<p>Cleaning of public spaces for human activities is commonly employed and reduces the microbial load in these spaces. However, the mechanisms underlying changes in microbial community structure following cleaning remain poorly understood. As a pilot study, we investigated the bacterial community in public toilets before and after cleaning using 16S rRNA sequencing and computational simulations, as well as bacterial colony growth assays and metabolic functional prediction. The results revealed that cleaning initially imposes a strong disturbance on bacterial communities, followed by continuous stochastic loss and a concurrent influx of bacteria, resulting in changes in community structure. Culturable bacterial communities before and after cleaning showed equivalent growth properties, indicating that cleaning did not alter the original environment. Cleaning time was positively correlated with colony number but not with colony growth capacity, suggesting that bacteria surviving cleaning were fast-growing or highly abundant taxa. Metabolisms related to methane and to diverse environments showed increased contributions to survival after cleaning. Taken together, cleaning-mediated changes in bacterial community structure in public toilets were driven by a stochastic process that renewed bacterial diversity. This study provides new insights into the disturbance and recovery dynamics of the microbiome in public environments subjected to cleaning.</p>