Guy Bélair

Genetic diversity of the golden potato cyst nematode Globodera rostochiensis and determination of the origin of populations in Quebec, Canada.

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.

Analysis of Putative Apoplastic Effectors from the Nematode, Globodera rostochiensis, and Identification of an Expansin-Like Protein That Can Induce and Suppress Host Defenses

The potato cyst nematode, Globodera rostochiensis, is an important pest of potato. Like other pathogens, plant parasitic nematodes are presumed to employ effector proteins, secreted into the apoplast as well as the host cytoplasm, to alter plant cellular functions and successfully infect their hosts. We have generated a library of ORFs encoding putative G. rostochiensis putative apoplastic effectors in vectors for expression in planta. These clones were assessed for morphological and developmental effects on plants as well as their ability to induce or suppress plant defenses. Several CLAVATA3/ESR-like proteins induced developmental phenotypes, whereas predicted cell wall-modifying proteins induced necrosis and chlorosis, consistent with roles in cell fate alteration and tissue invasion, respectively. When directed to the apoplast with a signal peptide, two effectors, an ubiquitin extension protein (GrUBCEP12) and an expansin-like protein (GrEXPB2), suppressed defense responses including NB-LRR signaling induced in the cytoplasm. GrEXPB2 also elicited defense response in species- and sequence-specific manner. Our results are consistent with the scenario whereby potato cyst nematodes secrete effectors that modulate host cell fate and metabolism as well as modifying host cell walls. Furthermore, we show a novel role for an apoplastic expansin-like protein in suppressing intra-cellular defense responses.

Current and potential use of pathogens in the management of turfgrass insects as affected by new pesticide regulations in North America

New legislation by governments and municipal authorities for the application of synthetic pesticides, including insecticides on turfgrass is currently changing the management of insect pests. Since the late 1990s, societal concerns over perceived health risks and environmental hazards of pesticides have led to tighter restrictions on insecticide use. In Canada, two provinces have banned the use of pesticides for cosmetic purposes and more than 152 municipalities have adopted some form of bylaw restricting or banning the use of landscape pesticides. Turfgrass managers are facing increasing constraints regarding the use of synthetic pesticides and only a few effective biological control agents are currently available on the markets to control turfgrass insects. Here we review the different pathogens that show potential for control and commercial use against turfgrass insect pests in North America. Even though numerous studies have shown the virulence of various pathogens against turfgrass insect pests, few of these pathogens are widely used. Limiting factors for their adoption include a narrow host range and, compared to synthetic insecticides, lower consistency, higher costs, and/or shorter shelf-life. However, new pesticide regulations, particularly in Canada, should lead to better opportunities for the biological control industry in turfgrass markets.

Pearl Millet for the Management of Pratylenchus penetrans in Flue-Cured Tobacco in Quebec

Two 1-year rotation experiments were conducted from 1998 to 2000 to assess the impact of forage and grain pearl millet (Pennisetum glaucum) on Pratylenchus penetrans populations in a tobacco field (Nicotiana tabacum cv. Delgold) in Quebec. Performance of these crops was compared with rye, the standard rotation crop. Rye plots were doubled to include a fumigated standard (application of metham sodium at 67.4 liters/ha on a band). Forage pearl millet increased tobacco yields by an average of 103% compared with nonfumigated rye. In 2000, leaf dry weights of tobacco following grain pearl millet CGPM H5 and CGPM H6 was increased by 70 and 73%, respectively, when compared with nonfumigated rye and were not significantly different from the fumigated plots following rye. Forage and grain pearl millet should be considered as a viable alternative to fumigation for controlling P. penetrans infestation in flue-cured tobacco production in Quebec.

A new method for studying population genetics of cyst nematodes based on Pool-Seq and genomewide allele frequency analysis

Cyst nematodes are important agricultural pests responsible for billions of dollars of losses each year. Plant resistance is the most effective management tool, but it requires a close monitoring of population genetics. Current technologies for pathotyping and genotyping cyst nematodes are time‐consuming, expensive and imprecise. In this study, we capitalized on the reproduction mode of cyst nematodes to develop a simple population genetic analysis pipeline based on genotyping‐by‐sequencing and Pool‐Seq. This method yielded thousands of SNPs and allowed us to study the relationships between populations of different origins or pathotypes. Validation of the method on well‐characterized populations also demonstrated that it was a powerful and accurate tool for population genetics. The genomewide allele frequencies of 23 populations of golden nematode, from nine countries and representing the five known pathotypes, were compared. A clear separation of the pathotypes and fine genetic relationships between and among global populations were obtained using this method. In addition to being powerful, this tool has proven to be very time‐ and cost‐efficient and could be applied to other cyst nematode species.

Analysis of Globodera rostochiensis effectors reveals conserved functions of SPRYSEC proteins in suppressing and eliciting plant immune responses.

Potato cyst nematodes (PCNs), including Globodera rostochiensis (Woll.), are important pests of potato. Plant parasitic nematodes produce multiple effector proteins, secreted from their stylets, to successfully infect their hosts. These include proteins delivered to the apoplast and to the host cytoplasm. A number of effectors from G. rostochiensis predicted to be delivered to the host cytoplasm have been identified, including several belonging to the secreted SPRY domain (SPRYSEC) family. SPRYSEC proteins are unique to members of the genus Globodera and have been implicated in both the induction and the repression of host defense responses. We have tested the properties of six different G. rostochiensis SPRYSEC proteins by expressing them in Nicotiana benthamiana and N. tabacum. We have found that all SPRYSEC proteins tested are able to suppress defense responses induced by NB-LRR proteins as well as cell death induced by elicitors, suggesting that defense repression is a common characteristic of members of this effector protein family. At the same time, GrSPRYSEC-15 elicited a defense responses in N. tabacum, which was found to be resistant to a virus expressing GrSPRYSEC-15. These results suggest that SPRYSEC proteins may possess characteristics that allow them to be recognized by the plant immune system.

Toxicity of spinosad and Beauveria bassiana to the black cutworm, and the additivity of subletal doses

Abstract The main objective of this study was to assess the susceptibility of the black cutworm (Agrotis ipsilon) to the biopesticide spinosad and to a commercial formulation (GHA strain) of the entomopathogenic fungus, Beauveria bassiana. Secondly, we quantified the effects of sublethal doses of spinosad on a number of A. ipsilon fitness parameters, and interactions resulting from simultaneous applications of sub-lethal doses of spinosad and B. bassiana. Under laboratory conditions, A. ipsilon third instar larvae were highly susceptible to spinosad, with an estimated LC50 of 50 ppm. The entomopathogenic fungus, B. bassiana had a lower efficacy with an estimated LC50 of 7×107 spores mL−1. Topical applications of 5, 7.5 and 10 ppm of spinosad on third instar larvae reduced larval size and increased time to pupation and to emergence. However, pupal and adult weights were not significantly different between treated and control individuals. Additivity was observed from most spinosad–B. bassiana combinations tested, thus indicating compatibility between products. We concluded that spinosad is a promising tool for controlling black cutworm larvae alone or in combination with other products.

Virulence of entomopathogenic nematodes (Rhabditida: Steinernematidae, Heterorhabditidae) against Tipula paludosa (Diptera: Tipulidae), a turfgrass pest on golf courses

Abstract The European crane fly (ECF), Tipula paludosa Meigen feeds on leaves, crowns, and roots of cool-season turfgrasses causing damage to residential lawns and golf courses. A laboratory study was conducted to determine the susceptibility of ECF larvae to four commercial entomopathogenic nematode (EPN) species (Heterorhabditis marelatus, H. megidis, Steinernema carpocapsae and S. feltiae). The virulence of four S. feltiae isolates recovered from golf courses in Quebec and Ontario were also compared to a commercial strain. LC50 values of EPN against late instar ECF larvae were 152, 562, 763, and 3584 for S. feltiae, H. megidis, H. marelatus and S. carpocapsae, respectively. When non-feeding (without grass seedling), ECF larvae mortalities decreased for all nematode species and concentrations tested. At 25°C, LC50 values for the two most virulent indigenous S. feltiae were 129 and 187 nematodes/larva, not different from the commercial strain. At 5°C, the commercial S. feltiae was more effective than both BIC14A and RE6A isolates against ECF larvae. However, at 15°C, BIC14A was the most virulent at the low concentration of 200 IJs/larva.

Natural occurrence of entomopathogenic nematodes (Rhabditida: Steinernematidae) on golf courses in eastern Canada

A survey of entomopathogenic nematodes from 38 golf courses was conducted in different climatic areas of Ontario and Quebec provinces, Canada, during autumn 2002 and 2003. Soil samples were collected on golf course greens, fairways and roughs. Entomopathogenic nematodes were isolated from soil using wax moth (Galleria mellonella) and carrot weevil (Listronotus oregonensis) larvae as bait. Of the 436 samples (218 sites), 11 contained entomopathogenic nematodes for this study. All entomopathogenic nematode-positive samples were isolated from roughs. Isolates were characterised using DNA satellite probes and 28S rDNA sequence data. Both methods confirmed the identities of nematode isolates as Steinernema carpocapsae, S. feltiae and S. kraussei.

Evaluation of cultural methods for the management of the golden nematode (Globodera rostochiensis) in Quebec, Canada

Abstract Potato cyst nematodes are a major threat to potato production worldwide. In 2006, the isolation of the golden nematode, Globodera rostochiensis, in Quebec, Canada, led to the establishment of a quarantine area and the initiation of a research programme for the sustainable management of this regulated pest. In this study, the field efficacy of crop rotations and soil amendments was assessed in microplots for their potential to reduce populations of G. rostochiensis in the quarantine area. Crop rotation with a resistant potato cultivar was very effective in decreasing the population density of G. rostochiensis in soil. A single year with a cultivar carrying the H1 resistance gene reduced nematode populations by 62–95%. After 3 consecutive years of cropping to resistant potato, the number of viable eggs was zero in several microplots, and the overall population reduction was around 95%. Natural population decline with the cultivation of a non-host crop such as corn was around 30% per year. Trap cropping also showed reduced G. rostochiensis populations with results similar to the resistant cultivar. Sticky nightshade could not be established under Quebec’s climatic conditions and therefore was not a viable alternative for managing golden nematode in Quebec. Soil amendment with high rates of urea provided some reduction in populations but was not an economically viable option for managing potato cyst nematodes. Chicken manure and pig slurry did not provide a significant reduction of G. rostochiensis under Quebec field conditions.