Morphological and Molecular Characterization of Ramularia beticola Causing Leaf Spot on Sugar Beet

Abstract

Leaf spot symptoms characterized by small angular necrotic lesions, chlorosis, premature defoliation, and reduced photosynthetic area were observed on sugar beet (Beta vulgaris) crops cultivated under humid temperate conditions. Diseased leaf samples collected from affected fields were used for pathogen isolation and characterization through morphological, pathogenicity, and molecular approaches. Fungal isolates consistently produced slow-growing colonies with pale to greyish mycelia and abundant conidiophores bearing hyaline, septate conidia typical of Ramularia beticola. Considerable variation in colony morphology, sporulation intensity, and lesion development was observed among isolates obtained from different sugar beet-growing regions. Pathogenicity assays conducted on healthy sugar beet plants reproduced characteristic leaf spot symptoms, including necrotic lesions and progressive foliar blighting, whereas control plants remained symptom-free. Re-isolation of the fungus from inoculated tissues fulfilled Koch’s postulates, confirming its pathogenic role. Molecular characterization was performed using amplification and sequencing of the internal transcribed spacer (ITS) region and additional conserved gene loci associated with fungal phylogenetic studies. Sequence analysis revealed high similarity with authenticated R. beticola isolates available in global sequence databases. Phylogenetic analysis grouped the isolates within the R. beticola clade with strong bootstrap support, confirming species identity and indicating limited genetic variation among regional populations. Disease development was strongly favored by prolonged leaf wetness, high humidity, and moderate temperatures, which enhanced spore germination and infection efficiency. Severe infections led to significant reduction in leaf longevity and sugar accumulation, ultimately affecting yield and quality of sugar beet production.