Beauveria brongniartii specifically infects the insect pest Melolontha melolontha and is commonly isolated from soil and larvae. Since 1990, it has been used commercially as a biological control agent (BCA) against the larvae. Recent research revealed that B. pseudobassiana was the most frequently isolated pathogen from adult beetles collected aboveground across 35 alpine sites. This led us to hypothesize that B. pseudobassiana primarily infects adults aboveground, while B. brongniartii mainly targets larvae in soil. To test these hypotheses, we investigated the occurrence and distribution of both species at two M. melolontha-infested sites in Switzerland using a combination of phylogenetic analyses and microsatellite markers. Species-specific microsatellite markers were developed for B. pseudobassiana, and existing markers were applied to B. brongniartii. From 399 Beauveria spp. isolates collected from adult beetles, soil, grassland plants, and tree leaves, 362 were identified as B. brongniartii, which was present in all sources except tree leaves, and 37 as B. pseudobassiana, found in all sources including tree leaves, except soil. The most common B. brongniartii multilocus genotypes (MLGs) were those of the BCAs BIPESCO 2 and BIPESCO 4, perhaps reflecting previous BCA applications at these locations. One B. pseudobassiana microsatellite-based MLG was isolated from both tree leaves and M. melolontha in the tree canopy. Additionally, B. pseudobassiana isolated from the phylloplane exhibited pathogenicity toward M. melolontha adults, suggesting the possibility of acquiring infections aboveground. To detail B. pseudobassiana intraspecific diversity, a sequence dataset was constructed including 13 unique haplotypes of the nuclear intergenic region Bloc sequenced in this study, 58 Bloc sequences downloaded from GenBank, and sphyngomyelin phosphodiesterase, glycosyl hydrolase, and AAA-ATPase midasin I sequences from 18 isolates illustrative of B. pseudobassiana genome diversity accessioned in GenBank. Phylogenetic analysis of this data reveals the species to comprise a cryptic complex with distinct genetic clusters that group isolates independently of their geographic origin and isolation substrate. Overall, this study shows B. brongniartii predominates in soil, while B. pseudobassiana occurs in the phylloplane, suggesting its potential as a complementary biological control for adult beetles aboveground.
