Frogeye leaf spot on soybeans is traditionally considered as a southern disease in the United States but its impact in North Central USA has been rising in recent years. In this study, we investigated the population structure and mating type distribution in the C. sojina population from Indiana, USA. Based on 27 single nucleotide polymorphism markers, 49 multi-locus genotypes (MLGs) were identified in 234 isolates collected from 29 counties in Indiana in 2020. Bayesian analysis grouped the 49 MLGs into three clusters. This grouping was supported by principal coordinate analysis and, in large part, by the unweighted pair group method with arithmetic mean and minimal spanning tree. Only one mating-type idiomorph was found in each isolate and in each MLG. The MAT1-1 idiomorph was found in 22 MLGs and the MAT1-2 idiomorph was found in 27 MLGs. Based on clone-corrected data, the distribution of mating-type idiomorphs did not deviate significantly from 1:1 ratio in Indiana as a whole and in 22 out of 24 counties where two or more MLGs were found. Thirty MLGs contained QoI-resistant isolates and 22 MLGs contained QoI-sensitive isolates, with three MLGs containing both types of isolates. MLG1, the most common MLG with 90 isolates, contained mostly QoI-resistant isolates. Interestingly, MLG1 was also the dominant genotype in the Tennessee population collected in 2015, suggesting that MLG1 has been a dominant genotype in a wider region for many years. Based on the standard index of association (r¯d), the Indiana population as a whole was in significant linkage disequilibrium. However, in five out of 16 counties where three or more MLGs were found, the null hypothesis of linkage equilibrium was not rejected. Tests of linkage disequilibrium between locus pairs showed that 33.3% of locus pairs on the same contigs were in significant disequilibrium and 17.7% of locus pairs on different contigs were in significant disequilibrium. The possibility of a cryptic sexual stage was discussed.
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