ObjectiveTo study the relationship between the mutations in rpoB gene of rifamycinresistant Mycobacterium tuberculosis(M. tuberculosis) and the level of rifabutin resistance.MethodsThe minimal inhibition concentrations(MICs) of 64 rifamycinresistant isolates and 6 sensitive isolates of M. tuberculosis to rifabutin were determined by the dilution method, their isonicotinyl hydrazide (INH) resistance were analyzed. The whole rpoB gene were amplified and sequenced, then the relationship between the mutations in rpoB gene and the level of rifabutin resistance and multidrug resistance were analyzed.ResultsThe sensitive isolates had no mutations in rpoB gene and their MICs were 0.25 mg/L -0.50 mg/L. All rifamycinresistant isolates had mutations in rpoB gene. Among 37 highlevel rifabutinresistant strains (MICs≥4 mg/L), mutations of S531L, H526R, and Y389C were found in 27,2 and 2 isolates respectively, comutation of S531W, H526Y, Q513K, V176F, D516Y and Q253R was in 1 strain, comutation of D516G and L511P was in 1 strain. Of 17 intermediatedlevel rifabutinresistant strains (MICs 2-4 mg/L), 16 isolates were with S531L mutation and 1 with D516G combined with L511P and S509R mutation,respectively . Of 10 lowlevel rifabutinresistant strains (MICs 0.25-1 mg/L), single mutation of L533P, H526L, H526S, D516V, and D516Y were found in 2 isolates respectively. 93.75％(60/64) of rifamycinresistant isolates were also resistant to INH.ConclusionDetection of rpoB mutation can preliminarily screen multidrugresistant M. tuberculosis; The predominant mutations of rpoB gene in highand middlelevel rifabutinresistant isolates are S531L, the mutation positions and types of rpoB gene in M. tuberculosis correlates with the level of rifabutin resistance.