Spatial Genetic Structure and Pathogenic Race Composition at the Field Scale in the Sunflower Downy Mildew Pathogen, Plasmopara halstedii

Yield losses in sunflower crops caused by Plasmopara halstedii can be up to 100%, depending on the cultivar susceptibility, environmental conditions, and virulence of the pathogen population. The aim of this study was to investigate the genetic and phenotypic structure of a sunflower downy mildew agent at the field scale. The genetic diversity of 250 P. halstedii isolates collected from one field in southern France was assessed using single-nucleotide polymorphisms (SNPs) and single sequence repeats (SSR). A total of 109 multilocus genotypes (MLG) were identified among the 250 isolates collected in the field. Four genotypes were repeated more than 20 times and spatially spread over the field. Estimates of genetic relationships among P. halstedii isolates using principal component analysis and a Bayesian clustering approach demonstrated that the isolates are grouped into two main genetic clusters. A high level of genetic differentiation among clusters was detected (FST = 0.35), indicating overall limited exchange between them, but our results also suggest that recombination between individuals of these groups is not rare. Genetic clusters were highly related to pathotypes, as previously described for this pathogen species. Eight different races were identified (100, 300, 304, 307, 703, 704, 707, and 714), with race 304 being predominant and present at most of the sites. The co-existence of multiple races at the field level is a new finding that could have important implications for the management of sunflower downy mildew. These data provide the first population-wide picture of the genetic structure of P. halstedii at a fine spatial scale.publishedVersio

What’s the meaning of local? Using molecular markers to define seed transfer zones for ecological restoration in Norway

-According to the Norwegian Diversity Act, practitioners of restoration in Norway are instructed to use seed mixtures of local provenance. However, there are no guidelines for how local seed should be selected. In this study, we use genetic variation in a set of alpine species (Agrostis mertensii, Avenella flexuosa, Carex bigelowii, Festuca ovina, Poa alpina and Scorzoneroides autumnalis) to define seed transfer zones to reduce confusion about the definition of ‘local seeds’. The species selected for the study are common in all parts of Norway and suitable for commercial seed production. The sampling covered the entire alpine region (7–20 populations per species, 3–15 individuals per population). We characterised genetic diversity using amplified fragment length polymorphisms. We identified different spatial genetic diversity structures in the species, most likely related to differences in reproductive strategies, phylogeographic factors and geographic distribution. Based on results from all species, we suggest four general seed transfer zones for alpine Norway. This is likely more conservative than needed for all species, given that no species show more than two genetic groups. Even so, the approach is practical as four seed mixtures will serve the need for restoration of vegetation in alpine regions in Norway

Transcriptional profiling of defense responses to Botrytis cinerea infection in leaves of Fragaria vesca plants soil-drenched with β-aminobutyric acid

Grey mold caused by the necrotrophic fungal pathogen Botrytis cinerea can affect leaves, flowers, and berries of strawberry, causing severe pre- and postharvest damage. The defense elicitor β-aminobutyric acid (BABA) is reported to induce resistance against B. cinerea and many other pathogens in several crop plants. Surprisingly, BABA soil drench of woodland strawberry (Fragaria vesca) plants two days before B. cinerea inoculation caused increased infection in leaf tissues, suggesting that BABA induce systemic susceptibility in F. vesca. To understand the molecular mechanisms involved in B. cinerea susceptibility in leaves of F. vesca plants soil drenched with BABA, we used RNA sequencing to characterize the transcriptional reprogramming 24 h post-inoculation. The number of differentially expressed genes (DEGs) in infected vs. uninfected leaf tissue in BABA-treated plants was 5205 (2237 upregulated and 2968 downregulated). Upregulated genes were involved in pathogen recognition, defense response signaling, and biosynthesis of secondary metabolites (terpenoid and phenylpropanoid pathways), while downregulated genes were involved in photosynthesis and response to auxin. In control plants not treated with BABA, we found a total of 5300 DEGs (2461 upregulated and 2839 downregulated) after infection. Most of these corresponded to those in infected leaves of BABA-treated plants but a small subset of DEGs, including genes involved in ‘response to biologic stimulus‘, ‘photosynthesis‘ and ‘chlorophyll biosynthesis and metabolism’, differed significantly between treatments and could play a role in the induced susceptibility of BABA-treated plants.publishedVersio

Transcriptional profiling of defense responses to Botrytis cinerea infection in leaves of Fragaria vesca plants soil-drenched with beta-aminobutyric acid

Grey mold caused by the necrotrophic fungal pathogen Botrytis cinerea can affect leaves, flowers, and berries of strawberry, causing severe pre- and postharvest damage. The defense elicitor beta-aminobutyric acid (BABA) is reported to induce resistance against B. cinerea and many other pathogens in several crop plants. Surprisingly, BABA soil drench of woodland strawberry (Fragaria vesca) plants two days before B. cinerea inoculation caused increased infection in leaf tissues, suggesting that BABA induce systemic susceptibility in F. vesca. To understand the molecular mechanisms involved in B. cinerea susceptibility in leaves of F. vesca plants soil drenched with BABA, we used RNA sequencing to characterize the transcriptional reprogramming 24 h post-inoculation. The number of differentially expressed genes (DEGs) in infected vs. uninfected leaf tissue in BABA-treated plants was 5205 (2237 upregulated and 2968 downregulated). Upregulated genes were involved in pathogen recognition, defense response signaling, and biosynthesis of secondary metabolites (terpenoid and phenylpropanoid pathways), while downregulated genes were involved in photosynthesis and response to auxin. In control plants not treated with BABA, we found a total of 5300 DEGs (2461 upregulated and 2839 downregulated) after infection. Most of these corresponded to those in infected leaves of BABA-treated plants but a small subset of DEGs, including genes involved in 'response to biologic stimulus', 'photosynthesis' and 'chlorophyll biosynthesis and metabolism', differed significantly between treatments and could play a role in the induced susceptibility of BABA-treated plants.Peer reviewe

Transcriptional profiling of defense responses to Botrytis cinerea infection in leaves of Fragaria vesca plants soil-drenched with β-aminobutyric acid

Grey mold caused by the necrotrophic fungal pathogen Botrytis cinerea can affect leaves, flowers, and berries of strawberry, causing severe pre- and postharvest damage. The defense elicitor β-aminobutyric acid (BABA) is reported to induce resistance against B. cinerea and many other pathogens in several crop plants. Surprisingly, BABA soil drench of woodland strawberry (Fragaria vesca) plants two days before B. cinerea inoculation caused increased infection in leaf tissues, suggesting that BABA induce systemic susceptibility in F. vesca. To understand the molecular mechanisms involved in B. cinerea susceptibility in leaves of F. vesca plants soil drenched with BABA, we used RNA sequencing to characterize the transcriptional reprogramming 24 h post-inoculation. The number of differentially expressed genes (DEGs) in infected vs. uninfected leaf tissue in BABA-treated plants was 5205 (2237 upregulated and 2968 downregulated). Upregulated genes were involved in pathogen recognition, defense response signaling, and biosynthesis of secondary metabolites (terpenoid and phenylpropanoid pathways), while downregulated genes were involved in photosynthesis and response to auxin. In control plants not treated with BABA, we found a total of 5300 DEGs (2461 upregulated and 2839 downregulated) after infection. Most of these corresponded to those in infected leaves of BABA-treated plants but a small subset of DEGs, including genes involved in ‘response to biologic stimulus‘, ‘photosynthesis‘ and ‘chlorophyll biosynthesis and metabolism’, differed significantly between treatments and could play a role in the induced susceptibility of BABA-treated plants

Spatial Genetic Structure and Pathogenic Race Composition at the Field Scale in the Sunflower Downy Mildew Pathogen, Plasmopara halstedii

Yield losses in sunflower crops caused by Plasmopara halstedii can be up to 100%, depending on the cultivar susceptibility, environmental conditions, and virulence of the pathogen population. The aim of this study was to investigate the genetic and phenotypic structure of a sunflower downy mildew agent at the field scale. The genetic diversity of 250 P. halstedii isolates collected from one field in southern France was assessed using single-nucleotide polymorphisms (SNPs) and single sequence repeats (SSR). A total of 109 multilocus genotypes (MLG) were identified among the 250 isolates collected in the field. Four genotypes were repeated more than 20 times and spatially spread over the field. Estimates of genetic relationships among P. halstedii isolates using principal component analysis and a Bayesian clustering approach demonstrated that the isolates are grouped into two main genetic clusters. A high level of genetic differentiation among clusters was detected (FST = 0.35), indicating overall limited exchange between them, but our results also suggest that recombination between individuals of these groups is not rare. Genetic clusters were highly related to pathotypes, as previously described for this pathogen species. Eight different races were identified (100, 300, 304, 307, 703, 704, 707, and 714), with race 304 being predominant and present at most of the sites. The co-existence of multiple races at the field level is a new finding that could have important implications for the management of sunflower downy mildew. These data provide the first population-wide picture of the genetic structure of P. halstedii at a fine spatial scale

Monitoring of the Apple Fruit Moth: Detection of Genetic Variation and Structure Applying a Novel Multiplex Set of 19 STR Markers

The apple fruit moth Argyresthia conjugella (Lepidoptera, Yponomeutidae) is a seed predator of rowan (Sorbus aucuparia) and is distributed in Europe and Asia. In Fennoscandia (Finland, Norway and Sweden), rowan fruit production is low every 2–4 years, and apple (Malus domestica) functions as an alternative host, resulting in economic loss in apple crops in inter-mast years. We have used Illumina MiSeq sequencing to identify a set of 19 novel tetra-nucleotide short tandem repeats (STRs) in Argyresthia conjugella. Such motifs are recommended for genetic monitoring, which may help to determine the eco-evolutionary processes acting on this pest insect. The 19 STRs were optimized and amplified into five multiplex PCR reactions. We tested individuals collected from Norway and Sweden (n = 64), and detected very high genetic variation (average 13.6 alleles, He = 0.75) compared to most other Lepidoptera species studied so far. Spatial genetic differentiation was low and gene flow was high in the test populations, although two non-spatial clusters could be detected. We conclude that this set of genetic markers may be a useful resource for population genetic monitoring of this economical important insect species

Monitoring of the Apple Fruit Moth: Detection of Genetic Variation and Structure Applying a Novel Multiplex Set of 19 STR Markers

The apple fruit moth Argyresthia conjugella (Lepidoptera, Yponomeutidae) is a seed predator of rowan (Sorbus aucuparia) and is distributed in Europe and Asia. In Fennoscandia (Finland, Norway and Sweden), rowan fruit production is low every 2–4 years, and apple (Malus domestica) functions as an alternative host, resulting in economic loss in apple crops in inter-mast years. We have used Illumina MiSeq sequencing to identify a set of 19 novel tetra-nucleotide short tandem repeats (STRs) in Argyresthia conjugella. Such motifs are recommended for genetic monitoring, which may help to determine the eco-evolutionary processes acting on this pest insect. The 19 STRs were optimized and amplified into five multiplex PCR reactions. We tested individuals collected from Norway and Sweden (n = 64), and detected very high genetic variation (average 13.6 alleles, He = 0.75) compared to most other Lepidoptera species studied so far. Spatial genetic differentiation was low and gene flow was high in the test populations, although two non-spatial clusters could be detected. We conclude that this set of genetic markers may be a useful resource for population genetic monitoring of this economical important insect species.publishedVersio

Genetic analyses of saprolegnia strains isolated from salmonid fish of different geographic origin document the connection between pathogenicity and molecular diversity

Saprolegnia parasitica is recognized as one of the most important oomycetes pests of salmon and trout species. The amplified fragment length polymorphism (AFLP) and method sequence data of the internal transcribed spacer (ITS) were used to study the genetic diversity and relationships of Saprolegnia spp. collected from Canada, Chile, Japan, Norway and Scotland. AFLP analysis of 37 Saprolegnia spp. isolates using six primer combinations gave a total of 163 clear polymorphic bands. Bayesian cluster analysis using genetic similarity divided the isolates into three main groups, suggesting that there are genetic relationships among the isolates. The unweighted pair group method with arithmetic mean (UPGMA) and principal coordinate analysis (PCO) confirmed the pattern of the cluster analyses. ITS analyses of 48 Saprolegnia sequences resulted in five well-defined clades. Analysis of molecular variance (AMOVA) revealed greater variation within countries (91.01%) than among countries (8.99%). We were able to distinguish the Saprolegnia isolates according to their species, ability to produce oogonia with and without long spines on the cysts and their ability to or not to cause mortality in salmonids. AFLP markers and ITS sequencing data obtained in the study, were found to be an efficient tool to characterize the genetic diversity and relationships of Saprolegnia spp. The comparison of AFLP analysis and ITS sequence data using the Mantel test showed a very high and significant correlation (r2 = 0.8317).publishedVersio

Genetic Diversity in Apple Fruit Moth Indicate Different Clusters in the Two Most Important Apple Growing Regions of Norway

The apple fruit moth (Argyresthia conjugella (A. conjugella)) in Norway was first identified as a pest in apple production in 1899. We here report the first genetic analysis of A. conjugella using molecular markers. Amplified fragment length polymorphism (AFLP) analysis was applied to 95 individuals from six different locations in the two most important apple-growing regions of Norway. Five AFLP primer combinations gave 410 clear polymorphic bands that distinguished all the individuals. Further genetic analysis using the Dice coefficient, Principal Coordinate analysis (PCO) and Bayesian analyses suggested clustering of the individuals into two main groups showing substantial genetic distance. Analysis of molecular variance (AMOVA) revealed greater variation among populations (77.94%) than within populations (22.06%) and significant and high FST values were determined between the two major regions (Distance = 230 km, FST = 0.780). AFLP analysis revealed low to moderate genetic diversity in our population sample from Norway (Average: 0.31 expected heterozygosity). The positive significant correlation between the geographic and the molecular data (r2 = 0.6700) indicate that genetic differences between the two major regions may be due to geographical barriers such as high mountain plateaus (Hardangervidda) in addition to isolation by distance (IBD)