Soil microorganisms play a critical role in influencing plant growth and managing soil pathogens. Tobacco mosaic virus (TMV) induces significant economic losses in global agriculture and can impact the composition and function of soil microbial communities. Despite its importance, the interactions between viruses and soil microbial communities remain inadequately understood. In this study, we employed 16S rRNA gene sequencing coupled with bioinformatics analyses to thoroughly investigate the bacterial communities and physicochemical properties of the rhizosphere soils of healthy tobacco plants under both sterilized (WJ) and non-sterilized (YJ) conditions, as well as TMV-infected tobacco plants under both sterilized (WT) and non-sterilized (YT) conditions. Our findings demonstrated that TMV infection significantly modifies the physicochemical properties and bacterial community structure of rhizosphere soils, with these changes being more pronounced in non-sterilized soils. Moreover, the YT samples exhibited a more intricate network of bacterial interactions. They showed significant differences from WT samples in key bacterial genera that might be involved in the response to or antagonism of TMV. The genera Burkholderia-Caballeronia-Paraburkholderia and Dyella were highlighted. These results suggest that rhizosphere microorganisms actively respond to TMV infection, with a more pronounced response observed in non-sterilized soils. This study provides novel insights into the microbial dynamics associated with TMV infection and underscores the importance of soil microbial communities in plant health and disease resistance. Additionally, it offers an experimental framework for future research on soil-borne diseases, emphasizing the pivotal role of soil microbiota in disease ecology and soil impact.