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Abstract：
In Mycobacterium tuberculosis, molecular predictions of ethambutol resistance rely primarily on the detection of mutations within embB. However, discordance between embB406 mutations and gold standard phenotypic drug sensitivity testing (DST) questions the significance of embB406 mutations used in molecular DST. This study tabulates embB mutations found in Canadian M. tuberculosis isolates and evaluates the impact of specific mutations on ethambutol resistance. The National Reference Centre for Mycobacteriology culture collection (n = 2796) was screened for isolates with embB mutations. Phenotypic DST was performed on the BACTEC™ MGIT™ 960 at ethambutol concentrations of 2-5 μg/mL. Whole genome sequencing was used for drug resistance predictions, phylogenomics and single nucleotide polymorphism analysis. Detection of resistance-associated embB mutations corresponded to a positive predictive value of 64.3%, negative predictive value of 99.2%, 98.7% specificity, and 73.3% sensitivity compared to phenotypic DST. Two embB406 mutation subtypes (Gly406Asp, Gly406Ala) were found among 16 isolates, of which 12 were sensitive at 5 µg/mL ethambutol with variable resistance between 2-4 µg/mL. A novel frameshift mutation in regulator embR (Gln258fs) was found in nine isolates. Mutations in embB406 were associated with low-level ethambutol resistance undetectable at the recommended critical concentration (5 μg/mL). These novel mutations may exacerbate variability in ethambutol resistance.
摘要：
在结核分枝杆菌中，乙胺丁醇抗性的分子预测主要依赖于embB内突变的检测。然而,embB406突变和金标准表型药物敏感性测试(DST)之间的不一致质疑分子DST中使用的embB406突变的重要性.本研究列出了在加拿大结核分枝杆菌分离株中发现的embB突变，并评估了特定突变对乙胺丁醇抗性的影响。国家分枝杆菌学培养物参考中心（n=2796）筛选了具有embB突变的分离株。表型DST在乙胺丁醇浓度为2-5μg/mL的BACTEC™MGIT™960上进行。全基因组测序用于耐药性预测，系统基因组学和单核苷酸多态性分析。检测到耐药性相关的embB突变对应的阳性预测值为64.3%，阴性预测值为99.2%，98.7%的特异性，与表型DST相比，敏感性为73.3%。两种embB406突变亚型（Gly406Asp，Gly406Ala)在16个分离株中发现，其中12对5µg/mL乙胺丁醇敏感，电阻在2-4µg/mL之间。在9个分离株中发现了调控因子embR（Gln258fs）的新移码突变。embB406中的突变与在推荐的临界浓度（5μg/mL）下无法检测到的低水平乙胺丁醇抗性有关。这些新的突变可能会加剧乙胺丁醇抗性的变异性。
