Morphological and Molecular Characterization of Colletotrichum nymphaeae Causing Anthracnose on Strawberry Fruit

Abstract

Isolates associated with anthracnose symptoms on strawberry (Fragaria × ananassa Duch.) fruits were investigated to determine the morphological and molecular identity of the causal pathogen Colletotrichum nymphaeae. Diseased fruit samples exhibiting sunken lesions, soft rot, and orange conidial masses were collected from commercial cultivation fields and subjected to standard mycological isolation procedures. Pure cultures developed on potato dextrose agar showed fast-growing colonies with gray to white mycelium and abundant acervuli formation under laboratory conditions. Conidia were hyaline, cylindrical to oblong, and aseptate, consistent with Colletotrichum species morphology. Pathogenicity tests on healthy strawberry fruits confirmed virulence, reproducing typical anthracnose lesions and satisfying Koch’s postulates. Molecular identification was performed using ITS rDNA, GAPDH, and β-tubulin gene sequencing, which confirmed high similarity with reference sequences of Colletotrichum nymphaeae. Phylogenetic analysis placed all isolates within a well-supported clade of the C. acutatum species complex, with limited but detectable genetic variability among isolates from different sampling locations. Observed sequence polymorphisms suggest localized adaptation and possible introduction of diverse genotypes through planting material exchange. Integrated morphological and multilocus molecular data provided robust confirmation of species identity and clarified its role as a dominant anthracnose pathogen in strawberry production systems. The study emphasizes the necessity of precise pathogen identification for effective disease management and resistance breeding programs. Early detection using molecular diagnostics can significantly improve disease forecasting and reduce postharvest losses in strawberry supply chains. These findings contribute to the global understanding of Colletotrichum diversity and its epidemiological significance in high-value horticultural crops under changing climatic and production conditions.