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Collar rot of vegetables by Sclerotium rolfsii is seriously a damaging disease worldwide. Tropical and sub-tropical weather with high temperature and soil moisture favors disease severity and its spread. At least 500 species of crops are the hosts of Sclerotium rolfsii. Variations in pathogenicity, aggressiveness or virulence, geographical locations and seasonal influence exist among the isolates of the pathogen. Eight isolates of S. rolfsii were established from the collection of the pathogen from eggplant, tomato, spinach and lentil from four districts of Bangladesh. The isolates were characterized on their morphology, growth and molecular aspects. Growth rate, lesion size on the inoculated plant and plant mortality rate goverened the virulence of the isolates as determined through RAPD analysis. Isolates from tomato proved most virulent.