In this study the effects of nutrient media, temperature, Relative Humidity, light and pH on mycelial growth of three species of Pestalotiopsis (P. fici,P. guepinii and P. palmarum) were evaluated for the better growth. PSA was most suitable for mycelial growth, whereas, the PLA appeared to be the best medium for conidiomata formation. The fungi grew from 10 to 40ºC, with optimum growth at 20-30ºC with no growth at 5ºC. The growth of the fungus was observed to increase with increase in relative humidity from 35% to 100%. Different light regimes had impact on mycelium growth Darkness was more suitable for the growth, while alternative light caused growth rings appearance in cultures. No visible growth at 4 pH. The fungus grew at pH 4.5 – 7. Optimum growth occurred at pH 7 with 94% for P. palmarum and 74% for P. guepinii after 5 days of incubation at 25oC. A notable decline in growth occurred when pH increased from 7.5 to 9.