWC)] and Meloidogyne incognita (0 and 12.000 eggs per pot)] and arbuscular mycorrhizal fungi – AMFs (Glomus etunicatum, Gigasporos albida, Acaulosporo longula and Scutellospora heterogama at 200 spores per pot in same proportion) on early morphological development and physiological activity of sugarcane (variety RB863129), as well as the effects on nematode and fungi multiplication. Sugarcane was inoculated with AMFs 20 days after seedlings acclimation in greenhouse and, with M. incognita, 30 days after the AMFs inoculation. Evaluations were carried out 45 days after nematode inoculation. Drought stress and M. incognita limit the initial vegetative development and fresh and dry biomass of sugarcane.

Abstracts 241 Drought stress also increased AMFs density in soil and roots. The AMFs increased sugarcane fresh stalk and root biomass but decreased gall and nematode numbers in roots. Drought stress as well M. incognita decreased ascorbate peroxidase contents in sugarcane, but the nematode infestation increased polyphenoloxidase and total soluble protein in the sugarcane plants. Drought stress decreased phosphorus content in the leaves and AMFs increased phosphorus content in stalk, however, the increase in phosphorus promoted by the AMFs did not occur under drought stress. Catalase, peroxidase and proline levels were not affected by drought stress, M. incognita and/or AMFs. NEMATODES AS BIOINDICATORS OF SOIL SUPPRESSIVENESS. Steel1,2, H. and H. Ferris1. 1Department of Entomology and Nematology, University of California Davis, One Shields Avenue, Davis, California 95616-8751, USA; 2 Department of Biology, Nematology Research Unit, Ghent University, K.L. Ledeganckstraat 31, 9000 Ghent. We tested the hypothesis that the numerical, biomass or functional abundance of predator nematodes, either specialists or generalists, are useful indicators of soil suppressiveness to opportunistic plant-feeding species. In concept, regulation or suppression of target pest species should be enhanced when an abundance of predator species is supported by ample availability of bacterial- fungal- and non-damaging plant-feeding prey species. Besides their direct effects on the prey, the predator nematodes are bioindicators of other soil organisms performing the same ecosystem services. We selected soils from natural and managed environments that represented different levels of resources and disturbance and introduced test prey not already resident in the soils (Meloidogyne incognita and Steinernema feltiae) into microcosm chambers of each soil either in its natural state or after heat defaunation. Survival of the test prey was determined after a 5 day bioassay exposure. Across the soils tested, predator abundance and biomass were greater in undisturbed soils with plentiful resources and lower in soils from agricultural sites. Suppressiveness to the two introduced species increased with both numerical abundance and metabolic footprint of the predator assemblages. The relationship between suppressiveness and metabolic footprint of the predators was best described by a logarithmic function which increased rapidly at first and then dampened to an asymptotic level possibly determined by temporal and spatial aspects of the bioassay system. The applied implications of this study are that soil suppressiveness to pest species may be enhanced by increasing resources, removing chemical and physical constraints to the survival and increase of predators, and altering management practices so that predators and target prey are colocated in time and space. HOST PLANT RESISTANCE IN GROUNDNUT TO DITYLENCHUS AFRICANUS. Steenkamp1, S., A.H. Mc Donald2 and D. de Waele3. 1Agricultural Research Council-Grain Crops Institute, Private Bag X1252, Potchefstroom, 2520 South Africa; 2Unit for Environmental Sciences and Management, North west University, Private Bag X 6001, Potchefstroom 2520, South Africa; 3Laboratory of Tropical Crop Improvement, Department of Biosystems, KU Leuven, Willem de Croylaan 42 – box 2455, 3001 Leuven, Belgium. Groundnut is an important cash crop for both commercial and small-scale farmers in South Africa. The main effect of Ditylenchus africanus on groundnut is qualitative, causing downgrading of groundnut consignments with significant financial losses for producers. This nematode is difficult to control with currently available management tools because of its high reproduction and damage potential. The objective of this study was to investigate the potential of host-plant resistance as an effective and economically-feasible alternative for the control of D. africanus on groundnut. Selected groundnut genotypes were evaluated in microplot and field trials over two consecutive growing seasons. PC254K1 and CG7 were identified as resistant to D. africanus under microplot and field conditions with resistance that was sustainable even under high infestation levels. These lines consistently produced high quality kernels throughout all of the trials and can therefore be used as major sources of resistance to D. africanus in groundnut breeding programmes for the development of D. africanusresistant groundnut cultivars. PC287K5 also showed a lesser degree of resistance and can be used in fields with a lower infestation rate. THE EFFICACY OF MULTIGUARD PROTECTÒ (A.I. FURFURAL) AS AN IN-SEASON TREATMENT FOR THE CONTROL OF CERTAIN NEMATODES ON TOMATOES, PEPPERS, CUCURBITS AND TURF UNDER FIELD CONDITIONS IN FLORIDA, USA. Steyn1, A., B. Booker2 and E. Buntting1. 1Illovo Sugar Ltd, South Africa, PO Box 31003, Merebank, 4059, Durban, South Africa; 2Florida Ag Research, 3001, North Kingsway Road, Thonotosassa, 33592, Florida, USA. With the phasing out or banning of certain toxic nematicides in the agricultural sector of many countries, the need for production of safer nematicides developed. Illovo Sugar South Africa has developed a number of new non systemic agricultural products derived from sustainable sugar cane resources. The active ingredient of these products is furfural, an aromatic aldehyde, which is naturally found in food crops and is also used in food flavouring. Furfural is a non systemic nematicide and breaks down fairly quick, making it ideal for use in all kinds of farming. The nematicide, Crop GuardÒ, is registered on multiple crops in South Africa while the nematicide, Multiguard ProtectÒ, is currently registered on turf, ornamentals and non-fruit bearing orchard crops in the USA. A number of efficacy trials on pre-plant application for tomatoes and peppers were executed in the United States of America to determine efficacy of Multiguard ProtectÒ on the control and

242 Journal of Nematology, Volume 46, No. 2, June 2014 suppression of certain plant parasitic nematodes. Results from field trials at Florida Ag Research station in Florida and from Pacific Ag in California, USA, have indicated that a single pre-plant treatment of 9 gal/A can control most nematodes during the early season. More trials are currently being executed on Multiguard ProtectÒ in the USA while registration on food crops is pending. THE EFFICACY OF MULTIGUARD PROTECTÒ (A.I. FURFURAL) AS A PRE-PLANT SOIL TREATMENT FOR THE CONTROL OF CERTAIN NEMATODES ON TOMATOES AND PEPPERS UNDER FIELD CONDITIONS IN FLORIDA, USA. Steyn1, A., B. Booker2, O. Cuevas3 and E. Buntting1. 1Illovo Sugar Ltd, South Africa, PO Box 31003, Merebank, 4059, Durban, South Africa; 2Florida Ag Research, 3001, North Kingsway Road, Thonotosassa, 33592, Florida, USA; 3Pacific Ag Research, 1840 Biddle Ranch Road, San Luis Obispo, 93401, California, USA. With the phasing out or banning of certain toxic nematicides in the agricultural sector of many countries, the need for production of safer nematicides developed. Illovo Sugar South Africa has developed a number of new non systemic agricultural products derived from sustainable sugar cane resources. The active ingredient of these products is furfural, an aromatic aldehyde, which is naturally found in food crops and is also used in food flavouring. Furfural is a non-systemic nematicide and breaks down fairly quick, making it ideal for use in all kinds of farming. The nematicide, Crop GuardÒ, is registered on multiple crops in South Africa while the nematicide, Multiguard ProtectÒ, is currently registered on turf, ornamentals and non-fruit bearing orchard crops in the USA. A number of efficacy trials on pre-plant application for tomatoes and peppers were executed in the United States of America to determine efficacy of Multiguard ProtectÒ on the control and suppression of certain plant parasitic nematodes. Results from field trials at Florida Ag Research station in Florida and from Pacific Ag in California, USA, have indicated that a single pre-plant treatment of 9 gal/A can control most nematodes during the early season. More trials are currently being executed on Multiguard ProtectÒ in the USA while registration on food crops is pending. VERIFICATION OF MELOIDOGYNE INCOGNITA RACE 2 RESISTANCE IN AMARANTHUS AND CAPSICUM GENOTYPES IN SOUTH AFRICA. Steyn1, W.P., M.S. Daneel1 and M.M. Slabbert2. 1Agricultural Research Council – Institute for Tropical and Sub Tropical Crops, Private Bag X11208, Nelspruit, 1200, South Africa; 2Department of Horticulture, Tshwane University of Technology, Private Bag X680, Pretoria, 0001, South Africa. Root-knot nematode resistance identified in crop genotypes in greenhouse studies should be verified for sustainability under natural environmental conditions. Therefore, the Amaranthus genotype ‘Local 33’ and Capsicum genotype ‘Tobasco’, identified in greenhouse studies as resistant to Meloidogyne. incognita race 2 together with susceptible Amaranthus and Capsicum genotypes ‘Bosbok Thepe’ and ‘Paprika’ respectively, were subjected to a range of initial nematode inoculum densities (Pi), namely 0 (control), 5 000, 20 000, 30 000 and 50 000 eggs and J2 per seedling at planting in a micro plot study. Reproduction factor values (Rf) were used as main criterion to evaluate for resistance. Results indicated that Amaranthus genotype, ‘Local 33’ displayed Rf-values of #1 at all inoculation levels. This confirms the resistance of this genotype against M. incognita race 2 as seen in the greenhouse trials. Capsicum genotype ‘Tobasco’ did however, not show resistance at all Pi levels used in this study. At inoculation levels of 30 000 and higher, ‘Tobasco’ exhibited Rf-values >1, indicating susceptibility to this root-knot nematode species. Strong relationships were obtained between the Pi and reproduction factor values for both Amaranthus and Capsicum genotypes and were described using the Logistic Dose Response Models (LDR) y=a+b/(1+(x/c)^d for both Amaranthus genotypes and y=a/(1+(x/b)^c for both Capsicum genotypes. Significant R2 values of 99% and 98% were obtained for Amaranthus genotypes ‘Bosbok Thepe’ and ‘Local 33’, respectively and 98% and 95% for Capsicum genotypes ‘‘Paprika’ and ‘Tobasco’, respectively. The need exists for more frequent and extensive screenings of these vegetable genotypes in order to provide resource-poor producers with better options for improved and sustainable yields. A NEW SUGARCANE FARMING SYSTEM IMPROVES SOIL HEALTH AND ENHANCES SUPPRESSIVENESS TO PLANT-PARASITIC NEMATODES. Stirling, G.R. Biological Crop Protection Pty. Ltd., 3601 Moggill Road, Moggill, Queensland, 4070, Australia. A multi-disciplinary team of scientists in Australia has shown that a sugarcane farming system based on permanent raised beds, residue retention, a leguminous rotation crop and controlled traffic using GPS guidance improves soil health and consistently increases yield. Soil physical properties improve markedly when beds are widened to accommodate the heavy machinery used to plant, harvest and transport the crop. The introduction of a non-grass rotation crop reduces populations of fungal and nematode pathogens that constrain productivity. A reduction in tillage increases earthworm populations, with consequential effects on macroporosity and water infiltration rates. Improved water capture in periods of low rainfall is another reason for increased yields, while improved percolation through macropores and retention of surface cover protects soils from erosion during intense tropical storms. From a biological perspective, suppressiveness to Pratylenchus zeae and Meloidogyne javanica, the main nematode pests of sugarcane, is enhanced by components of the new farming system. Populations of plant-parasitic nematodes do not resurge as strongly when tillage is eliminated, while a blanket of crop residue

Abstracts 243 on the soil surface creates a layer of soil that is amenable to root growth and highly suppressive to Pratylenchus (and possibly other root pathogens). Initial results suggest that numerous organisms (including nematophagous fungi and predatory nematodes) are contributing to the suppressiveness observed. Future research aims to determine whether predatory microarthropods also contribute regulatory services when compaction is removed through traffic control; and whether Pasteuria penetrans and Pasteuria thornei play a greater role in suppressing their hosts when soil is no longer tilled. POTENTIAL OF ENTOMOPATHOGENIC NEMATODES TO CONTROL WOOLLY APPLE APHID (ERIOSOMA LANIGERUM). Stokwe1,2, N.F. and A.P. Malan2. 1Stellenbosch University, Department of Conservation Ecology and Entomology, Private Bag X1, Matieland, Stellenbosch 7602, South Africa; 2Agricultural Research Council, ARC InfruitecNietvoorbij, Private Bag X 5026, Stellenbosch 7600, South Africa. Woolly apple aphid, Eriosoma lanigerum, is an important pest of apples, occurring throughout the apple-growing regions of the world. It feeds above ground, and on the roots of apple trees, causing hypertrophic gall formation, which affects the transport of water and nutrients. Entomopathogenic nematodes (EPN) of the two families, Steinernematidae and Heterorhabditidae, and their symbiotic bacteria have generated extensive interest as inundative biological control agents of insects. With the development of resistance of woolly apple aphid to certain chemicals, export restrictions, and the inability of parasitoids to control the aphid successfully early in the season, it is important to consider EPN as an alternative biocontrol agent. Apple roots from an orchard, infested with woolly apple aphids, were kept in large plastic containers to maintain a steady supply of insects. Seven EPN species were tested for their pathogenicity against E. lanigerum, includingSteinernema khoisanae, Steinernema yirgalemense, Steinernema citrae, Steinernema feltiae, Heterorhabditis zealandica, Heterorhabditis bacteriophora, and Heterorhabditis safricana. Laboratory bioassays identified S. yirgalemense and H. zealandica as being the most virulent against the subterranean stage of the woolly apple aphid, with a mortality rate of 48% and 38%, respectively. Studies on the effect of wooly apple aphid size showed that the last instar is most susceptible to infection by S. yirgalemense and H. zealandica, whereas smaller instars appear to be too small for nematode penetration and infection. The haemolymph of woolly aphid apple showed an inhibitory effect on the development of the symbiotic bacteria Xenorhabdus sp. and Photorhabdus sp., which are associated with S. yirgalemense and H. zealandica, respectively. THE IMPORTANCE OF PRATYLENCHUS SPP. ON THE FIELD TO FORK PRODUCTION CHAIN OF APPLE TREES IN THE WESTERN CAPE, SOUTH AFRICA. Storey, S.G. and L. van der Walt. Nemlab, cnr. of R44 and Anyswortelrug Rd, Klapmuts, 7625, South Africa. Agriculture forms a key component of the Western Cape’s economy. The export of fresh fruit is vital to this economy as it brings in much needed foreign currency and ensures job security. The production of pome fruit (apples and pears) is the largest sector of the agricultural fruit production sector. The export of fruit to Europe, China, Russia and Japan provides important foreign exchange. Of the total apple production during the 2011/2012 growing season exports amounted to 45.34%. The field-to-fork chain in apple production must be maximised to ensure the sustainability of the economic wellbeing of this sector. Nematodes impact heavily on various facets of this chain. Pratylenchus spp. play a key role in the demise of nursery trees, apple replant disease and the size of the fruit exported. The impact is already measurable at nursery level, as affected trees have to be held back as the stems are not yet strong enough to sell to producers. When the trees are planted they are more susceptible to apple replant disease. Trees with this disease bear fruit 2-3 years later than uninfected trees and also have lower yields throughout their shortened life span. Once bearing, potassium-uptake is affected by Pratylenchus, which is directly linked to fruit size. Export prices are determined by size, thus direct impact on income is expected. GlobalGAP and European retailers have strict regulations regarding minimum residue levels making control strategies complex. THE EFFECT OF CONSERVATION AGRICULTURE ON NEMATODE POPULATIONS AND SOIL MICROBIAL COMMUNITY DYNAMICS. Swart1, A., M. Marais1, and J. Habig2. 1National Collection of Nematodes, Biosystematics Programme, ARC-Plant Protection Research Institute, Private Bag X134, Queenswood 0121, South Africa; 2Soil Microbiology, Soil Microbiology and Plant Pathology Programme, ARC-Plant Protection Research Institute, Private Bag X134, Queenswood 0121, South Africa. The ARC Zeekoegat experimental farm at Roodeplaat, Gauteng, was selected for a conservation agriculture trial in which several conservation agriculture practices were tested. A multidisciplinary approach was followed and the focus area included soil, and other biological parameters. The key plots examined included six treatments consisting of maize monoculture, maize and soybean rotation, and maize and cowpea intercropping. All these treatments were combined with either low of high fertiliser levels, and either reduced or conventional tillage. Nematodes and soil microbial communities play important roles in nutrient cycling and have been used as indicators of soil health. Population numbers of different nematode trophic groups were compared with each other during the 2010-2013 seasons. In most treatments a decline in the incidence and population numbers of free-living nematodes were noticed from 201-2013. Overall, the reduced tillage treatments show a slightly higher percentage of fungivores than bacterivores, while the conventional tillage plots slightly favoured bacterivores over fungivores. The population numbers of the free-living nematodes in conventional tilled plots were generally higher than

244 Journal of Nematology, Volume 46, No. 2, June 2014 those in reduced tillage plots. Of concern is the very high population numbers of herbivores compared to free-living nematodes in all the treatments. As mentioned previously, soil microbial community diversity and activity play an integral part in maintaining ecosystem function. Carbon source utilisation profiles indicated differences between different rotation systems and tillage practices. Overall enzyme activity was higher under reduced tillage practices, compares to conventional tillage practices. This presentation will combine all findings on nematodes as well as soil microbial data. FORMULATING A NEW VISION AND DIRECTING CHANGE IN NEMATOLOGY EDUCATION IN SUBSAHARAN AFRICA TO REFLECT A SCIENCE OF THE 21ST CENTURY. Talwana, H. Department of Agricultural Production College of Agricultural and Environmental Sciences Makerere University. P.O. Box 7062 Kampala Uganda. Nematology is a fascinating cross-disciplinary science. However, in sub-Saharan Africa, Nematology is at crossroads or lost completely due to lack of a critical mass of nematologists; nematologists rarely working full time on the nematode problems; trained nematologists rarely remaining in the discipline, and most importantly, higher learning institutions in Africa are not teaching Nematology at all and/or where offered, it is taught as part of Plant Pathology or Entomology. Consequently, there is a general lack of awareness of even the existence of nematodes by the public – policy makers, farmers, formative education teachers, etc. On the other hand, there is growing apathy in today’s students of Biological Sciences about Nematology, which in most cases is a result of the content and style of delivery by trainers. With little practice in laboratories and in the field due to lack of appropriate equipment, limited local contemporary examples due to low research output, and limited inference to modern day biosciences trends and achievements, there is a negative perception of the applicability and transferability of Nematology to ‘real-world’ situations and employment opportunities. Thus, even the most interested students are turned away. This calls for change in the way the content of Nematology is delivered and the onus is on the trainers to inspire and entice a new generation into Nematology as an ultimate career path. The opportunity that the convergence of advances in Nematology and emerging (bio)-technologies provides should particularly be harnessed to deliver Nematology as a model biological science. TOWARDS A REFERENCE TRANSCRIPTOME OF ANGIOSTRONGYLUS CANTONENSIS. Tang1,2, P., T-W. Chen2, C-C. Lee2, K-Y. Chen1, S-H. Chang1, J.-W. Shin3 and L-C. Wang1. 1Department of Parasitology, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; 2Bioinformatics Center, Chang Gung University, Taoyuan 333, Taiwan; 3 Department of Parasitology, National Cheng Kung University, Tainan 701, Taiwan. Although Angiostrongylus cantonensis is the most common cause of eosinophilic meningitis in Southeast Asia and the Pacific Basin, relatively little is known about this neglected parasite at molecular and genomic level. In order to understand the regulation of development, adaptations to different microenvironments, and the interaction between the host, the Chang Gung University initiated an A. cantonensis systems biology project that aimed to study the genome, transcriptome and proteome of this parasite. For the past two years, we have performed pilot studies on gene, miRNA and protein expression profiling. In the present investigation, we focus on the generation of a reference transcriptome of A. cantonensis. More than 300 million paired-end reads (28Gbp) were generated from the Illumina Hiseq sequencing platform from different developing stages. Assembling of these reads resulted in more than 44,000 contigs with a N50 of 1,023bp. BLAST search revealed that 31.5% of these putative transcripts showed significant sequence homology to known protein or protein motifs/domains. Functional classifications indicated that most of these putative transcripts are involved in embryo development or egg hatching, larval development, regulation of growth rate and locomotion. The protein expression levels of these transcripts were verified by using two-dimensional electrophoresis and LC/MS/MS. The reference transcriptome established in the present study will provide a foundation for the ongoing whole genome sequencing project and comparative transcriptomic/ proteomic analyses. COMPARISON OF EXTRACTION METHODS FOR EXTRACTION OF MIGRATORY ENDOPARASITIC NEMATODES FROM ROOTS. Tangchitsomkid1, N., T. Chanmalee2 and M. Hodda3. 1Biotechnology Research and Development Office, Department of Agriculture, Chatuchak 10900 Bangkok, Thailand; 2Centre for Agricultural Biotechnology, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140, Thailand; 3CSIRO Ecosystem Sciences, GPO Box 1700 Canberra, ACT 2601, Australia. This poster reports tests of the efficiency of ultrasonic methods in extracting the nematode, Radopholus similis, from the roots of the aquatic plant, Anubias sp. The first test compared different extraction methods. In water, with a duration of 10 minutes and frequency of 50/60 KHz, ultrasonic extraction recovered a little more than three times the number of nematodes as a mist chamber running for 48 hours (a mean of 26.4 nematodes versus 8.2 nematodes). The second test compared the extraction rate and damage to both plant roots and extracted nematodes at different durations of sonication. Duration times of 5, 10, 20, 40 and 60 minutes were investigated. Minimal root damage and nearly maximal extraction rates were obtained after sonication for 20 minutes, allowing for the plants to be replanted and the nematodes to be identified easily. The ultrasonic technique can require less time and effort for root preparation than most other techniques for migratory endoparasites, yet has a similar or superior extraction efficiency per sample, thus allowing more samples to be processed more cheaply.

Abstracts 245 A NEW NEMATODE EXTRACTION KIT FOR FIELD WORK USING ULTRASONIC TECHNOLOGY. Tangchitsomkid1, N., T. Chanmalee2 and M. Hodda3. 1Biotechnology Research and Development Office, Department of Agriculture, Chatuchak 10900 Bangkok, Thailand; 2Centre for Agricultural Biotechnology, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140 Thailand; 3CSIRO Ecosystem Sciences, GPO Box 1700 Canberra, ACT 2601 Australia. An efficient, easy-to-use extraction kit was developed for extracting root migratory endoparasites—such as Hirshmanniella, Radopholus and Pratylenchus—in the field. Ultrasonic waves in a water medium at 40 KHz causes alternate compression and decompression inducing nematodes to leave the roots and enter the water within 20 minutes. Consolidated into a single unit for fieldwork, the kit is a cube of about 20 cm size, made of stainless steel for durability. It has a water chamber of 9 cm wide by 15 cm long by 6 cm deep, with a single-frequency ultrasonic wave generator connected to the bottom. It has an automatic timer and operates at 220 volts. It is also equipped with a mini microscope fixed at 50 x magnification 10 cm long. The kit comes with a carry bag into which it fits for convenient transportation. The kit can be used by farmers to detect nematode outbreaks in cultivated fields, especially in the roots of plants for export (such as aquatic plants, Philodendron species, orchids, Anthurium species, Zamioculcas zamiifolia and other ornamental plants). The kit is also suitable for quarantine inspection and certification to enable rapid examination and detection of nematodes, thus saving both time and expense. The kit was tested and compared with other methods available, with extraction efficiencies equal to, or better than, the sucrose-centrifugation and mist-chamber techniques. However, we consider it more practical and convenient than either. The kit and its basic technology was invented in Thailand and is now patent pending (Application no.1101000659). ULTRASONIC EXTRACTION OF PLANT-PARASITIC NEMATODES FROM PLANT ROOTS. Tangchitsomkid1, N., T. Chanmalee2 and M. Hodda3. 1Plant Protection Research and Development Office Department of Agriculture, 50 Phaholyothin Road, Chatuchak, Bangkok, 10900 Thailand; 2Kasetsart University, Kamphaeng Saen Campus, 1 Malaiman Rd, Kamphaeng Saen District, Nakhon Pathom 73140, Thailand; 3CSIRO Ecosystem Sciences, GPO Box 1700, Canberra ACT 2601 Australia. Extraction of nematodes from plant roots is a very time- and labour-intensive process, so new methods offering advantages for either speeding or lessening workloads are highly desired, but must be tested before adoption. A novel method using ultrasonic waves is fast, simple and non-destructive to roots. The method was tested and optimized for extraction of burrowing nematodes (Radopholus similis) from the roots of the aquatic plant Anubias sp. in water. Different durations of treatment and ultrasonic frequencies were tested, and the number of nematodes and plant damage recorded. The highest extraction efficiencies per sample were obtained after 40 minutes or longer sonication at a frequency of 40 KHz, but the condition of the extracted nematodes was poor and not suitable for identification.. Considering efficiency as number of nematodes extracted per unit time, optimal extraction was obtained with a duration of 20 minutes. With this duration there was little observable damage to the nematodes (allowing use in experiments or inoculation trials), or the plants (so that successful replanting is possible). The ultrasonic method was compared with other extraction techniques, and was much more efficient than root submersion, shaking or mist chamber techniques, but a little less efficient per sample than the sucrosecentrifuge method. Ultrasonic extraction requires less time and effort for root preparation than other methods, so it can be a lower cost method for routine use. Furthermore, it can be incorporated into a simple, cheap, mass-manufactured, portable and automated kit, which can be made widely available. DIFFERENT CITRUS JUICES AND OILS AS SOIL AMENDMENTS USED IN CONTROL OF MELOIDOGYNE INCOGNITA ON TOMATO. Tefu, G., M.S. Daneel, W.P Steyn, C. Arries and T. Selabela. Agricultural Research CouncilInstitute for Tropical and Subtropical Crops, Private Bag X11208, Nelspruit 1200, South Africa. Glasshouse experiments were conducted to evaluate the effect of soil amendments with extracted juices and oils of different citrus plants (grapefruit, lemon, lime, naartjie and sweet orange) for the control of Meloidogyne incognita. Soil was inoculated with 3000 root-knot nematode eggs after which the treatments were applied and 3 days later tomato seedlings were transplanted in each bag. The height of the tomato plants was measured bi-weekly and root weight, plant mass, root system, gall index and nematode numbers were determined at 56 days after transplanting. The organic amendment consisting of orange juice gave persistently the best improvement in growth as well as best reduction of nematodes. Citrus oils consistently performed weaker than the juice and a clear difference was observed between the different juices with orange followed by naartjie, grapefruit, lime and lemon. Lime and lemon were similar to the control. Further studies are required to confirm the potential of these organic amendments in the control of Meloidogyne incognita in the field. THE EFFECT OF TEMPERATURE AND TIME ON THE POPULATION DYNAMICS OF MELOIDOGYNE. CHITWOODI IN POTATO TUBERS DURING STORAGE. Teklu, M.G., T.H. Been, C.H. Schomaker and J.E. Beniers. Plant Research International, Wageningen University and Research Centre, Plant Sciences Group, 6700 AA, Wageningen, The Netherlands.

246 Journal of Nematology, Volume 46, No. 2, June 2014 This experiment was carried out to assess the population dynamics of Meloidogyne chitwoodi in tubers under different storage conditions used in The Netherlands. Infected tubers were produced by growing the susceptible cv. De´sire´e in 100, 10 kg pots at a density (Pi) of = 12 J2 g dry soil-1. Infected tubers of all pots were combined and sorted into classes based on their tuber-knot index. Thirteen batches of infected tubers were composed of equal amounts of tubers from each index. The infected tubers were stored at 98% relative humidity at temperatures of 4, 8 and 12 8C and nematode numbers were assessed after 0, 2, 4, 6 and 8 months storage time. After hardening, peeling and mixing of the skins, replications were placed in the spray-mist chamber for 7 weeks to extract the juveniles. The logistic model was fitted to the cumulative hatch to follow the hatching process of M. chitwoodi from the peel and the estimated maximum cumulative hatch was used to evaluate the treatments. Both storage temperature and time influenced the population dynamics of M. chitwoodi in potato tubers. Storage temperatures of 4 and 8o C reduced populations of M. chitwoodi up to 96% and 89% after 8 months of storage. However, at 128C the population density remained at the same level over time. Recovered juveniles of M. chitwoodi from tubers after 6 and 8 months of storage, at all three temperatures, proved to be still infective on the susceptible cv. De´sire´e. TESTING FOR TUBER RESISTANCE AGAINST M. CHITWOODI IN PARTIALLY RESISTANT POTATO GENOTYPES WITH AVENA STRIGOSA. Teklu1, M.G., T.H. Been1, C.H. Schomaker1, L.P.G. Molendijk2, T.G. van Beers2 and J. E. Beniers1. 1Plant Research International, Wageningen University and Research Centre, Plant Sciences Group, 6700 AA, Wageningen, The Netherlands; 2Applied Plant Research International, 8219 PH Lelystad, The Netherlands. A pilot experiment was carried out using two partially resistant genotypes AR04-4096, 2011M1 and the control cv. De´sire´e to test whether these genotypes possess both tuber and root resistance. Four plants of Avena strigosa, and one potato plant, consisting of a single sprout, were sown in 10 kg pots. The resistant genotypes were tested with and without A. strigosa and cv. De´sire´e was used as a susceptible control. Pots were inoculated with a moderate nematode density: 16 J2 g dry soil-1. Final population densities were estimated from the soil, roots and tuber peel. The roots and peel were incubated in a spraymist chamber for 4 and 7 weeks, respectively. The numbers of nematodes extracted from soil, roots and tubers in the pots with A. strigosa and the resistant genotypes increased compared to those without A. strigosa. The relative susceptibilities (%RS) of the genotypes AR04-4096 and 2011M1, without A. strigosa, were 0.18 and 0.52, respectively, which is consistent with previous results. The presence of A. strigosa increased the relative combined host status (RH) for AR04-4096 compared to De´sire´e till 25%. Avena strigosa slightly increased the estimated relative susceptibility of tubers of the tested genotypes: %RST of AR04-4096 increased from 0 to 0.04%; that of 2011M1 from 0.56 to 2.96%. The negative effect of A. strigosa on the percentage of nematode-free tubers of AR04-4096 was negligible, but nematode-free tubers decreased from 84% to 21% in 2011M1. It can be concluded that the tested partially resistant genotypes possessed partial resistance in both roots and tubers. SURVEY FOR THE SOYBEAN CYST NEMATODE IN MANITOBA, CANADA. Tenuta1, M., M. Madani1 and D. Lange2. 1Department of Soil Science, University of Manitoba, Winnipeg, MB, R3T 2N2, Canada; 2Manitoba Agriculture, Food and Rural Development, Altona, MB, R0B 0B0, Canada. We report the findings of a survey conducted in 2012/2013 of soybean fields for the soybean cyst nematode (Heterodera glycines) in Manitoba. In total, 48 fields were sampled with fields prone to infestation selected. Fields were sectioned into areas that could be responsible for pest introduction. In total, 282 composite soil samples were analyzed from 48 fields. Overall, 37 of the samples from 22 fields had nematode cysts. Sixty cysts were recovered. Most cysts were heavily damaged but 26 were sufficient for examination. Of these, 23 had circumfenestrate vulval cone structures. Three cysts had bifenestrate vulval cone structures characteristic of the genus Heterodera. Not all the cysts contained nematodes either as juveniles or eggs. Most were empty except for 15 circumfenestrate and one bifenestrate cysts. These 16 cysts were then followed up with molecular analyses by ITS sequencing, three SCN species-specific PCR approaches (Ou et al. 1993, Subbotin et al. 2001, Madani et al. 2013), and in one case, by 28s rDNA gene sequencing. ITS sequencing was possible for 14 of the circumfenestrate cysts. These best matched the sequence database for the cyst nematode, Cactodera. Only one of the bifenestrate cysts yielded DNA for analysis and its identification was ambiguous between H, glycines and Cactodera. Cactodera is not a pest of soybean or other crop plants in Manitoba. These cyst nematodes are likely naturally present, living on weeds and grasses in the sampled fields. The findings of the current study mean H. glycines has still not been detected in Manitoba. DETERMINING THE SPECIES OF STEM NEMATODE ON GRAIN PEA AND CREEPING THISTLE IN THE CANADIAN PRAIRIE PROVINCES. Tenuta1, M., M. Madani1, S. Briar1, and S.A. Subbotin2. 1Department of Soil Science, University of Manitoba, Winnipeg, MB, R3T 2N2 Canada; 2Plant Pest Diagnostic Center, California Department of Food and Agriculture, Sacramento, CA, 95832-1448, USA. The stem nematode (Ditylenchus dipsaci) was first described as a parasite of creeping thistle (Cirsium arvense) in the Canadian Prairied in 1979. Recently, Ditylenchus on creeping thistle in Russia was shown to be Ditylenchus weischeri, which is not an agricultural pest. This study examined if D. weischeri and not D. dipsaci is present in yellow pea (Pisum sativum) grain harvest samples in 2009 and 2010 and from creeping thistle in Saskatchewan, Alberta and Manitoba. Samples from 538

Abstracts 247 fields (mainly yellow pea) were provided by 151 pea growers. Two percent of the samples were Ditylenchus positive. Numbers ranged from 4-1500 nematodes kg-1 pea and were related to presence of creeping thistle seeds. Positive samples occurred in 2009 but not 2010 and were from different geographical locations. The nematode was morphologically dissimilar to D. dipsaci. Creeping thistle plants were then collected from yellow pea fields in the Provinces. All plants from Saskatchewan and Manitoba were infested with Ditylenchus. The ITS-rRNA regions ITS1, 5.8S and ITS2 was amplified using the TW 81 and AB 28 primer set. For some samples the ITS1 and ITS2 were separately amplified using TW81 and Dit58SR or Dit58SF and AB28 primers, respectively. A fragment of hsp90 was also obtained with U831 and L1110 primers. PCR-ITS products were subjected to RFLP using Bsh1236I, HinfI, MspI, RsaI and TaqI. The results match those published for D. weischeri. The results indicate the stem nematode found in yellow pea grain is D. weischeri and that creeping thistle seeds in grain its source. A TRANSCRIPTOMIC STUDY OF COLD TOLERANCE MECHANISMS IN THE ANTARCTIC NEMATODE PANAGROLAIMUS DAVIDI. Thorne1, M., M. Clark1, C. Marshall2 and D. Wharton3. 1British Antarctic Survey, Cambridge, UK; 2Department of Biochemistry, University of Otago; 3Department of Zoology, University of Otago, PO Box 56, Dunedin, NZ. Panagrolaimus davidi is an Antarctic nematode from coastal terrestrial habitats of the McMurdo Sound region, where it survives both desiccation and freezing. It can survive complete water loss (anhydrobiosis) and is the only organism known to survive intracellular ice formation. It has several cold tolerance strategies, including; freeze avoidance, cryoprotective dehydration, freezing tolerance and anhydrobiosis. We have conducted a study to determine gene expression during cold acclimation (+58C), freezing at -18C (which induces cryoprotective dehydration), freezing at -108C (which produces intracellular freezing) and during recovery from freezing. After exposure to these conditions the total RNA was extracted and sequenced on on an Illumina HiSeq 2000 resulting in 286,447,212 paired-end reads of >100bp, after quality control, and a transcriptome constructed from the data from all the treatments. To determine upregulation in specific treatments, expression levels were compared by fold difference and a ratio test of mapping counts. I will present here the data on acclimation and cryoprotective dehydration. Most prominent amongst genes upregulated during acclimation are those associated with cuticle synthesis and embryonic development. This is surprising since there is no population growth in cultures below +6.88C. Maintenance of cuticle structure may be important for preventing inoculative freezing during cryoprotective dehydration. Genes upregulated during freezing at -18C, and recovery from freezing, include those associated with enzymes and transport, signalling and control, and lipid metabolism. Genes associated with stress responses are present in the transcriptome, suggesting that the main survival mechanisms are expressed constitutively. SEASONAL VARIATIONS OF NEMATODE ASSEMBLAGES AND DIVERSITY IN VERTISOLS, CAMBISOLS AND ARENOSOLS SOIL GROUPS IN KENYA. Thuo1, A.K., J.W. Kimenju1, G.M. Kariuki2, G.N. Karuku1, P.K. Wendot2 and H. Melakeberhan3. 1Department of Plant Science and Crop Protection, University of Nairobi, P.O Box 30197, G.P.O, Kenya; 2Kenyatta University, Department of Agricultural Science and Technology, P.O. Box 43844-00100 Nairobi, Kenya; 3Agricultural Nematology Laboratory, Department of Horticulture, Michigan State University (MSU), East Lansing, Michigan 48824, USA. Soil health assessment has been based on narrow disciplinary approaches that overlook the multiple and interacting biological processes that are the basis of sustainable crop productivity. This study was carried out to determine the influence of seasonal variations in nematode assemblages in different soil groups, sites and disturbance levels as an indicator of soil health. Sampling was done in areas characterized by small scale subsistence agriculture in Kenyan northern and southern sites over three distinct seasons. The sampling points included disturbed (tilled) and the adjoining undisturbed (untilled) soils within three soil groups, namely vertisols, cambisols and arenosols. Nematodes were extracted using the centrifugal-floatation technique, enumerated and assigned to their respective trophic groups. Total nematode abundance in the three seasons varied significantly with a mean of 68.4, 93.1 and 51.6 nematodes in 200 cm3 of soil in season 1, season 2 and season 3, respectively. Nematodes abundance in the undisturbed soils was higher (97.74) compared to the disturbed soils (62.08 nematodes per 200 cm3. Mean abundance of nematodes was highest in Ccmbisols. In addition, nematode abundances, in all trophic levels across the three seasons, were significantly higher in the northern compared to the southern sites. Bacterivores (28%) had the highest incidence followed by herbivores (27%) and fungivores (21%) while omnivores (11%) had the least. The study demonstrated that nematode communities do vary in the different soil groups and between seasons, suggesting that they can be utilized as viable bio-indicators of soil health. EFFECT OF TILLAGE AND FUMIGATION ON PASTEURIA PENETRANS. Timper, P. USDA ARS, P.O. Box 748, Tifton, Georgia 31794, USA. The endospore-forming bacterium Pasteuria penetrans is a parasite of Meloidogyne spp. In this study, the effect of tillage and the fumigant 1,3-dichloropropene (1,3-D) on numbers of P. penetrans and suppression of M. incognita was evaluated from 2011-2013. A split-plot experiment was established in a field infested with both the nematode and bacterium. The main

248 Journal of Nematology, Volume 46, No. 2, June 2014 plots were tillage (strip vs conventional) with five fumigation treatments in 4-year sequences within each tillage treatment (C=no fumigant; F=fumigant): C-C-C-C, F-F-F-F, F-C-F-C, F-F-C-F, and C-F-F-C. Abundance of P. penetrans endospores was determined in the spring after tillage/fumigation and average root-gall indices on cotton plants were determined at the end of the season. Additionally, a greenhouse bioassay was conducted to determine reproductive potential of M. incognita in soil collected from each plot at planting time. Abundance of P. penetrans was greater in C-C-C-C than in most of the fumigated treatments and greater in strip tillage than in conventional tillage in 2012 and 2013. Nevertheless, the reproductive potential of M. incognita in the greenhouse was not affected by the fumigation or tillage treatments. Endospore abundance was greater in 2012 than in the other two years and was negatively correlated with reproductive potential only in that year. In the no-fumigation treatment, endospore abundance was negatively correlated with gall index indicating that P. penetrans was contributing to nematode suppression. Tillage mixes and redistributes soil and may move endospores away from the planting furrow. Although1,3-D had no direct effect on P. penetrans, it reduced the number of available hosts leading to lower endospore numbers. DISTINCTION OF ’TRYPOPHLOEI’ GROUP - A NEW TYPOLOGICAL GROUP AMONG WOOD NEMATODES OF THE GENUS BURSAPHELENCHUS FUCHS, 1937. Tomalak, M., A. Filipak, T. Malewski and J. Pomorski. Department of Biological Pest Control and Quarantine, Institute of Plant Protection - National Research Institute, Wladys1awa Wegorka 20, 60-318 Poznan, Poland. The nearly 90 known species of the genus Bursaphelenchus Fuchs, 1937 have been divided into 13 typological groups. Bursaphelenchus xylophilus is considered the most dangerous pest of pine trees worldwide and the xylophilus group seems to be the most extensively studied. At present it contains 15 species characterized by a relatively uniform set of distinctive morphological characters, such as the morphology of male spicules, extended anterior vulval lip, lateral fields with four incisures, and number and arrangement of male caudal papillae. Recent inclusion of new species gradually widened the morphological variation within this group. Additional data on the nematode genetics and ecology clearly indicate that species presently assigned to the xylophilus group may represent two closely related phylogenetic groups which also differ in their ecological relations with vector insects. We therefore suggest the establishment of a new typological group designated as trypophloei. This group would comprise species located in a separate but adjacent to main xylophilus group clade, and at the present it would include Bursaphelenchus trypophloei, Bursaphelenchus masseyi, Bursaphelenchus tokyoensis, Bursaphelenchus fagi, and two other species, presently under description, which are located in the same phylogenetic clade. These nematodes are morphologically most similar to the xylophilus group with some variation in spicule morphology. They all are associated with bark beetles (Curculionidae: Scolytinae). In contrast, all species of the classical xylophilus group with known insect association use long horned beetles (Cerambycidae) as vectors. MOLECULAR CHARACTERIZATION OF HETERODERA SPP AND PRATYLENCHUS SPP. FROM WINTER WHEAT IN THE EAST ANATOLIAN REGION OF TURKEY. Toktay1, H., M. Imren2, A. Ocal3, E.B. Kasapoglu4, A.A. Dababad5 and H. Elekcioglu4. 1University of Nigde, Faculty of Agricultural Sciences and Technologies, Nigde, _ Turkey; 2 Abant Izzet Baysal University, Faculty of Agriculture, Department of Plant Protection Bolu, Turkey; 3West _ Mediterranean Agricultural Research Institute. Antalya, Turkey; 4University of C xukurova, Faculty of Agriculture, Department of Plant Protection, 01360, Adana Turkey, Adana, Turkey; 5CIMMYT (International Maize and Wheat Improvement Centre), Wheat Improvement Program, Ankara, Turkey. Turkey is one of the top ten wheat producing countries in the world, with the East Anatoloian region being one of the most important winter wheat areas in the country. Plant parasitic nematodes are one of most important biotic agents that cause economical yield loss in wheat growing regions. Cereal cyst nematodes (CCN) and root lesion nematodes (RLN) are some of the most important pest of wheat worldwide. Limited knowledge of the nematode pests of wheat exists in Turkey. This study aims to characterize Heterodera and Pratylenchus species in this region by using morphological and molecular tools and to establish phylogenetic relationships between the populations. Forty-three Heterodera and thirty Pratylenchus populations were identified using sequence analysis of the Internal Transcribed Spacer region of the ribosomal DNA (ITS-rDNA), phylogenetic analysis using ITS-rDNA sequences, and morphological methods. Forty poulations were identified as Heterodera filipjevi and five populations as H. Latipons. Pratylenchus species identified were P. thornei, P.penetrans and P. neglectus. DEVELOPMENT OF QPCR ASSAYS FOR QUANTITATIVE DETECTION OF HETERODERA AVENAE, H. FILIPJEVI AND H. LATIPONS. Toumi1,2, F., L. Waeyenberge1,2, N. Viaene1,3, A. Dababat4, J.M. Nicol4, F.C. Ogbonnaya5 and M. Moens1,2. 1Institute for Agricultural and Fisheries Research (ILVO), Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium; 2 Faculty of Bio engineering, Ghent University, Coupure links 653, 9000 Ghent, Belgium; 3Faculty of Sciences, Ghent University, K. L. Ledeganckstraat 35, 9000 Ghent, Belgium; 4International Maize and Wheat Improvement Centre (CIMMYT), Ankara, Turkey; 5International Center for Agricultural Research in the Dry Areas, PO Box 5466, Aleppo, Syria.

Abstracts 249 Heterodera avenae, H. filipjevi and H. latipons are considered of major economic significance in cereals. Precise identification and quantification of these nematodes are necessary to develop effective integrated pest control. We report the results of a qPCR assay that we developed for the quick detection and quantification of the three species. Three qPCR primer sets comprising two primers and a probe, were designed and optimized. All developed assays were able to detect a single second-stage juvenile (J2). Their specificity was confirmed by the lack of amplification of 13 other Heterodera species. A qPCR using DNA extracted from 106, 114 and 114 J2 + eggs of H. avenae, H. filipjevi and H. latipons resulted in steady Ctvalues (Ct = 22.33 ± 0.1, Ct = 21.83 ± 0.05 and Ct= 18.6 ± 0.12, respectively). Dilution series of DNA extracted from known numbers of J2 + eggs of the three species were made. The assays resulted in a standard curve showing a highly significant linearity between the Ct-values and the dilution rates (R2 = 0.99; slope = -3.05, R2 = 0.99; slope = -3.4 and R2 = 0.99; slope = -3.5 for H. avenae, H. filipjevi and H. latipons respectively). The three qPCR assays provide a sensitive and valid tool for the rapid detection and quantification of the three species whether they occur alone or in mixtures with other species. Unfortunately, the assay for the detection and quantification of H. filipjevi was not successful for all H. filipjevi populations, which could be contributed to DNA polymorphism. DEVELOPMENT OF REAL-TIME PCR PRIMERS FOR MAJOR PLANT PARASITIC NEMATODES IN RICE FIELDS IN MYANMAR. Toyota1, K. and Y.Y. Min2. 1Tokyo University of Agriculture and Technology, Japan; 2Yezin Agricultural University, Japan. Myanmar has 100% self-sufficiency rate of cereals and is the seventh largest rice producing country. Rice yield increased from 1997 to 2007 in major rice producing countries, while decreasing in Myanmar, suggesting the presence of obstacles against rice production. The objectives of this study were to identify major plant-parasitic nematodes in rice fields in Myanmar and to estimate possible damage to yield. Soil was collected from 26 paddy fields in different regions in Myanmar and nematodes were extracted with the Baermann-funnel method. The D2/D3 regions of 28S rDNA was amplified from individual plant-parasitic nematodes and sequenced. The rice root nematode Hirschmanniella oryzae and the root-knot nematode Meloidogyne graminicola were identified. In addition, the rice stem nematode (ufra) Ditylenchus angustus was also detected in some fields. Then, real-time PCR primers were designed to quantify the species in soil. H. oryzae and M. graminocola were detected in 23 and 3 fields, respectively. Nematicide was applied into two paddy fields infested with H. oryzae and D. angustus to estimate potential damage by nematodes. The results showed 20% to 70% increase in the grain yields by nematicide application. This study enabled the quantification of the major plant parasitic nematodes in Myanmar, and suggested that nematodes cause a significant reduction in rice yields in certain regions of Myanmar. TRANSGENIC NEMATODE RESISTANCE FOR AFRICAN FOOD SECURITY: NEMATODE RESISTANT BANANAS AS A CASE STUDY. Tripathi1, L., H. Roderick2, A. Babirye1 and H.J. Atkinson2. 1International Institute of Tropical Agriculture, PO Box 7878, Kampala, Uganda; 2Centre for Plant Sciences, University of Leeds, Leeds LS2 9JT, UK. Transgenic crops can support future food security when they overcome agronomic challenges refractory to other approaches. Examples include nematode resistant plantains and cooking bananas for Africa that prevent current losses of up to 70%. The crop’s sterility enhances both the biosafety of this biotechnological intervention and its rate of progress relative to that of classical plant breeding. Our efficient genetic transformation system for plantain produced over 200 transgenic lines expressing a maize cystatin that limits nematode feeding success and/or a synthetic peptide that suppresses root invasion. Nematode challenge of plantlets in the glasshouse identified lines with promising levels of resistance. Twelve of these lines were grown for an authorised, contained field trial in Uganda. Nematodes from infected banana roots were added to the pot soil of selected lines before transplanting to the field in a randomised block design. Subsequent plant growth was measured non-destructively using digital hemispherical photography to calculate the leaf area index (LAI). Radopholus similis dominated the nematode population at 7 months post-planting and caused severe necrosis to control plant roots. Several transgenic lines had significantly larger LAI values and less root necrosis than the control non-transgenic plants. A significant reduction in recovered nematodes for three lines corresponded to 89-98% resistance. The experiment is continuing for a further 2-3 harvests. Plantains and cooking bananas are a vital food for about 100 million Africans. The future challenges for this not-for-profit public research are to ensure biosafe uptake for plantain and then application to other subsistence crops that suffer appreciable nematode damage. INTEGRATED MANAGEMENT OF ROOT-KNOT NEMATODES ON TOMATO WITH ARBUSCULAR MYCORRHIZAL FUNGI, PAECILOMYCES LILACINUS AND MUCUNA SPP. SOIL AMENDMENTS. Udo1, I.A., M.I. Uguru2 and R.O. Ogbuji2. 1Department of Crop Science, University of Calabar, PMB 1115, Calabar, Nigeria; 2Department of Crop Science, University of Nigeria, Nsukka, Enugu State, Nigeria. A greenhouse trial was conducted to evaluate the efficacy of five arbuscular mycorrhizal fungi (AMF), five Mucuna spp. soil amendments and bioformulated Paecilomyces lilacinus singly and combined in the management of Meloidogyne incognita race 1 on tomato. The treatments involved the combination of five AMF (Glomus etunicatum, Glomus mosseae, Glomus clarum, Glomus deserticola and Gigaspora gigantea), five Mucuna spp (Mucuna pruriens utilis, Mucuna ghana,

250 Journal of Nematology, Volume 46, No. 2, June 2014 Mucuna cochichinensis, Mucuna jaspaeda and Mucuna pruriens IR2) and inoculation with bioformulated P. lilacinus. For the respective factors, pots without Mucuna amendment, not inoculated with P. lilacinus or arbuscular mycorrhizal fungi served as the controls. The tomato seedlings were inoculated with 5000 eggs of M. incognita per plant. The plants were grown to full maturity. The Mucuna spp. differed in their mineral contents. M. ghana and M. jaspaeda had the highest nitrogen (N) content (>4%) and the lowestt C : N ratio (7-9). The combination of the three control agents significantly (P