Spot blotch, incited by Bipolaris sorokiniana, is one of the serious foliar diseases affecting wheat productivity worldwide, significantly. The cultural, morphological, pathogenic, and molecular variability were studied comprehensively among 51 B. sorokiniana isolates collected from diverse wheat-growing regions of India. Regarding cultural variability, the highest radial growth rate was observed in isolate Bs-9 (8.45 mm/day), whereas isolate Bs-27 exhibited the lowest growth rate (2.50 mm/day).