Highlights: • TSM-pathogenic strains of B. bassiana were not selective against adults of three Neoseiulus species. • Neoseiulus barkeri was less susceptible to fungal infections than N. californicus and N. idaeus. • Very low RH conditions negatively affect the biology and activity of Neoseiulus spp. • The predatory mites N. californicus and N. idaeus were more tolerant to drought conditions. Abstract: Predatory mites and invertebrate-pathogenic fungi have been used as biological control agents in a complementary way to suppress two-spotted spider mite (TSM) populations, which are a key pest of horticultural crops worldwide. However, their performance can be lowered by both biotic (e.g., intraguild interactions) and abiotic stress (especially relative humidity - RH). The present study evaluated the susceptibility of three predatory phytoseiid mite species in the genus Neoseiulus – N. californicus, N. idaeus, and N. barkeri – to infection by selected spider mite-pathogenic Beauveria bassiana strains as well as their predation capacity and resilience under simulated drought conditions. The two selected B. bassiana strains induced TSM mortality above 70 %. All three Neoseiulus mite species also showed susceptibility to both strains; however, N. barkeri was less susceptible to fungal infections than the other Neoseiulus species (with mortality < 65 % and without reduction of oviposition). All three Neoseiulus species were negatively affected by low RH conditions. Neoseiulus barkeri was the most affected with a drastic reduction in TSM egg consumption (about 40 %) and preventing the deposition of eggs by females. The egg consumption by N. californicus and N. idaeus was not affected by low RH conditions; however, both predatory mites reduced the number of eggs laid per female. Thus, the efficiency of the conversion of food into egg biomass for all three Neoseiulus species decreased under low RH conditions, dropping to zero for N. barkeri. We showed that the simultaneous use of B. bassiana and Neoseiulus had a detrimental effect on predatory mite survival and progeny, indicating the need for further development of new strategies which combine these biological control agents.