The gray mold fungus Botrytis cinerea is a major threat to fruit and vegetable production. Strawberry fields usually receive several fungicide treatments against Botrytis per season. Gray mold isolates from several German strawberry-growing regions were analyzed to determine their sensitivity against botryticides. Fungicide resistance was commonly observed, with many isolates possessing resistance to multiple (up to six) fungicides. A stronger variant of the previously described multidrug resistance (MDR) phenotype MDR1, called MDR1h, was found to be widely distributed, conferring increased partial resistance to two important botryticides, cyprodinil and fludioxonil. A 3-bp deletion mutation in a transcription factor-encoding gene, mrr1, was found to be correlated with MDR1h. All MDR1h isolates and the majority of isolates with resistance to multiple fungicides were found to be genetically distinct. Multiple-gene sequencing confirmed that they belong to a novel clade, called Botrytis group S, which is closely related to B. cinerea and the host-specific species B. fabae. Isolates of Botrytis group S genotypes were found to be widespread in all German strawberry-growing regions but almost absent from vineyards. Our data indicate a clear subdivision of gray mold populations, which are differentially distributed according to their host preference and adaptation to chemical treatments.

cBotrytis cinerea is one of the most important pathogens worldwide, causing gray mold on a large variety of crops. Botrytis pseudocinerea has been found previously to occur together with B. cinerea in low abundance in vineyards and strawberry fields. Here, we report B. pseudocinerea to be common and sometimes dominant over B. cinerea on several fruit and vegetable crops in Germany. On apples with calyx end rot and on oilseed rape, it was the major gray mold species. Abundance of B. pseudocinerea was often negatively correlated with fungicide treatments. On cultivated strawberries, it was frequently found in spring but was largely displaced by B. cinerea following fungicide applications. Whereas B. cinerea strains with multiple-fungicide resistance were common in these fields, B. pseudocinerea almost never developed resistance to any fungicide even though resistance mutations occurred at similar frequencies in both species under laboratory conditions. The absence of resistance to quinone outside inhibitors in B. pseudocinerea was correlated with an intron in cytB preventing the major G143A resistance mutation. Our work indicates that B. pseudocinerea has a wide host range similar to that of B. cinerea and that it can become an important gray mold pathogen on cultivated plants. Botrytis cinerea Pers.:Fr. is one of the most important plant pathogens worldwide, causing gray mold rot on far more than 200 plant species, including cultivated fruits, vegetables, and ornamental flowers. In addition to its wide host range, typical features of B. cinerea are a necrotrophic, macerating mode of infection, abundant production of asexual macroconidia on the surface of colonized host tissue, and formation of melanized sclerotia for long-term survival and sexual reproduction. Control of gray mold often relies on the use of fungicides although the efficacy of such treatments is threatened worldwide by the abundance of fungicide-resistant B. cinerea field strains (1). Based on gene sequence comparisons, the genus Botrytis has been divided into two phylogenetically separate clades (2). Clade 1 includes B. cinerea and B. pseudocinerea as well as the host-specific species B. fabae, B. calthae, and B. sinoviticola; all of these infect only dicot plants. Clade 2 is phylogenetically more diverse and currently comprises 23 host-specific Botrytis species that infect predominantly monocots (2-4).B. cinerea shows considerable variability in mycelial growth phenotype, conidiation, and sclerotium formation. Numerous studies have also documented a high degree of genetic variability within the species. Based on the analysis of PCR-restricted fragment length polymorphism (RFLP) patterns, fungicide resistance, and the detection of transposable elements, Giraud et al. provided evidence of the existence of genetically distinct groups within B. cinerea (5, 6). In particular, the presence or absence of the long terminal repeat retrotransposon Boty (7) and of the DNA transposon Flipper (8) was used to divide isolates into different transposon types, e...

Botrytis calthae is a necrotrophic plant pathogen, closely related to the ubiquitous broad host range fungus Botrytis cinerea, but highly host specific. Botrytis isolates from lesions of Caltha palustris grown at different locations were classified with genetic markers as either B. calthae or Botrytis pseudocinerea, or less frequently as B. cinerea. A PCR-based identification of B. calthae was developed. Seven haplotypes of B. calthae could be distinguished. Compared to B. cinerea, mycelium growth of B. calthae was similar, but conidiation less abundant, and sclerotia formation was only partially repressed by light. Conidia of B. calthae germinated more slowly, and showed a highly acidic optimum (pH 2Á5) compared to B. cinerea conidia (pH 5Á3). All B. calthae isolates were sensitive to common anti-Botrytis fungicides, but showed partial resistance to the succinate dehydrogenase inhibitors boscalid, fluopyram and carboxin. Infection experiments revealed a weak capability of B. calthae to induce necrotic lesions on plants that are hosts for B. cinerea. On C. palustris leaves, B. calthae induced similar lesions to B. cinerea. These data provide a basis for comparative molecular investigation of the physiology and host specificity of B. calthae and closely related Botrytis species.

Botrytis cinerea is a world-wide occurring plant pathogen, causing pre- and post-harvest gray mold rot on a large number of fruit, vegetable, and flower crops. B. cinerea is closely related to Botrytis pseudocinerea, another broad host range species which often occurs in sympatry with B. cinerea, and to several host-specific species including Botrytis fabae and Botrytis calthae. B. cinerea populations have been shown to be genetically heterogeneous, and attempts have been made to correlate genetic markers to virulence and host adaptation. Here, we present the development of a multilocus sequence typing (MLST) scheme, with 10 genes selected for high variability and phylogenetic congruence, to evaluate the genetic diversity of B. cinerea, B. fabae, and B. pseudocinerea. Using PacBio-assisted simultaneous mass sequencing of PCR products, MLST analysis of about 100 strains from diverse geographical origins and years of isolation was performed, which resulted in high-resolution strain differentiation and robust species separation. Several B. cinerea strains formed an as yet unknown population, referred to as group B, which was well separated from all other B. cinerea strains. Furthermore, the gene cluster for biosynthesis of the phytotoxin botcinic acid was missing in B. cinerea B strains. B. cinerea strains from the monocot Iris pseudacorus were found to form a genetically distinct population, and contained an intact gene cluster for production of the red pigment bikaverin, which is usually degenerated in B. cinerea. Remarkably, these strains were much more aggressive on Iris than other B. cinerea strains, which is the first unequivocal example for host specialization in B. cinerea. Our data reveal new insights into the genetic diversity of B. cinerea and provide evidence for intraspecific differentiation and different degrees of host adaptation of this polyphagous necrotrophic pathogen.

causes pre- and postharvest decay of many fruit and vegetable crops. A survey of German strawberry fields revealed strains that differed from in diagnostic PCR markers and growth appearance. Phylogenetic analyses showed that these strains belong to an undescribed species in clade 2, named sp. nov. Isolates of were detected in strawberry fields throughout Germany, sometimes at frequencies similar to those of, and in the southeastern United States. was isolated from overwintering strawberry tissue but not from freshly infected fruit. invaded strawberry tissues with an efficiency similar to or lower than that of but showed poor colonization of inoculated nonhost plant tissues. These data and the exclusive occurrence of this fungus on strawberry plants indicate that is host specific and has a tissue preference different from that of Various fungicide resistance patterns were observed in populations. Many strains showed resistance to one or several chemical classes of fungicides and an efflux-based multidrug resistance (MDR1) phenotype previously described in Resistance-related mutations in were identical or similar to those of for carbendazim (E198A mutation in ), azoxystrobin (G143A in), iprodione (G367A+V368F in ), and MDR1 (gain-of-function mutations in the transcription factor gene and overexpression of the drug efflux transporter gene ). The widespread occurrence of indicates that this species is adapted to fungicide-treated strawberry fields and may be of local importance as a gray mold pathogen alongside Gray mold is the most important fruit rot on strawberries worldwide and requires fungicide treatments for control. For a long time, it was believed to be caused only by, a ubiquitous pathogen with a broad host range that quickly develops fungicide resistance. We report the discovery and description of a new species, named , that is widely distributed in commercial strawberry fields in Germany and the southeastern United States. It was observed on overwintering tissue but not on freshly infected fruit and seems host specific on the basis of its occurrence and artificial infection tests. has also developed resistance to several fungicides that is caused by mutations similar to those known in , including an efflux-based multidrug resistance. Our data indicate that could be of practical importance as a strawberry pathogen in some regions where its abundance is similar to that of .