Ocheretina Oksana, Escuyer Vincent E, Mabou Marie-Marcelle, Royal-Mardi Gertrude, Collins Sean, Vilbrun Stalz C, Pape Jean W, Fitzgerald Daniel W
Center for Global Health, Division of Infectious Diseases, Department of Medicine, Weill Cornell Medical College, New York, New York, United States of America; Les Centres GHESKIO, Port-au-Prince, Haiti.
Laboratory of Mycobacteriology, Wadsworth Center, New York State Department of Health, Albany, New York, United States of America.
PLoS One. 2014 Mar 5;9(3):e90569. doi: 10.1371/journal.pone.0090569. eCollection 2014.
The World Health Organization has recommended use of molecular-based tests MTBDRplus and GeneXpert MTB/RIF to diagnose multidrug-resistant tuberculosis in developing and high-burden countries. Both tests are based on detection of mutations in the Rifampin (RIF) Resistance-Determining Region of DNA-dependent RNA Polymerase gene (rpoB). Such mutations are found in 95-98% of Mycobacterium tuberculosis strains determined to be RIF-resistant by the "gold standard" culture-based drug susceptibility testing (DST). We report the phenotypic and genotypic characterization of 153 consecutive clinical Mycobacterium tuberculosis strains diagnosed as RIF-resistant by molecular tests in our laboratory in Port-au-Prince, Haiti. 133 isolates (86.9%) were resistant to both RIF and Isoniazid and 4 isolates (2.6%) were RIF mono-resistant in MGIT SIRE liquid culture-based DST. However the remaining 16 isolates (10.5%) tested RIF-sensitive by the assay. Five strains with discordant genotypic and phenotypic susceptibility results had RIF minimal inhibitory concentration (MIC) close to the cut-off value of 1 µg/ml used in phenotypic susceptibility assays and were confirmed as resistant by DST on solid media. Nine strains had sub-critical RIF MICs ranging from 0.063 to 0.5 µg/ml. Finally two strains were pan-susceptible and harbored a silent rpoB mutation. Our data indicate that not only detection of the presence but also identification of the nature of rpoB mutation is needed to accurately diagnose resistance to RIF in Mycobacterium tuberculosis. Observed clinical significance of low-level resistance to RIF supports the re-evaluation of the present critical concentration of the drug used in culture-based DST assays.
世界卫生组织已建议在发展中国家和高负担国家使用基于分子的检测方法MTBDRplus和GeneXpert MTB/RIF来诊断耐多药结核病。这两种检测方法均基于对依赖DNA的RNA聚合酶基因（rpoB）的利福平（RIF）耐药决定区突变的检测。在通过“金标准”基于培养的药敏试验（DST）确定为对RIF耐药的结核分枝杆菌菌株中，95%-98%存在此类突变。我们报告了在海地太子港我们实验室中通过分子检测诊断为对RIF耐药的153株连续临床结核分枝杆菌菌株的表型和基因型特征。在基于MGIT SIRE液体培养的DST中，133株分离株（86.9%）对RIF和异烟肼均耐药，4株分离株（2.6%）为RIF单耐药。然而，其余16株分离株（10.5%）通过该检测对RIF敏感。5株基因型和表型药敏结果不一致的菌株，其RIF最低抑菌浓度（MIC）接近表型药敏试验中使用的1μg/ml的临界值，并通过固体培养基上的DST确认为耐药。9株菌株的RIF MIC处于亚临界范围，为0.063至0.5μg/ml。最后，2株菌株对所有药物敏感，但携带沉默的rpoB突变。我们的数据表明，为准确诊断结核分枝杆菌对RIF的耐药性，不仅需要检测rpoB突变的存在，还需要鉴定其性质。观察到的对RIF低水平耐药的临床意义支持重新评估目前基于培养的DST试验中使用的该药物的临界浓度。
