Twenty five samples from tomato cultivated soil were collected from April to October, 2010 in different areas of Nagpur, India. Three centimeter of the top soil was removed and 5 subsamples were then taken at random at a depth of 20 cm for each site. The soil samples were then transferred into sterile polyethylene bags and transported to the laboratory. All subsamples from one site were combined to yield one composite sample representing the location, exposed to room temperature with a humidity degree of 50% and sieved through a mesh of 2mm.
The soil samples were diluted into different concentration solution and vortexed well. The supernatant were then poured into plates of PDA (potato dextrose agar, with 10mg/ml stock solution of chloramphenicol) and incubated at 28 °C ±1 °C. Colonies appearing on the plates were isolated and re-inoculated into a new plate. After 7 days, single spore colonies were obtained by subculturing at 28 °C ±1 °C. The isolate was maintained on PDA medium at 4°C.
Isolation of pathogenic fungi
Fusarium oxysporum f.sp. lycopersici was isolated from stems and root tissues of infected tomato plants then sections of in lateral stem washed with three changes of sterilized distilled water and dried on filter paper. They were inoculated on PDA plates and incubated at 28 °C ±1 °C. Single spore culture was prepared[11]. Pathogenicity of the isolate toward tomato plant was estimated[12]. The isolate was maintained on PDA medium at 4°C.