In this work, we conducted bacterial population profile studies to assess trends of rifampin (RIF) resistance of Mycobacterium tuberculosis isolates collected across China from 2005 to 2015. Totals of 273 and 269 randomly selected M. tuberculosis isolates from 2005 and 2015, respectively, were analyzed. The rates of RIF resistance (36.4%), isoniazid resistance (39.0%), and levofloxacin resistance (25.7%) in 2015 were significantly higher than those in 2005 (28.2%, 30.0%, and 15.4%, respectively; P < 0.05). Genotypic data revealed 256 (95.2%) Beijing-type isolates in 2015, a rate significantly higher than that in 2005 (86.4%) (P < 0.01). A higher proportion of mutations was identified within the rifampin resistance-determining region (RRDR) of rpoB in isolates from 2015 (99.0%) than in 2005 isolates (85.7%, P < 0.01). In addition, a significantly higher proportion of RIF-resistant isolates carrying compensatory mutations was observed in 2015 (31.6%) than in 2005 (7.8%). Notably, the great majority of these compensatory mutations (91.9%) were observed in isolates that harbored a mutation of codon 531 of the rpoB gene. In conclusion, our data demonstrate that resistance to RIF, isoniazid, and levofloxacin has become significantly more prevalent during the past decade. In addition, the prevalence of the Beijing genotype significantly increased from 2005 to 2015. Notably, a significantly increased frequency of strains with mutations in rpoC or rpoA is observed among those that have codon 531 mutations, which suggests that they may be compensatory and may play a role in facilitating transmission.