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目的 研究结核分枝杆菌在不同pH值液体培养基中对吡嗪酰胺(PZA)敏感度的影响,同时比较不同pH值下结核分枝杆菌对吡嗪酰胺的敏感度与测序结果的一致率,探讨最适合对吡嗪酰胺进行药物敏感性试验(简称“药敏试验”)的pH值.方法 本研究采用简单随机抽样法中的直接抽选法,从2007年全国结核病耐药性基线调查的3929株结核分枝杆菌菌株中抽取348株;采用BACTEC MGIT 960系统在液体培养基pH值为5.3、5.5、5.7、5.9和6.1时,分别检测结核分枝杆菌对吡嗪酰胺的敏感度;同时对吡嗪酰胺耐药的结核分枝杆菌相关基因pmcA和rpsA进行测序,分析其突变特征;采用卡方检验统计2种方法检测吡嗪酰胺耐药率的差别,以P＜0.05为差异有统计学意义.结果 在348株结核分枝杆菌中,用MGITTM 960 PZA试剂盒(pH值为5.9)检测的吡嗪酰胺总耐药率为14.4％ (50/348),DNA测序结果有43株耐药,其中有30(69.8％)株检测到吡嗪酰胺药物耐药相关基因pncA发生突变,1株(2.3％)耐药分离株仅在rpsA有突变.在液体培养基pH值为6.1、5.9、5.7、5.5和5.3时,DNA测序结果与所有菌株表型药敏试验结果的一致率分别为94.8％ (330/348)、95.1％ (331/348)、96.8％(337/348)、95.2％ (316/332)和93.5％ (287/307).并且pH值为5.7时2种方法的耐药一致率(92.1％,35/38)明显高于pH值为5.9(76.0％,38/50)时(x2=3.961,P=0.047);而pH值为6.1 (72.7％,40/55)、pH值为5.5(93.3％,28/30)和pH值为5.3(95.0％,19/20)时的耐药一致率与pH值为5.9时的耐药一致率相比,差异无统计学意义(x2 =0.147,P=0.702;x2 =0.366,P=0.545;x2 =3.410,P=o.065).结论 液体培养基pH值为5.7时,结核分枝杆菌对吡嗪酰胺耐药与遗传突变检测结果有较好的相关性.“,”Objective To evaluate the effect of pH on the pyrazinamide (PZA) susceptibility of Mycobac terium tuberculosis strains in the BACTEC MGIT 960 liquid medium,to demonstrate the relationship between phenotype susceptibility and genetic changes among M.tuberculosis isolates,and to determine the most suitable pH value for evaluating pyrazinamide susceptibility.Methods A total of 348 isolates were randomly selected from those collected during the national drug resistance baseline survey conducted in 2007,and the PZA susceptibility of each M.tuberculosis isolate was detected using the BACTEC MGIT 960 automated system at pH 5.3,5.5,5.7,5.9 and 6.1.The pncA and rpsA genes from all isolates were sequenced for genetic mutations conferring PZA resistance.The concordance rate between phenotypic DST in different pH environments and genetic changes was evaluated using the Chi-square test,P＜0.05 being considered statistically significant.Results Of the 348 M.tuberculosis isolates,50 (14.4％) isolates resistant to PZA were detected using a commercial BACTEC MGIT 960 PZA kit at pH 5.9,and 43 using DNA sequencing,30 (69.8％) isolates harboring pncA mutations,and 1 (2.3％) isolate har boring mutations only in rpsA.The overall concordance rate between DNA sequencing and phenotypic DST results at different pH values was 94.8％ (330/348),95.1％ (331/348),96.8％ (337/348),95.2％ (316/332) and 93.5％ (287/307) at pH 6.1,pH 5.9,pH 5.7,pH 5.5 and pH 5.3,respectively.The difference between the pH 5.9 and pH 5.7 groups was statistically significant (x2 =3.961,P=0.047),but that between the pH 6.1,pH 5.5,pH 5.3 and pH 5.9 groups (x2 =0.147,P=0.702;x2 =0.366,P=0.545;x2 =3.410,P=0.065,respectively) was not statistically significant.Conclusion The strongest correlation between genetic changes in M.tuberculosis isolates and PZA resistance was obtained using liquid culture at pH 5.7.Using a pH 5.7 liquid medium may be the best approach to resolve the dilemma between PZase activity and mycobacterial growth during PZA susceptibility testing.