Morphological and Molecular Identification of Colletotrichum fructicola Causing Crown Rot on Strawberry Transplants

Abstract

Crown rot symptoms characterized by tissue browning, vascular discoloration, wilting, and collapse of strawberry transplants were observed in nurseries and production fields, leading to significant plant mortality and reduced stand establishment. Diseased crown tissues were collected and subjected to pathogen isolation and characterization using morphological and molecular approaches. Fungal isolates consistently produced fast-growing grayish to white colonies with abundant acervuli and setae typical of Colletotrichum species. Microscopic examination revealed hyaline, aseptate, cylindrical conidia consistent with descriptions of Colletotrichum fructicola. Considerable variation in colony morphology, sporulation intensity, and growth rate was observed among isolates obtained from different strawberry production sites. Pathogenicity assays conducted on healthy strawberry transplants reproduced typical crown rot symptoms, including crown necrosis, reduced root development, and plant collapse, whereas control plants remained symptom-free. Re-isolation of the fungus from inoculated tissues fulfilled Koch’s postulates and confirmed its pathogenic role. Molecular identification was performed through amplification and sequencing of the internal transcribed spacer (ITS) region along with additional conserved loci including β-tubulin and actin gene regions. Sequence analysis showed high similarity with reference C. fructicola isolates available in public databases. Phylogenetic analysis clustered the isolates within the C. fructicola clade with strong bootstrap support, confirming species identity and revealing limited genetic variability among regional populations. Disease development was favored by warm temperatures and high humidity conditions typical of strawberry nurseries and transplant production systems. Infected plants exhibited reduced vigor, poor establishment, and significant losses in commercial transplant quality. Integrated morphological and molecular approaches provided accurate identification of the pathogen and improved understanding of its epidemiology.