Aims: To investigate the thermal biology of entomopathogenic fungi being examined as potential microbial control agents of Varroa destructor , an ectoparasite of the European honey bee Apis mellifera . Methods and Results: Colony extension rates were measured at three temperatures (20, 30 and 35degreesC) for 41 isolates of entomopathogenic fungi. All of the isolates grew at 20 and 30degreesC but only 11 isolates grew at 35degreesC. Twenty-two isolates were then selected on the basis of appreciable growth at 30-35degreesC (the temperature range found within honey bee colonies) and/or infectivity to V. destructor , and their colony extension rates were measured at 10 temperatures (12.5-35degreesC). This data were then fitted to Schoolfield et al . [J Theor Biol (1981)88:719-731] re-formulation of the Sharpe and DeMichele [J Theor Biol (1977)64:649-670] model of poikilotherm development. Overall, this model accounted for 87.6-93.9% of the data variance. Eleven isolates exhibited growth above 35degreesC. The optimum temperatures for extension rate ranged from 22.9 to 31.2degreesC. Only three isolates exhibited temperature optima above 30degreesC. The super-optimum temperatures (temperature above the optimum at which the colony extension rate was 10% of the maximum rate) ranged from 31.9 to 43.2degreesC. Conclusions: The thermal requirements of the isolates examined against V. destructor are well matched to the temperatures in the broodless areas of honey bee colonies, and a proportion of isolates, should also be able to function within drone brood areas. Significance and Impact of the Study: Potential exists for the control of V. destructor with entomopathogenic fungi in honey bee colonies. The methods employed in this study could be utilized in the selection of isolates for microbial control prior to screening for infectivity and could help in predicting the activity of a fungal control agent of V. destructor under fluctuating temperature conditions.