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Globodera ellingtonae was originally described from populations collected in the United States. In the original description, ribosomal DNA loci from Globodera sp. collected in Chile and Argentina were similar to G. ellingtonae, suggesting this nematode originated in this region of South America. In an effort to find additional populations of G. elllingtonae, collection trips were conducted in 2017 and 2020 in the Antofagasta and Arica y Parinacota Regions in Northern Chile, respectively. Globodera sp. were more prevalent in Antofagasta (17 samples collected, 53% positive for Globodera sp.) than in Arica y Parincota (16 samples collected, 13% positive for Globodera sp.). The genomes of single cysts (N ≥ 3) from four fields were sequenced. Additionally, the genomes of the G. ellingtonae population from Oregon and a Globodera sp. population originally collected in Antofagasta Region but maintained in culture in France were also sequenced. Based upon a HSP90 sequenced data mined from WSG data, all of the populations from the Antofagasta Region were G. ellingtonae and grouped in a monophyletic clade. A population collected from the Arica y Parincota Region was identified as G. rostochiensis based upon HSP90 data. Genome-wide SNP patterns of the G. ellingtonae populations showed strong clustering based on geographic location indicating that G. ellingtonae has high genetic diversity within Chile. A phylogenetic tree derived from 168,354 binary SNPs in the nuclear genome showed separate but distinct clustering of the Oregon population and the population from Antofagasta maintained in France. The Oregon G. ellingtonae population subtended the Chilean clades and placed on a long branch representing approximately twice the genetic variation observed among all Chilean G. ellingtonae populations. The possibility remains that G. ellingtonae from Chile may be sufficiently diverged to constitute a new species from G. ellingtonae originally described from a population collected in Oregon.

Our knowledge of the diversity of potato cyst nematodes in their native areas still remains patchy and should be improved. A previous study based on 42 Peruvian Globodera pallida populations revealed a clear south to north phylogeographic pattern, with five well‐supported clades and maximum diversity observed in the south of Peru. In order to investigate this phylogeographic pattern more closely, we genotyped a larger collection of Peruvian populations using both cathepsin L gene sequence data and a new set of 13 microsatellite loci. Using different genetic analyses (STRUCTURE, DAPC), we consistently obtained the same results that led to similar conclusions: the presence of a larger genetic diversity than previously known suggesting the presence of cryptic species in the south of Peru. These investigations also allowed us to clarify the geographic borders of the previously described G. pallida genetic clades and to update our knowledge of the genetic structure of this species in its native area, with the presence of additional clades. A distance‐based redundancy analysis (dbRDA) was also carried to understand whether there was a correlation between the population genetic differentiation and environmental conditions. This analysis showed that genetic distances observed between G. pallida populations are explained firstly by geographic distances, but also by climatic and soil conditions. This work could lead to a revision of the taxonomy that may have strong implications for risk assessment and management, especially on a quarantine species.

Plant-parasitic nematodes are a major, and in some cases a dominant, threat to crop production in all agricultural systems. The relative scarcity of classical resistance genes highlights a pressing need to identify new ways to develop nematode-resistant germplasm. Here, we sequence and assemble a high-quality genome of the model cyst nematode Heterodera schachtii to provide a platform for the first system-wide dual analysis of host and parasite gene expression over time, covering all major stages of the interaction. This novel approach enabled the analysis of the hologenome of the infection site, to identify metabolic pathways that were incomplete in the parasite but complemented by the host. Using a combination of bioinformatic, genetic, and biochemical approaches, we show that the highly atypical completion of vitamin B5 biosynthesis by the parasitic animal, putatively enabled by a horizontal gene transfer from a bacterium, is critically important for parasitism. Knockout of either the plant-encoded or the now nematode-encoded steps in the pathway blocks parasitism. Our experiments establish a reference for cyst nematodes, use this platform to further our fundamental understanding of the evolution of plant-parasitism by nematodes, and show that understanding congruent differential expression of metabolic pathways represents a new way to find nematode susceptibility genes, and thereby, targets for future genome editing-mediated generation of nematode-resistant crops.

International audience; Identification of plant parasitic nematode species is usually achieved following morphobiometric analysis, which requires a certain level of expertise and remains time consuming. Moreover, molecular and morphological discrimination of a number of emergent or cryptic species is sometimes difficult. Finding a way to achieve morphological characterisation quickly and accurately would greatly advance nematology science. Here, we developed a complete method in order to identify the two quarantine nematode species Globodera pallida and Globodera rostochiensis. First, we chose discriminative metrics on the stylet of nematodes that are able to be used by algorithms in order to build an automated process. Second, we used a custom computer vision algorithm (CCVA) and a convolutional neural network (CNN) to measure our metrics of interest. Third, we compared the CCVA and CNN predictions and their discriminative power to distinguish closely related species. Results show accurate identification of G. pallida and G. rostochiensis with the two methods, despite small-scale divergence (one to five µm depending on the metric used). However, the error rate is higher for Globodera mexicana, suggesting that the algorithms are too specific. Nonetheless, these methods represent a promising novel approach to automated morphological identification of nematodes and Globodera species in particular.

International audience; The cyst nematode Heterodera carotae, which parasitizes carrot roots, has been recorded in many countries in Europe (Italy, The Netherlands, Switzerland, France, Denmark, …), in South Africa and in North America (Canada, USA). To date, there is a lack of knowledge about the genetic structure of the populations of this economically important nematode. The aim of this work was to study the structuration of the genetic diversity of the carrot cyst nematode at the European scale. We have developed a set of thirteen polymorphic microsatellite markers and used it to genotype seventeen European populations of H. carotae coming from France, Switzerland, Italy, Denmark and one non-European population from Canada. As previously showed for other cyst nematode species, the H. carotae populations were characterised by a strong heterozygote deficit. A Bayesian clustering analysis revealed two distinct genetic clusters, with one group located in the north of Europe and a second one located in the south of Europe. Moreover, our results highlighted rather limited gene flow at small spatial scale and some events of long distance migration. This first investigation of the genetic diversity of H. carotae populations would be useful to develop sustainable control strategies.

International audience; Plant-parasitic nematodes are a costly burden of crop production. Ubiquitous in nature, phytoparasitic nematodes are associated with nearly every important agricultural crop and represent a significant constraint on global food security. Population genetics is a key discipline in plant nematology to understand aspects of the life strategies of these parasites, in particular their modes of reproduction, geographic origins, evolutionary histories, and dispersion abilities. Advances in high-throughput sequencing technologies have enabled a recent but active effort in genomic analyses of plant-parasitic nematodes. Such genomic approaches applied to multiple populations are providing new insights into the molecular and evolutionary processes that underpin the establishment of these nematodes and into a better understanding of the genetic and mechanistic basis of their pathogenicity and adaptation to their host plants. In this review, we attempt to update information about genome resources and genotyping techniques useful for nematologists who are thinking about initiating population genomics or genome sequencing projects. This review is intended also to foster the development of population genomics in plant-parasitic nematodes through highlighting recent publications that illustrate the potential for this approach to identify novel molecular markers or genes of interest and improve our knowledge of the genome variability, pathogenicity, and evolutionary potential of plant-parasitic nematodes.

International audience; Global trading of plant materials, in combination with agricultural practices, may facilitate the spreading of cyst nematodes to so far non-infected areas. Recently Potato Cyst Nematode (PCN) was recognized to be present in Indonesia and both diversity and distribution require further study. Assessment of PCN populations was done by collecting soil samples, determination of morphological characteristics in combination with ITS rDNA and COI mtDNA sequencing. Thirty-seven soil samples were collected from potato fields in the Indonesia archipelago. The results showed the presence ofGlobodera rostochiensisin 22 out of 37 sampling fields, namely North Sumatra (6 fields), Central Java (12 fields), East Java (3 fields), and -for the first time- in Sulawesi (North Sulawesi) (1 field). The highest observed density was found in Banjarnegara (Central Java), i.e., 872 cysts 100 ml soil(-1).Globodera pallidawas not recovered. Both ITS and COI characterisation of Indonesian PCN (G. rostochiensis) revealed the virtual absence of sequence variation as compared to most PCN from the rest of the world; the COI sequences were identical to the most common and mostly distributed haplotype around the world. Microsatellite genotyping indicated a higher genetic diversity for populations from East Java than for populations from North Sumatra, suggesting that cysts at the origin of populations in North Sumatra were coming from populations in East Java. These data on species identification, population density, genetic diversity, and distribution of potato cyst nematode over the Indonesian archipelago constitute the very basis for the design of environmentally-sound and effective PCN control strategies.

International audience; Management of plant pests is probably the most serious challenge in sustainable food production and the maintenance of food security. Due to the strict regulation of or ban on major categories of pesticide, the potato cyst nematode Globodera pallida has been managed by a combination of crop rotation and the potato resistance locus Grp1, a relatively narrow range resistance gene which was introgressed into a range of commercial potato cultivars in Europe. However, in 2014, G. pallida populations were described that can no longer be controlled by Grp1. Most likely similar highly virulent populations will also emerge in all major potato growing areas in North Western Europe where production practices are very similar. Except for laborious, costly and often moderately accurate pot experiments, there is currently no rapid and reliable method to identify virulent populations. This represents a strong limitation and prevents an accurate and durable management of infestations. The PalAdapt project funded by EFSA represents the first step of a European battle plan against the emergence of virulent G. pallida populations and aims at improving the methods and tools for a fast identification of virulence outbreaks. Four main research questions were investigated during the project: (i) Do resistance breaking populations correspond to novel introductions into Europe? (ii) Can miniaturized in vitro tests be used to get more rapidly an accurate identification of the virulence status?, (iii) Is cyst size a life history trait useful to estimate the virulence status of a population?, (iv) Can we identify polymorphism to design molecular tools for an accurate virulence monitoring? The EFSA partnering grants initiative was an accurate way to improve the EU risk assessment capacity through a knowledge exchange among partners having complementary resources and expertise.

International audience; Plant-parasite coevolution has generated much interest and studies to understand and manage diseases in agriculture. Such a reciprocal evolutionary process could lead to a pattern of local adaptation between plants and parasites. Based on the phylogeography of each partner, the present study tested the hypothesis of local adaptation between the potato cyst nematode Globodera pallida and wild potatoes in Peru. The measured fitness trait was the hatching of cysts which is induced by host root exudates. Using a cross-hatching assay between 13 populations of G. pallida and root exudates from 12 wild potatoes, our results did not show a strong pattern of local adaptation of the parasite but the sympatric combinations induced better hatching of cysts than allopatric combinations, and there was a negative relationship between the hatching percentage and the geographical distance between nematode populations and wild potatoes. Moreover, a strong effect of the geographic origin of root exudates was found, with root exudates from south of Peru inducing better hatching than root exudates from north of Peru. These results could be useful to develop new biocontrol products or potato cultivars to limit damages caused by G. pallida.

International audience; Evidence of strong gene flow among French populations of the carrot cyst nematode [i]Heterodera carotae[/i]

Considering the growing trade of seed potato, reliable diagnostic protocols are required for the detection of regulated nematode species. In this study, a specific and sensitive multiplex Taqman-based real-time PCR method was developed in order to detect and identifyGlobodera pallida,G. rostochiensisandHeterodera schachtii. The newly designed primers and probes enabled the detection of all the target populations tested and with no cross-reaction for closely related non-target species (55 populations tested). The limit of detection (LOD) was one juvenile forG. rostochiensisandG. pallidaand five juveniles forH. schachtii. For monitoring potato cyst nematodes, this analytical tool would extend the number of cyst investigated as five juveniles can be detected among 50 cysts in a sample. Furthermore, this multiplex assay detects DNA of the three targeted species in template DNA obtained directly from float material after nematode extraction from soil.

The effective size of a population is the size of an ideal population which would undergo genetic drift at the same rate as the real population. The balance between selection and genetic drift depends on the effective population size (Ne), rather than the real numbers of individuals in the population (N). The objectives of the present study were to estimateNein the potato cyst nematodeGlobodera pallidaand to explore the causes of a lowNe/Nratio in cyst nematodes using artificial populations. Using a temporal analysis of 24 independent populations, the medianNewas 58 individuals (minNe= 25 and maxNe= 228).Neis commonly lower thanNbut in the case of cyst nematodes, theNe/Nratio was extremely low. Using artificial populations showed that this low ratio did not result from migration, selection and overlapping generations, but could be explain by the fact thatG. pallidapopulations deviate in structure from the assumptions of the ideal population by having unequal sex ratios, high levels of inbreeding and a high variance in family sizes. The consequences of a lowNe, resulting in a strong intensity of genetic drift, could be important for their control becauseG. pallidapopulations will have a low capacity to adapt to changing environments.

Background In hyperspecialized parasites, the ability to grow on a particular host relies on specific virulence factors called effectors. These excreted proteins are involved in the molecular mechanisms of parasitism and distinguish virulent pathogens from non-virulent related species. The potato cyst nematodes (PCN) Globodera rostochiensis and G. pallida are major plant-parasitic nematodes developing on numerous solanaceous species including potato. Their close relatives, G. tabacum and G. mexicana are stimulated by potato root diffusate but unable to establish a feeding site on this plant host. Results RNA sequencing was used to characterize transcriptomic differences among these four Globodera species and to identify genes associated with host specificity. We identified seven transcripts that were unique to PCN species, including a protein involved in ubiquitination. We also found 545 genes that were differentially expressed between PCN and non-PCN species, including 78 genes coding for effector proteins, which represent more than a 6-fold enrichment compared to the whole transcriptome. Gene polymorphism analysis identified 359 homozygous non-synonymous variants showing a strong evidence for selection in PCN species. Conclusions Overall, we demonstrated that the determinant of host specificity resides in the regulation of essential effector gene expression that could be under the control of a single or of very few regulatory genes. Such genes are therefore promising targets for the development of novel and more sustainable resistances against potato cyst nematodes. Electronic supplementary material The online version of this article (10.1186/s12864-019-5853-4) contains supplementary material, which is available to authorized users.

Studying phenotypic and genomic modifications associated with pathogen adaptation to resistance is a crucial step to better understand and anticipate resistance breakdown. This short review summarizes recent results obtained using experimentally evolved populations of the potato cyst nematode Globodera pallida. In a first step, the variability of resistance durability was explored in four different potato genotypes carrying the resistance quantitative trait loci (QTL) GpaV(vrn) originating from Solanum vernei but differing by their genetic background. The consequences of the adaptation to resistance in terms of local adaptation, cross-virulence and virulence cost were then investigated. Finally, a genome scan approach was performed in order to identify the genomic regions involved in this adaptation. Results showed that nematode populations were able to adapt to the QTL GpaV(vrn), and that the plant genetic background has a strong impact on resistance durability. A trade-off between the adaptations to different resistant potato genotypes was detected, and we also showed that adaptation to the resistance QTL GpaV(vrn) from S. vernei did not allow adaptation to the colinear locus from S. sparsipilum (GpaV(spl)). Unexpectedly, the adaptation to resistance led to an increase of virulent individual's fitness on a susceptible host. Moreover, the genome scan approach allowed the highlighting of candidate genomic regions involved in adaptation to host plant resistance. This review shows that experimental evolution is an interesting tool to anticipate the adaptation of pathogen populations and could be very useful for identifying durable strategies for resistance deployment.

Among plant-parasitic nematodes (PPNs), the species Meloidogyne chitwoodi constitutes an important agricultural issue and is listed as a quarantine species in the EU. This study investigated whether PPN communities can affect the establishment and expansion of M. chitwoodi during a simulated introduction. An original experimental design was developed on potato in the glasshouse involving four PPN communities derived from a single natural one that was initially grown on different host plants. Meloidogyne chitwoodi was inoculated at two different densities (50 or 1000 second-stage juveniles (J2) per pot). After 4 months, M. chitwoodi was more abundant in the community showing a low overall abundance of PPNs, decreased in the other PPN communities and failed to establish in five pots out of 80. Results showed that establishment - even starting from a very low inoculum - can occur in all communities even if indigenous PPNs in the community affect the expansion through interspecific competition. The reverse interaction was also observed as PPN taxa were generally less abundant in the 1000 M. chitwoodi J2 inoculation treatments. The proportion of M. chitwoodi males produced was higher in the community showing low overall PPN abundance, suggesting a strong sensitivity of M. chitwoodi to intraspecific competition. The presented results suggest that a low abundance of indigenous PPNs presents a higher risk of expansion of M. chitwoodi even if the introduced inoculum is low.

International audience; Agricultural practices shaping plant-parasitic nematode (PPN) assembly are still unclear, and this limits our understanding of the impact of anthropic disturbances on the resilience of PPN communities and the emergence of agronomic problems. Here the abundance and diversity of PPN in France's oilseed rape production area was determined by sampling 72 fields over two consecutive years. We identified and counted PPN taxa and collected anthropic and environmental variables for the past 5 years. PPN were assigned to seven genera and one family including PPN that have not been identified to genus level. Using multiple correspondence analyses, we selected the main variables and tested their effect on the abundance of each taxon with mixed generalised linear models. We emphasise that at the landscape scale investigated, crop rotations were no longer a major factor impacting the PPN communities. However, we observed that tillage and pesticides had a significant impact on several taxa.

SummaryThe effective size of a population is the size of an ideal population which would drift at the same rate as the real population. The balance between selection and genetic drift depends on the population size expressed as the genetically effective population size (Ne), rather than the real numbers of individuals in the population (N).The objectives of the present study were to estimate Nein the potato cyst nematodeGlobodera pallidausing artificial populations and to explore the link between Neand the durability of plant resistances.Using a temporal method on 24 independent pairs of initial and final populations, the median Newas 58 individuals.Neis commonly lower than N but in our case the Ne/N ratio was extremely low becauseG. pallidapopulations deviate in structure from the assumptions of the ideal population by having unequal sex-ratios, high levels of inbreeding and a high variance in family sizes. The consequences of a low Necould be important for the control of phytoparasitic nematodes becauseG. pallidapopulations will have a low capacity to adapt to changing environments unless selection intensity is very strong, which could be greatly beneficial for long-term use of plant resistances.

Routine postoperative radiographs are commonly obtained at the initial postoperative visit. The purpose of this study was to demonstrate the clinical utility of routine postoperative radiographs and quantify the radiation exposure and cost to the health care system. All orthopaedic surgeries performed during 2007 at a level I trauma center were retrospectively analyzed. Surgical procedures that were likely to require follow-up radiographs were included. Analysis demonstrated 878 procedures with 471 procedures (54%) receiving initial postoperative radiographs. Routine radiographs were performed in 455 (96.6%) procedures with 4/455 (0.879%) resulting in a change in management. Nonroutine radiographs were performed in 16 (3.40%) procedures with 2/16 (12.5%) triggering a change in management. Subjects receiving radiographs at the initial postoperative visit obtained a mean of 2.54 radiographs per procedure with a mean exposure of 0.199 mSv with a median cost of $29.98 per radiographic series in 2013 Medicare reimbursement dollars. (Journal of Surgical Orthopaedic Advances.

International audience; Improving resistance durability involves to be able to predict the adaptation speed of pathogen populations. Identifying the genetic bases of pathogen adaptation to plant resistances is a useful step to better understand and anticipate this phenomenon. Globodera pallida is a major pest of potato crop for which a resistance QTL, GpaV(vm), has been identified in Solanum vernei. However, its durability is threatened as G. pallida populations are able to adapt to the resistance in few generations. The aim of this study was to investigate the genomic regions involved in the resistance breakdown by coupling experimental evolution and high-density genome scan. We performed a whole-genome resequencing of pools of individuals (Pool-Seq) belonging to G. pallida lineages derived from two independent populations having experimentally evolved on susceptible and resistant potato cultivars. About 1.6 million SNPs were used to perform the genome scan using a recent model testing for adaptive differentiation and association to population-specific covariables. We identified 275 outliers and 31 of them, which also showed a significant reduction in diversity in adapted lineages, were investigated for their genic environment. Some candidate genomic regions contained genes putatively encoding effectors and were enriched in SPRYSECs, known in cyst nematodes to be involved in pathogenicity and in (a) virulence. Validated candidate SNPs will provide a useful molecular tool to follow frequencies of virulence alleles in natural G. pallida populations and define efficient strategies of use of potato resistances maximizing their durability.

In the current agronomical context of pesticide use reduction, deciphering the genetic bases of pathogen adaptation to plant resistances is of major importance to improve their durability. Knowledge of the virulence gene frequencies in pathogens populations could indeed allow the prediction of their durability before deployment. Globodera pallida is a major pest of potato crop for which a promising resistance QTL, GpaVvrn, has been identified in Solanum vernei. An experimental evolution study, in which G. pallida lineages evolved on resistant or susceptible potato genotypes for up to eight generations, previously showed that G. pallida was able to rapidly overcome GpaVvrn resistance. However it was not known if enough genetic mixing occurred in these lineages to be able to detect islands of differentiation in a genome scan approach. Here, we investigated this question using 53 polymorphic microsatellite markers distributed along the genome and three different tests based on genetic differentiation and heterozygosity. We identified eight outlier loci, indicative of genomic regions putatively involved in host adaptation. Several loci were identified by multiple detection methods and/or in two independent adapted lineages. We also showed that some identified candidate genomic regions seem to be also involved in the overcoming of nematode resistance in a genotype harbouring the same resistance QTL in a different genetic background. These results validate the feasibility of a genome scan approach on a biological material coming from short experimental evolution, and encourage to perform a high coverage genome scan using whole genome resequencing. This article is protected by copyright. All rights reserved.

Can social media be an effective way for merchants to create new business? We believe that more and more businesses are using Social Media Promotions (SMPs) as an inexpensive way to market their business and/or new items to new, old and existing customers. We examined the effects of SMPs on businesses by surveying 111 customers and over 30 businesses. We found that customers mainly purchased a promotion to either try out a new place or enjoy the value of the deal. On the business side, we found that businesses have had a good experience with offering SMPs. Businesses can maintain profitability while offering a promotion via a social media site. The customer experience at the business is mostly the deciding factor in determining if they will return without the coupon. This study concludes that businesses were able to be successful offering discounts via a promotion on social media sites, such as Groupon and LivingSocial, as long as they follow some key steps in their strategy and prepare themselves for the positive and negative effects that they might experience. Customers in today’s economy are always looking for deals and an easy way for businesses to increase their profits and bring in new customers is to offer deals on social media sites.

In the current agronomical context of reduced use of pesticides, deciphering the genetic bases of pathogen adaptation to plant resistances is of major importance as it could help us to improve durability of these resistances. Globodera pallida is a major pest of potato and for which the promising resistance factor, QTL GpaVvrn, has been identified in Solanum vernei and introduced into several new resistant cultivars in Europe.

To understand the mechanisms leading to the breakdown of plant resistance by potato cyst nematodes (PCN; Globodera rostochiensis and G. pallida), an understanding of the genetic structure and variation in populations and the processes that govern their evolution is required.

International audience; Tobacco cyst nematode is a sexually diploid species composed of three subspecies, Globodera tabacum subsp. tabacum (Gtt), G. tabacum subsp. virginiae (Gtv) and G. tabacum subsp. solanacearum (Gts). In Europe, only Gtt have been reported to date. Here, we report from a plant in a French tobacco field, the identification of a mixture between cysts of the subspecies Gtt and cysts genetically different. The objective of the present study was to determine the subspecies status of those particular cysts. To this aim, we performed CAPS analyses on two genes and a principal component analysis on microsatellite data. The results obtained with both CAPS and microsatellite markers demonstrate for the first time the occurrence of G. tabacum subsp. virginiae in France.

International audience; In this article, we describe the analysis of over 9000 expressed sequence tags (ESTs) from cDNA libraries obtained from various life cycle stages of Globodera pallida. We have identified over 50 G. pallida effectors from this dataset using bioinformatics analysis, by screening clones in order to identify secreted proteins up-regulated after the onset of parasitism and using in situ hybridization to confirm the expression in pharyngeal gland cells. A substantial gene family encoding G. pallida SPRYSEC proteins has been identified. The expression of these genes is restricted to the dorsal pharyngeal gland cell. Different members of the SPRYSEC family of proteins from G. pallida show different subcellular localization patterns in plants, with some localized to the cytoplasm and others to the nucleus and nucleolus. Differences in subcellular localization may reflect diverse functional roles for each individual protein or, more likely, variety in the compartmentalization of plant proteins targeted by the nematode. Our data are therefore consistent with the suggestion that the SPRYSEC proteins suppress host defences, as suggested previously, and that they achieve this through interaction with a range of host targets.

Native to South America, the potato cyst nematode Globodera pallida is one of the principal pests of Andean potato crops and is also an important global pest following its introduction to Europe, Africa, North America, Asia and Oceania. Building on earlier work showing a clear south to north phylogeographic pattern in Peruvian populations, we have been able to identify the origin of Western European populations with high accuracy. They are all derived from a single restricted area in the extreme south of Peru, located between the north shore of the Lake Titicaca and Cusco. Only four cytochrome b haplotypes are found in Western Europe, one of them being also found in some populations of this area of southern Peru. The allelic richness at seven microsatellite loci observed in the Western European populations, although only one-third of that observed in this part of southern Peru, is comparable to the allelic richness observed in the northern region of Peru. This result could be explained by the fact that most of the genetic variability observed at the scale of a field or even of a region is already observed at the scale of a single plant within a field. Thus, even introduction via a single infected potato plant could result in the relatively high genetic variability observed in Western Europe. This finding has important consequences for the control of this pest and the development of quarantine measures.

Plant-parasitic nematodes are major agricultural pests worldwide and novel approaches to control them are sorely needed. We report the draft genome sequence of the root-knot nematode Meloidogyne incognita, a biotrophic parasite of many crops, including tomato, cotton and coffee. Most of the assembled sequence of this asexually reproducing nematode, totaling 86 Mb, exists in pairs of homologous but divergent segments. This suggests that ancient allelic regions in M. incognita are evolving toward effective haploidy, permitting new mechanisms of adaptation. The number and diversity of plant cell wall-degrading enzymes in M. incognita is unprecedented in any animal for which a genome sequence is available, and may derive from multiple horizontal gene transfers from bacterial sources. Our results provide insights into the adaptations required by metazoans to successfully parasitize immunocompetent plants, and open the way for discovering new antiparasitic strategies

Using a complementary (c)DNA-amplified fragment length polymorphism (AFLP) approach, we investigated differential gene expression linked to resistance mechanisms during the incompatible potato – Globodera pallida interaction. Expression was compared between a resistant and a susceptible potato clone, inoculated or not inoculated with G. pallida. These clones were issued from a cross between the resistant Solanum sparsipilum spl329.18 accession and the susceptible dihaploid S. tuberosum Caspar H3, and carried, respectively, resistant and susceptible alleles at the resistance quantitative trait loci (QTLs). Analysis was done on root fragments picked up at 4 time points, during a period of 6 days after infection, from penetration of the nematode in the root to degradation of the feeding site in resistant plants. A total of 2560 transcript-derived fragments (TDFs) were analyzed, resulting in the detection of 46 TDFs that were up- or downregulated. The number of TDFs that were up- or downregulated increased with time after inoculation. The majority of TDFs were upregulated at only 1 or 2 time points in response to infection. After isolation and sequencing of the TDFs of interest, a subset of 36 sequences were identified, among which 22 matched plant sequences and 2 matched nematode sequences. Some of the TDFs that matched plant genes showed clear homologies to genes involved in cell-cycle regulation, transcription regulation, resistance downstream signalling pathways, and defense mechanisms. Other sequences with homologies to plant genes of unknown function or without any significant similarity to known proteins were also found. Although not exhaustive, these results represent the most extensive list of genes with altered RNA levels after the incompatible G. pallida–potato interaction that has been published to date. The function of these genes could provide insight into resistance or plant defense mechanisms during incompatible potato-cyst nematode interactions.

International audience; Deviations of genotypic frequencies from Hardy-Weinberg equilibrium (HWE) expectations could reveal important aspects of the biology of populations. Deviations from HWE due to heterozygote deficits have been recorded for three plant-parasitic nematode species. However, it has never been determined whether the observed deficits were due (i) to the presence of null alleles, (ii) to a high level of consanguinity and/or (iii) to a Wahlund effect. The aim of the present work was, while taking into the possible confounding effect of null alleles, to disentangle consanguinity and Wahlund effect in natural populations of those three economically important cyst nematodes using microsatellite markers: Globodera pallida, G. tabacum and Heterodera schachtii, pests of potato, tobacco and sugar beet, respectively. The results show a consistent pattern of heterozygote deficiency in the three nematode species sampled at the spatial scale of the host plant. We demonstrate that the prevalence of null alleles is weak and that heterozygote deficits do not have a single origin. Our results suggested that it is restricted dispersal that leads to heterozygote deficits through both consanguinity and substructure, which effects can be linked to soil movement, cyst density, and the number of generations per year. We discuss potential implications for the durability of plant resistances that are used to protect crops against parasites in which mating between relatives occur. While consanguineous mating leads to homozygosity at all loci, including loci governing avirulence/virulence, which favours the expression of virulence when recessive, the Wahlund effect is expected to have no particular effect on the adaptation of nematodes to resistances.

A homologue of a Ran binding protein to microtubules ( ranbpm ) gene, initially known as IC5, was identified and characterised in second-stage juveniles (J2s) of the potato cyst nematode Globodera pallida. The full-length cDNA (937 bp) was obtained by 5′ and 3′ rapid amplification of cDNA ends (RACE) and specific primers were designed to amplify the genomic sequences of 2396 bp containing six introns. The ORF (798 bp) encodes a putative 265 amino acid sequence with a predicted SPRY domain and a signal peptide of 23 amino acids on the N-terminal part of the protein. In situ hybridisation experiments showed that the transcript is located in the dorsal gland of the J2s, suggesting that the encoded protein has an extracellular function and can be involved in the late stages of parasitism such as feeding site establishment. This gene was specifically over-expressed in the juveniles before and during parasitism, but not in adult developmental stages. As this gene is presumed to be involved in plant–nematode interaction, particularly in the development and maintenance of the feeding structure that allows the nematode to achieve parasitic development, homologous genes were sought in other cyst nematode species. One of them was cloned and sequenced in the closely related species Globodera ‘ mexicana ’.

Summary Globodera pallida and G.'mexicana' are closely related nematode species that can mate and form viable hybrids on tomato but usually develop on different Solanaceous plants. Identification of nematode genes involved in parasitism is important for elucidation of disease resistance mechanisms in plants. In this study, we have used suppression subtractive hybridization (SSH) to investigate differences between the transcriptomes of G. pallida and G.'mexicana' J2s. This provides a basis for further studies characterizing pathogenicity factors in these nematodes. None of the cDNA fragments isolated in the SSH experiments appeared to be completely absent from the other transcriptome. Differences in expression levels of some of the isolated cDNAs between the two species were detected. Sequence analysis revealed that nearly 85% of the cloned sequences are nematode specific and a high proportion were pioneer genes for which no putative homologues were present in the databases. However, homologues of a cellulase and a putative pathogenicity factor previously described from G. rostochiensis were isolated. The putative roles of these sequences in parasitism are discussed.

Session 6: GenomicsSession 6: Genomics; absent

International audience; 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 of significant yield reductions. Nine microsatellites markers were used to explore the dispersal and genetic structure of 18 French G. tabacum populations. All the populations sampled belong to the "tabacum" subspecies and low level of gene flow was observed among G. tabacum populations in France. Bayesian genetic assignments revealed two distinct genetic groups that matched with the geographic limits of two agricultural cooperative societies. An important part of the genetic variability was observed between these agricultural cooperative societies and also within populations. Those results highlight the impact of the human organization of agricultural practices on the genetic structure of G. tabacum populations and complement previous results obtained on other cyst nematodes, showing the major contribution of human activities and soil transports during harvest in the passive dispersion of these organisms. (c) 2014 Elsevier B.V. All rights reserved.

Peruvian potato cyst nematode populations were analysed to assess both their inter- and intraspecific similarities. ITS--RFLP and two satellite DNA sequences were used as taxonomic tools. Both techniques have confirmed that the Peruvian populations have as their closest relatives the European Globodera pallida, despite the detection of clear differences that prevents us from assigning these South American populations unambiguously to any Globodera species. A more precise study of the variability of these Peruvian populations was investigated and they were compared with the imported European populations using protein (2-DGE) and DNA (RAPD) datasets. The clear distinction between the Peruvian and the European populations was confirmed and, inside each group, no correlation was found between the pathotype classification and the observed clustering of the populations. Surprisingly, while RAPDs revealed a higher variability in the Peruvian group than in the European one, some characteristic proteins were found by 2-DGE in some European populations, whereas it was impossible to find any in the Peruvian populations. It is concluded that the primary founders of the European populations may have an origin other than that of the Peruvian populations involved in this study.

For the first time, mariner elements were found in insect parasitic nematodes. Full-length elements were isolated from the rhabditid Heterorhabditis bacteriophora. They were 1279 bp long, flanked by two 30-bp inverted repeats, and were able to encode a putative 358-amino acid transposase. These elements were present in about 30 copies in the H. bacteriophora genome, but their distribution among closely related Heterorhabditis and Steinernema genera was patchy. DNA and encoded peptide sequences of H. bacteriophora mariners showed greater similarity to the mariner of the coleopteran Carpelimus sp. than to the mariners of the rhabditid Caenorhabditis elegans. The possibility of horizontal transfer was investigated by examination of a host for Heterorhabditis nematodes, a beetle of the Phyllophaga sp. Mariner elements were found in this insect, but they were not very similar to the H. bacteriophora elements. Finally, the H. bacteriophora mariners formed a group with those of invertebrates, suggesting vertical transmission from a common ancestor.

International audience; The golden cyst nematode (Globodera rostochiensis), native to South America, has been introduced in many parts of the world, including Europe and North America. Recently, it was found for the first time in the province of Quebec, Canada in the locality of St. Amable near Montreal. To date, very few studies have examined the population genetics of this pest. Consequently, there is a lack of knowledge about the genetic structure and evolution of this nematode. In this study, twelve new microsatellite markers were developed in order to explore these questions. These markers were used to genotype fifteen populations originating from different regions of the world, including five from Canada. Within populations, the highest genetic diversity was consistently observed in the populations from Bolivia, the postulated region of origin of the golden nematode, and the lowest in populations from British Columbia (Canada) and New York (USA). The two Quebec populations were very similar to each other and to the population found in Newfoundland, but surprisingly, they were significantly different from three other North American populations including those from New York and British Columbia. Based on our results, we conclude that the golden cyst nematode has been introduced in North America at least twice from distinct regions of the world. Crown Copyright (C) 2013 Published by Elsevier Inc. All rights reserved.

Background The Ran GTPase Activating Protein 2 (RanGAP2) was first described as a regulator of mitosis and nucleocytoplasmic trafficking. It was then found to interact with the Coiled-Coil domain of the Rx and GPA2 resistance proteins, which confer resistance to Potato Virus X (PVX) and potato cyst nematode Globodera pallida, respectively. RanGAP2 is thought to mediate recognition of the avirulence protein GP-RBP-1 by GPA2. However, the Gpa2-induced hypersensitive response appears to be relatively weak and Gpa2 is limited in terms of spectrum of efficiency as it is effective against only two nematode populations. While functional and evolutionary analyses of Gp-Rbp-1 and Gpa2 identified key residues in both the resistance and avirulence proteins that are involved in recognition determination, whether variation in RanGAP2 also plays a role in pathogen recognition has not been investigated. Results We amplified a total of 147 RanGAP2 sequences from 55 accessions belonging to 18 different di-and tetraploid Solanum species from the section Petota. Among the newly identified sequences, 133 haplotypes were obtained and 19.1% of the nucleotide sites were found to be polymorphic. The observed intra-specific nucleotide diversity ranges from 0.1 to 1.3%. Analysis of the selection pressures acting on RanGAP2 suggests that this gene evolved mainly under purifying selection. Nonetheless, we identified polymorphic positions in the protein sequence at the intra-specific level, which could modulate the activity of RanGAP2. Two polymorphic sites and a three amino-acid deletion in RanGAP2 were found to affect the timing and intensity of the Gpa2-induced hypersensitive response to avirulent GP-RBP-1 variants even though they did not confer any gain of recognition of virulent GP-RBP-1 variants. Conclusions Our results highlight how a resistance gene co-factor can manage in terms of evolution both an established role as a cell housekeeping gene and an implication in plant parasite interactions. StRanGAP2 gene appears to evolve under purifying selection. Its variability does not seem to influence the specificity of GPA2 recognition but is able to modulate this activity by enhancing the defence response. It seems therefore that the interaction with the plant resistance protein GPA2 (and/or Rx) rather than with the nematode effector was the major force in the evolution of the RanGAP2 locus in potato. From a mechanistic point of view these results are in accordance with a physical interaction of RanGAP2 with GPA2 and suggest that RBP-1 would rather bind the RanGAP2-GPA2 complex than the RanGAP2 protein alone.

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