Mythimna separata is a destructive polyphagous pest of field crops. Insecticides are generally applied for its control which not only negatively affect natural enemies and the environment and cause resistance in the insect pests. There is a need for the friendly method which is safe for the environment and life. Currently, entomopathogenic fungi are being used as biological control agents for different insects. The influence of Isaria fumosorosea on survival, life table parameters, and enzymatic activities of M. separata were assessed. On the seventh day post-treatment, the highest concentration 3 × 108 spores/mL-1 caused the 92.5% larval mortality. The effect of LC15 and LC50 of I. fumosorosea were recorded on parental generation (F0) and first filial generation (F1) of M. separata. The life table parameters of F1 showed a decreasing trend in the intrinsic rate (r), net reproductive rate (Ro), mean generational time (T), total larval duration, and fecundity ratio in treated groups. In LC15 and LC50, groups the average fecundity ratio was 319.2 and 191.18 eggs/female, respectively. The activities of detoxifying enzymes were concentration-dependent and highest activities were recorded on the third day. I. fumosorosea negatively affected the growth parameters of M. separata and can be included in M. separata management program.