Insecticides have been broadly used to control cockroaches but these insects have developed resistance to several pesticides. Entomopathogenic fungi can be ideal for the biocontrol of cockroaches as it is environment-friendly microbial pesticide. The aim of the present study is to determine the effect of Hirsutella thompsonii, an entomopathogenic fungus on mortality and antioxidant enzymes of Periplaneta Americana. Conidial suspensions of fungi were given to cockroaches through different modes for 24 hours. We observed the positive effect of H. thompsonii on both mortality and the antioxidant enzyme (catalase, superoxide dismutase and glutathione peroxidase) activity in different tissues of cockroaches. The mortality rate in cockroaches ranged between 54.98% and 63.45% when treated with H. thompsonoii through different modes. The results showed that the enzyme activity of catalase (1.23±0.1 to 17±0.3 U/mg pot.), superoxide dismutase (3.3±0.3 to 20.1±0.3 U/mg prot.) and glutathione peroxidase (2.2±0.1 to 11.7±0.2 U/mg prot.) varied in different tissues of cockroaches. The present study reveals that the H. thompsonii have great potential for biocontrol against insects.