BackgroundPleurotus ostreatus is an edible mushroom popularly cultivated worldwide. Distilled grain waste (DGW) is a potential substrate for P. ostreatus cultivation. However, components in DGW restrict P. ostreatus mycelial growth. Therefore, a cost-effective approach to facilitate rapid P. ostreatus colonization on DGW substrate will benefit P. ostreatus cultivation and DGW recycling.ResultsFive dominant indigenous bacteria, Sphingobacterium sp. X1, Ureibacillus sp. X2, Pseudoxanthomonas sp. X3, Geobacillus sp. X4, and Aeribacillus sp. X5, were isolated from DGW and selected to develop a consortium-based microbial agent to compost DGW for P. ostreatus cultivation. Microbial agent inoculation led to faster carbohydrate metabolism, a higher temperature (73.2 vs. 71.2 °C), a longer thermophilic phase (5 vs. 3 days), and significant dynamic changes in microbial community composition and diversity in composts than those of the controls. Metagenomic analysis showed the enhanced microbial metabolisms, such as xenobiotic biodegradation and metabolism and terpenoid and polyketide metabolism, during the mesophilic phase after microbial agent inoculation, which may facilitate the fungal colonization on the substrate. In accordance with the bioinformatic analysis, a faster colonization of P. ostreatus was observed in the composts with microbial inoculation than in control after composting for 48 h, as indicated from substantially higher fungal ergosterol content, faster lignocellulose degradation, and higher lignocellulase activities in the former than in the latter. The final mushroom yield shared no significant difference between composts with microbial inoculation and control, with 0.67 ± 0.05 and 0.60 ± 0.04 kg fresh mushroom/kg DGW, respectively (p > 0.05).ConclusionThe consortium-based microbial agent comprised indigenous microorganisms showing application potential in composting DGW for providing substrate for P. ostreatus cultivation and will provide an alternative to facilitate DGW recycling.