Abstract: Objective To understand the prevalence of drug-resistant Mycobacterium(M.) tuberculosis in Wenzhou, Zhejiang province, and provide evidence for the prevention and control of drug-resistant tuberculosis (TB). Methods The susceptibility tests of 196 M. tuberculosis strains isolated from smear positive TB patients to four first line anti-TB drugs (isoniazid, rifampicin, streptomycin and ethambutol) and nine second-line anti-TB drugs (kanamycin, amikacin, capreomycin, p-aminosalicylic acid, protionamide, cyclocerine, ofloxacin, levofloxacin, moxifloxacin) were performed. Results A total of 191 M. tuberculosis strains were cultured. The overall drug-resistant rate was 25.1% (48/191); the initial drug-resistant rate was 21.9% (36/164) and the acquired drug-resistant rate was 44.4% (12/27), the difference was significant (2=6.23,P0.05).The multidrugresistant (MDR) rate was 6.8% (13/191), in which the initial and the acquired MDR rates were 4.3%(7/164) and 22.2% (6/27), respectively, the difference was significant (2=9.12,P0.01). The resistant rates to streptomycin, isoniazid, rifampicin and ethambutol were 19.9%, 14.7%, 7.9% and 3.1%, respectively. The resistant rates to one, two, three and four first line anti TB drugs were 14.7%, 5.2%, 2.6% and 2.6%, respectively. The overall drugresistant rate and MDR rate were 25.5%(36/141) and 6.4%(9/141), respectively, in males and 24.0% (12/50) and 8.0% (4/50) in females,the differences were not significant (2=0.05,2=0.93,allP0.05). The age specific overall drug resistant rate showed no significant differences (2=5.34,P0.05). In 13 MDR strains, 8 (61.5%) showed resistance to at least one secondline anti-TB drugs tested. For second line anti-TB drugs, the resistant rate to ofloxacin and p-aminosalicylic acid was highest(5/13), followed by that to cyclocerine (4/13), that to levofloxacin, moxifloxacin (1/4) and that to capreomycin (1/13). Conclusion The prevalence of MDR-TB was high in Wenzhou, it is necessary to strengthen the prevention and control of drugresistant TB by conducting the susceptibility test to the second line anti-TB drugs and improving the detection and treatment of drug resistant TB patients