Bacillus licheniformis (B. licheniformis) is a microorganism with a wide range of probiotic properties and applications. Isolation and identification of novel strains is a major aspect of microbial research. Besides, the role of different carbon sources affects B. licheniformis in regulating micro-environment and the mechanisms need to be further investigated. In this study, we first isolated and identified a new strain of B. licheniformis from bovine rumen fluid. Microcrystalline cellulose (MC) and cellobiose (CB) as the certain carbon sources to treat strain. Further, a combination of transcriptome and proteome analyses was used to different carbon sources effects. The results showed that B. licheniformis ABC transporter proteins, antibiotic synthesis, flagellar assembly, cellulase-related pathways and proteins were significantly up-regulated in the MC treatment compared to the CB treatment, and lactate metabolism was inhibited. In addition, MC was used as a certain carbon source to improve bacterial inhibition of B. licheniformis, its own disease resistance and to regulate the rumen micro-environment. In conclusion, our research provides a potential new probiotic for feed research and a theoretical basis for investigating the mechanisms by which bacteria respond to different carbon sources.