The efficacy of silver generated larvicide with the help of entomopathogenic fungi, Isaria fumosorosea (Ifr) against major vector mosquitoes Culex quinquefasciatus and Aedes aegypti. The Ifr-silver nanoparticles (AgNPs) were characterized structurally and functionally using UV-visible spectrophotometer followed by scanning electron microscopy (SEM), energy-dispersive X-ray (EDX) spectroscopy and Fourier transform infrared (FTIR) spectra. The optimum pH (alkaline), temperature (30 °C) and agitation (150 rpm) for AgNP synthesis and its stability were confirmed through colour change. Ae. aegypti larvae (I-IV instars) were found highly susceptible to synthesized AgNPs than the larvae of Cx. quinquefasciatus. However, the mortality rate was indirectly proportional to the larval instar and the concentration. The lethal concentration that kills 50% of the exposed larvae (LC50) and lethal concentration that kills 90% of the exposed larvae (LC90) values of the tested concentration are 0.240, 0 0.075.337, 0.430, 0.652 and 1.219, 2.210, 2.453, 2.916; 0.065, 0.075, 0.098, 0.137 and 0.558, 0.709, 0.949, 1.278 ppm with respect to 0.03 to 1.00 ppm of Ifr-AgNPs against first, second, third and fourth instars of Cx. quinquefasciatus and Ae. aegypti, respectively. This is the first report for synthesis of AgNPs using Ifr against human vector mosquitoes. Hence, Ifr-AgNPs would be significantly used as a potent mosquito larvicide.