Georghiou S B, Seifert M, Catanzaro D, Garfein R S, Valafar F, Crudu V, Rodrigues C, Victor T C, Catanzaro A, Rodwell T C
Department of Medicine, University of California, San Diego, La Jolla, California, USA
Department of Medicine, University of California, San Diego, La Jolla, California, USA.
Antimicrob Agents Chemother. 2016 Jun 20;60(7):3994-4004. doi: 10.1128/AAC.00222-16. Print 2016 Jul.
Molecular diagnostic assays, with their ability to rapidly detect resistance-associated mutations in bacterial genes, are promising technologies to control the spread of drug-resistant tuberculosis (DR-TB). Sequencing assays provide detailed information for specific gene regions and can help diagnostic assay developers prioritize mutations for inclusion in their assays. We performed pyrosequencing of seven Mycobacterium tuberculosis gene regions (katG, inhA, ahpC, rpoB, gyrA, rrs, and eis) for 1,128 clinical specimens from India, Moldova, and South Africa. We determined the frequencies of each mutation among drug-resistant and -susceptible specimens based on phenotypic drug susceptibility testing results and examined mutation distributions by country. The most common mutation among isoniazid-resistant (INH(r)) specimens was the katG 315ACC mutation (87%). However, in the Eastern Cape, INH(r) specimens had a lower frequency of katG mutations (44%) and higher frequencies of inhA (47%) and ahpC (10%) promoter mutations. The most common mutation among rifampin-resistant (RIF(r)) specimens was the rpoB 531TTG mutation (80%). The mutation was common in RIF(r) specimens in Mumbai (83%) and Moldova (84%) but not the Eastern Cape (17%), where the 516GTC mutation appeared more frequently (57%). The most common mutation among fluoroquinolone-resistant specimens was the gyrA 94GGC mutation (44%). The rrs 1401G mutation was found in 84%, 84%, and 50% of amikacin-resistant, capreomycin-resistant, and kanamycin (KAN)-resistant (KAN(r)) specimens, respectively. The eis promoter mutation -12T was found in 26% of KAN(r) and 4% of KAN-susceptible (KAN(s)) specimens. Inclusion of the ahpC and eis promoter gene regions was critical for optimal test sensitivity for the detection of INH resistance in the Eastern Cape and KAN resistance in Moldova. (This study has been registered at ClinicalTrials.gov under registration number NCT02170441.).
分子诊断检测技术能够快速检测细菌基因中与耐药性相关的突变，是控制耐药结核病（DR-TB）传播的有前景的技术。测序检测可为特定基因区域提供详细信息，并有助于诊断检测开发者确定检测中应优先纳入的突变。我们对来自印度、摩尔多瓦和南非的1128份临床标本的七个结核分枝杆菌基因区域（katG、inhA、ahpC、rpoB、gyrA、rrs和eis）进行了焦磷酸测序。我们根据表型药物敏感性检测结果确定了耐药和敏感标本中每种突变的频率，并按国家检查了突变分布情况。耐异烟肼（INH(r)）标本中最常见的突变是katG 315ACC突变（87%）。然而，在东开普省，INH(r)标本中katG突变的频率较低（44%），而inhA（47%）和ahpC（10%）启动子突变的频率较高。耐利福平（RIF(r)）标本中最常见的突变是rpoB 531TTG突变（80%）。该突变在孟买（83%）和摩尔多瓦（84%）的RIF(r)标本中很常见，但在东开普省不常见（17%），在东开普省516GTC突变出现得更频繁（57%）。耐氟喹诺酮标本中最常见的突变是gyrA 94GGC突变（44%）。rrs 1401G突变分别在84%、84%和50%的耐阿米卡星、耐卷曲霉素和耐卡那霉素（KAN(r)）标本中被发现。eis启动子突变-12T在26%的KAN(r)标本和4%的卡那霉素敏感（KAN(s)）标本中被发现。纳入ahpC和eis启动子基因区域对于在东开普省检测INH耐药性以及在摩尔多瓦检测KAN耐药性的最佳检测敏感性至关重要。（本研究已在ClinicalTrials.gov注册，注册号为NCT02170441。）
