Thirty-eight cows in a herd were determined to be positive for bovine tuberculosis (TB). The bacterial isolation and characterization with multiplex PCR identified six Mycobacterium tuberculosis isolates. The Mycobacterium bovis and M. tuberculosis infection induced comparable pathology in cattle in both gross pathology and histopathology based on the qualitative assessment of the sampled lung tissues. The spoligotyping demonstrated that cow M. tuberculosis isolates belonged to Beijing family strains. Meanwhile, the isolates from tuberculosis patients hospitalized in the local hospitals were assessed. No M. bovis strains were identified in 186 human isolates. Eighty-two percent (153/186) of M. tuberculosis isolates were Beijing-family strains. The 12 loci MIRU genotyping revealed that the first three prevalent patterns were 2232-2517-3533, 2233-2517-3533, and 2223-2517-3533. The bovine M. tuberculosis isolates were the third dominant MIRU pattern. The further 16 loci MIRU-VNTR assay confirmed that the bovine M. tuberculosis strains shared the same pattern suggesting there was a common source causing cow infection and an epidemiological link between cow and human M. tuberculosis infection. On the other hand, the retrospective investigation for the past three years' cases of TB patients from local hospitals revealed 0.34% (17/5011) prevalence of M. bovis infection in local people. In conclusion, in TB high-burden countries like China where bovine and human TB coexists, the fact that cattle maintain both M. bovis and M. tuberculosis would be a potential challenge to both Stop TB Plan of humans and bovine TB eradication scheme.