In order to achieve the food demand of a growing population, agricultural productivity needs to be increased by employing safe strategies. In the present study we have evaluated ZnONPs that were synthesized from the culture supernatant of Bacillus subtilis. Bio mimetically synthesized ZnONPs showed a surface resonance peak of 355 nm corresponding to NPs formation. Further, NPs were examined for their size, shape and element confirmation by DLS, AFM, SEM, TEM and EDAX, which confirmed the synthesized NPs were nearly spherical in size with average diameter of 32 nm by TEM. Surface charge of + 34.3 mV was observed for NPs with a low poly-dispersity index of 0.21. In vitro efficacy studies against fungi Colletotrichum capsici, Sclerotium rolfsii, Alternaria solani and Fusarium oxysporum f. sp. cicero showed up to 99% mycelial growth inhibition at 0.125% ZnONPs. Further, in-vitro disk-diffusion assay showed inhibition zones of 23 ± 0.4 mm and 12.67 ± 0.24 mm for Xanthomonas axonopodis pv. punicae (Xap) and Xanthomonas oryzae pv. oryzae (Xoo) bacterial cultures. Plant toxicity study was observed that ≤ 0.14% NPs concentration was safe under greenhouse conditions. Overall, the present study emphasizes the potential effect of ZnONPs against agricultural pathogens which play an important role in agriculture production.