Diversity and Specific Detection of <i>Pseudomonas syringae</i> Associated with Zucchini Squash Seed Production

Seed infestation is an important factor contributing to the dissemination of Pseudomonas syringae in vegetable crops. However, the broad ecological and genetic diversity found within the bacterial species has hindered a precise understanding of the populations responsible for phytosanitary disorders associated with commercial zucchini squash seed production. Here, we genotyped an international collection of over 260 P. syringae strains, as well as other cucurbit-associated bacteria that were primarily isolated from zucchini squash seedling grow-out assays using BOX and REP PCRs as well as through sequence analysis of the partial citrate synthase gene. While the population associated with zucchini squash seeds spanned the diversity of P. syringae phylogroup 2, the hybrid clade 2b-a was the dominant genetic group and comprised 78% of this population, followed by clade 2a at 13%. Seed and spray inoculation assays indicated that clade 2a and 2b-a strains induced severe foliar blighting and stunting of zucchini squash seedlings, while strains from other genetic groups were significantly less aggressive. Subsequently, a dual-probe real-time polymerase chain reaction assay targeting the syringolin biosynthesis gene, sylC, was developed to distinguish between P. syringae populations deemed to pose either a high or low phytosanitary risk to zucchini squash seed, based on prevalence and pathogenicity phenotyping. The sylC assay was then multiplexed with an internal positive control to enable use in routine seed health testing. The results of this study elucidate the diversity of the P. syringae population associated with zucchini squash seed production and help to improve seed-health testing methods.