Genetic diversity of the spinach downy mildew pathogen based on hierarchical sampling

Downy mildew, caused by the obligate oomycete pathogen Peronospora effusa, is the most economically important disease of spinach. In the past 30 years, 14 new races and 13 strains with novel virulence have been identified. However, the mechanism(s) driving the rapid evolution of virulence remains unknown. To understand reproductive strategies potentially driving the emergence of new races in P. effusa, 67 composite isolates (a collection of symptomatic leaves from a single cultivar grown in a defined area) of P. effusa obtained from 13 states between 2010 and 2018 were used to analyze the population genetic diversity hierarchically. Genotypes at 33 SNP loci of 719 lesions from these 67 isolates were determined by targeted sequencing. Diversity was then evaluated among individual lesions within the composite isolates, between isolates, host cultivars, geographic locations, and years of isolates collected. A total of 380 genotypes were identified from 719 individual lesions. Of the 380 genotypes, 350 (92%) were unique while the most common genotype was identified in 110 lesions of 16 isolates collected from 13 cultivars from CA and AZ in 2016. Variation within composite isolates ranged from none (a single genotype among lesions from a composite isolate) to 38 unique genotypes recovered from 39 lesions of a composite isolate. An index of association analysis suggested asexual (clonal) and sexual reproduction play important roles in population structure. Based on discriminant analysis of principal components, four distinct subpopulations were identified. Host cultivar, origin, and time of collection had an effect on population differentiation, and genotypes specific to a certain location or collection period were identified. Some subpopulations were unique to certain areas, and were only detected after 2014-2016. The co-existence of sexual and asexual reproduction strategies may partially explain the rapid emergence and spread of new races and novel strains of P. effusa.