Division of Infection and Immunity, University College London, London, UK.
UCL Genetics Institute, University College London, London, UK.
Lancet Microbe. 2020 Aug;1(4):e165-e174. doi: 10.1016/S2666-5247(20)30031-8.
Bedaquiline and clofazimine are important drugs in the treatment of drug-resistant tuberculosis and are commonly used across southern Africa, although drug susceptibility testing is not routinely performed. In this study, we did a genotypic and phenotypic analysis of drug-resistant isolates from cohort studies in hospitals in KwaZulu-Natal, South Africa, to identify resistance-associated variants (RAVs) and assess the extent of clofazimine and bedaquiline cross-resistance. We also used a comprehensive dataset of whole-genome sequences to investigate the phylogenetic and geographical distribution of bedaquiline and clofazimine RAVs in southern Africa.
In this study, we included isolates reported from the PRAXIS study of patients with drug-resistant tuberculosis treated with bedaquiline (King Dinuzulu Hospital, Durban) and three other cohort studies of drug-resistant tuberculosis in other KwaZulu-Natal hospitals, and sequential isolates from six persistently culture-positive patients with extensively drug-resistant tuberculosis at the KwaZulu-Natal provincial referral laboratory. Samples were collected between 2013 and 2019. Microbiological cultures were done as part of all parent studies. We sequenced whole genomes of included isolates and measured bedaquiline and clofazimine minimum inhibitory concentrations (MICs) for isolates identified as carrying any variant or previously published , and RAVs, which were the subject of the phenotypic study. We combined all whole-genome sequences of obtained in this study with publicly available sequence data from other tuberculosis studies in southern Africa (defined as the countries of the Southern African Development Community), including isolates with variants identified by screening public genomic databases. We used this extended dataset to reconstruct phylogenetic relationships across lineage 2 and 4 isolates.
We sequenced the whole genome of 648 isolates from 385 patients with drug-resistant tuberculosis recruited into cohort studies in KwaZulu-Natal, and 28 isolates from six patients from the KwaZulu-Natal referral laboratory. We identified 30 isolates with RAVs from 16 (4%) of 391 patients. We did not identify any , or RAVs. MICs were measured for 21 isolates with RAVs. MICs were above the critical concentration for bedaquiline resistance in nine (43%) of 21 isolates, in the intermediate category in nine (43%) isolates, and within the wild-type range in three (14%) isolates. Clofazimine MICs in genetically wild-type isolates ranged from 0·12-0·5 μg/mL, and in isolates with RAVs from 0·25-4·0 μg/mL. Phylogenetic analysis of the extended dataset including isolates from southern Africa resolved multiple emergences of variants in lineages 2 and 4, documented two likely nosocomial transmission events, and identified the spread of a possibly bedaquiline and clofazimine cross-resistant clone in eSwatini. We also identified four patients with frameshift mutations that may confer resistance.
Bedaquiline and clofazimine cross-resistance in southern Africa is emerging repeatedly, with evidence of onward transmission largely due to mutations in . Roll-out of bedaquiline and clofazimine treatment in the setting of limited drug susceptibility testing could allow further spread of resistance. Designing strong regimens would help reduce the emergence of resistance. Drug susceptibility testing is required to identify where resistance does emerge.
Wellcome Trust, National Institute of Allergy and Infectious Diseases and National Center for Advancing Translational Sciences of the National Institutes of Health.
贝达喹啉和氯法齐明是治疗耐药结核病的重要药物，在整个南部非洲都有广泛应用，尽管尚未常规进行药物敏感性测试。在这项研究中，我们对来自南非夸祖鲁-纳塔尔省医院队列研究的耐药分离株进行了基因和表型分析，以确定耐药相关变异（RAV），并评估氯法齐明和贝达喹啉的交叉耐药程度。我们还使用了一个全面的全基因组序列数据集，调查了南部非洲贝达喹啉和氯法齐明 RAV 的系统发育和地理分布。
在这项研究中，我们纳入了来自贝达喹啉治疗耐药结核病患者的 PRAXIS 研究（德班 King Dinuzulu 医院）和其他三个夸祖鲁-纳塔尔省其他医院耐药结核病队列研究报告的分离株，以及来自夸祖鲁-纳塔尔省转诊实验室 6 例广泛耐药结核病持续培养阳性患者的连续分离株。样本采集于 2013 年至 2019 年期间。所有的母研究均进行了微生物培养。我们对纳入的分离株进行了全基因组测序，并测量了被鉴定为携带任何变异或先前发表的 、 和 RAV 的分离株的贝达喹啉和氯法齐明最小抑菌浓度（MIC），这些变异是表型研究的对象。我们将在这项研究中获得的所有 全基因组序列与来自南部非洲（定义为南部非洲发展共同体国家）的其他结核病研究的公开序列数据进行了组合，包括通过筛选公共基因组数据库鉴定的 变异的分离株。我们使用这个扩展数据集重建了谱系 2 和 4 分离株的系统发育关系。
我们对来自夸祖鲁-纳塔尔省队列研究的 385 名耐药结核病患者中的 648 个分离株进行了全基因组测序，并对来自夸祖鲁-纳塔尔省转诊实验室的 6 名患者中的 28 个分离株进行了测序。我们从 16 名（4%）391 名患者中鉴定出 30 株带有 RAV 的分离株。我们没有发现任何 或 RAV。我们测量了 21 株带有 RAV 的分离株的 MIC。在 9 株（43%）分离株中，贝达喹啉的 MIC 高于耐药临界值，在 9 株（43%）分离株中处于中间类别，在 3 株（14%）分离株中处于野生型范围内。遗传野生型分离株的氯法齐明 MIC 范围为 0.12-0.5μg/ml，而 RAV 分离株的 MIC 范围为 0.25-4.0μg/ml。包括来自南部非洲的 分离株的扩展数据集的系统发育分析解决了谱系 2 和 4 中多个 变异的出现，记录了两个可能的医院内传播事件，并确定了一种可能的贝达喹啉和氯法齐明交叉耐药克隆在斯威士兰的传播。我们还鉴定了 4 名可能携带耐药性的患者存在框移突变。
南部非洲的贝达喹啉和氯法齐明交叉耐药正在反复出现，证据表明主要是由于 突变导致的传播。在药物敏感性测试有限的情况下推出贝达喹啉和氯法齐明治疗可能会进一步导致耐药性的传播。设计强有力的治疗方案有助于减少耐药性的出现。需要进行药物敏感性测试以确定耐药性的出现。
惠康信托基金会、美国国立过敏和传染病研究所以及美国国立卫生研究院的国家中心转化科学。
