Department of Immunology and Molecular Biology, School of Biomedical Sciences, Makerere University College of Health Sciences, Kampala, Uganda.
Department of Medical Microbiology, School of Biomedical Sciences, Makerere University College of Health Sciences, Kampala, Uganda.
PLoS One. 2018 May 30;13(5):e0198091. doi: 10.1371/journal.pone.0198091. eCollection 2018.
Accurate diagnosis of tuberculosis, especially by using rapid molecular assays, can reduce transmission of drug resistant tuberculosis in communities. However, the frequency of resistance conferring mutations varies with geographic location of Mycobacterium tuberculosis, and this affects the efficiency of rapid molecular assays in detecting resistance. This has created need for characterizing drug resistant isolates from different settings to investigate frequencies of resistance conferring mutations. Here, we describe the prevalence and patterns of rifampicin- and isoniazid- resistance conferring mutations in isolates from Uganda, which could be useful in the management of MDR-TB patients in Uganda and other countries in sub-Saharan Africa.
Ninety seven M. tuberculosis isolates were characterized, of which 38 were MDR, seven rifampicin-resistant, 12 isoniazid-mono-resistant, and 40 susceptible to rifampicin and isoniazid. Sequence analysis of the rpoB rifampicin-resistance determining region (rpoB/RRDR) revealed mutations in six codons: 588, 531, 526, 516, 513, and 511, of which Ser531Leu was the most frequent (40%, 18/45). Overall, the three mutations (Ser531Leu, His526Tyr, Asp516Tyr) frequently associated with rifampicin-resistance occurred in 76% of the rifampicin resistant isolates while 18% (8/45) of the rifampicin-resistant isolates lacked mutations in rpoB/RRDR. Furthermore, sequence analysis of katG and inhA gene promoter revealed mainly the Ser315Thr (76%, 38/50) and C(-15)T (8%, 4/50) mutations, respectively. These two mutations combined, which are frequently associated with isoniazid-resistance, occurred in 88% of the isoniazid resistant isolates. However, 20% (10/50) of the isoniazid-resistant isolates lacked mutations both in katG and inhA gene promoter. The sensitivity of sequence analysis of rpoB/RRDR for rifampicin-resistance via detection of high confidence mutations (Ser531Leu, His526Tyr, Asp516Tyr) was 81%, while it was 77% for analysis of katG and inhA gene promoter to detect isoniazid-resistance via detection of high confidence mutations (Ser315Thr, C(-15)T, T(-8)C). Furthermore, considering the circulating TB genotypes in Uganda, the isoniazid-resistance conferring mutations were more frequent in M. tuberculosis lineage 4/sub-lineage Uganda, perhaps explaining why this genotype is weakly associated with MDR-TB.
Sequence analysis of rpoB/RRDR, katG and inhA gene promoter is useful in detecting rifampicin/isoniazid resistant M. tuberculosis isolates in Uganda however, about ≤20% of the resistant isolates lack known resistance-conferring mutations hence rapid molecular assays may not detect them as resistant.
准确诊断结核病，尤其是使用快速分子检测方法，可以减少耐药结核在社区中的传播。然而，耐药相关突变的频率因结核分枝杆菌的地理位置而异，这影响了快速分子检测方法检测耐药性的效率。这就需要对来自不同环境的耐药分离株进行特征描述，以研究耐药相关突变的频率。在这里，我们描述了乌干达分离株中利福平-和异烟肼耐药相关突变的流行情况和模式，这对于乌干达和撒哈拉以南非洲其他国家的耐多药结核病患者的管理可能是有用的。
对 97 株结核分枝杆菌进行了特征描述，其中 38 株为耐多药，7 株为利福平耐药，12 株为异烟肼单耐药，40 株对利福平和异烟肼敏感。rpoB 区（rpoB/RRDR）的序列分析显示，在 6 个密码子中存在突变：588、531、526、516、513 和 511，其中 Ser531Leu 是最常见的（40%，18/45）。总的来说，与利福平耐药相关的三个突变（Ser531Leu、His526Tyr、Asp516Tyr）在 76%的利福平耐药分离株中出现，而 18%（8/45）的利福平耐药分离株中 rpoB/RRDR 缺失突变。此外，katG 和 inhA 基因启动子的序列分析显示，主要是 Ser315Thr（76%，38/50）和 C（-15）T（8%，4/50）突变。这两个与异烟肼耐药相关的突变，在 88%的异烟肼耐药分离株中同时出现。然而，20%（10/50）的异烟肼耐药分离株在 katG 和 inhA 基因启动子中均缺乏突变。rpoB/RRDR 序列分析检测高可信度突变（Ser531Leu、His526Tyr、Asp516Tyr）对利福平耐药的敏感性为 81%，而分析 katG 和 inhA 基因启动子检测高可信度突变（Ser315Thr、C（-15）T、T（-8）C）对异烟肼耐药的敏感性为 77%。此外，考虑到乌干达的结核分枝杆菌流行株，结核分枝杆菌 4 亚群/乌干达亚群中的异烟肼耐药相关突变更为常见，这可能解释了为什么该基因型与耐多药结核病的相关性较弱。
rpoB/RRDR、katG 和 inhA 基因启动子的序列分析可用于检测乌干达的利福平/异烟肼耐药结核分枝杆菌分离株，但约 20%的耐药分离株缺乏已知的耐药相关突变，因此快速分子检测方法可能无法检测到这些耐药分离株。
