Wassihun Wedajo Aragaw, Tewodros T Gebresilase, Dereje A Negatu, Véronique Dartois, Thomas Dick
{"title":"脓肿分枝杆菌抗σ因子RshA功能缺失突变带来的多药耐受性","authors":"Wassihun Wedajo Aragaw, Tewodros T Gebresilase, Dereje A Negatu, Véronique Dartois, Thomas Dick","doi":"10.1128/aac.01051-24","DOIUrl":null,"url":null,"abstract":"<p><p>Low-level drug resistance in noncanonical pathways can constitute steppingstones toward acquisition of high-level on-target resistance mutations in the clinic. To capture these intermediate steps in <i>Mycobacterium abscessus</i> (Mab), we performed classic mutant selection experiments with moxifloxacin at twofold its minimum inhibitory concentration (MIC) on solid medium. We found that low-level resistance emerged reproducibly as loss-of-function mutations in RshA (MAB_3542c), an anti-sigma factor that negatively regulates activity of SigH, which orchestrates a response to oxidative stress in mycobacteria. Since oxidative stress is generated in response to many antibiotics, we went on to show that deletion of <i>rshA</i> confers low to moderate resistance-by measure of MIC-to a dozen agents recommended or evaluated for the treatment of Mab pulmonary infections. Interestingly, this moderate resistance was associated with a wide range of drug tolerance, up to 1,000-fold increased survival of a Δ<i>rshA</i> Mab mutant upon exposure to several β-lactams and DNA gyrase inhibitors. Consistent with the putative involvement of the SigH regulon, we showed that addition of the transcription inhibitor rifabutin (RBT) abrogated the high-tolerance phenotype of Δ<i>rshA</i> to representatives of the β-lactam and DNA gyrase inhibitor classes. In a survey of 10,000 whole Mab genome sequences, we identified several loss-of-function mutations in <i>rshA</i> as well as non-synonymous polymorphisms in two cysteine residues critical for interactions with SigH. Thus, the multidrug multiform resistance phenotype we have uncovered may not only constitute a step toward canonical resistance acquisition during treatment but also contribute directly to treatment failure.</p>","PeriodicalId":8152,"journal":{"name":"Antimicrobial Agents and Chemotherapy","volume":" ","pages":"e0105124"},"PeriodicalIF":4.1000,"publicationDate":"2024-12-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11619451/pdf/","citationCount":"0","resultStr":"{\"title\":\"Multidrug tolerance conferred by loss-of-function mutations in anti-sigma factor RshA of <i>Mycobacterium abscessus</i>.\",\"authors\":\"Wassihun Wedajo Aragaw, Tewodros T Gebresilase, Dereje A Negatu, Véronique Dartois, Thomas Dick\",\"doi\":\"10.1128/aac.01051-24\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Low-level drug resistance in noncanonical pathways can constitute steppingstones toward acquisition of high-level on-target resistance mutations in the clinic. To capture these intermediate steps in <i>Mycobacterium abscessus</i> (Mab), we performed classic mutant selection experiments with moxifloxacin at twofold its minimum inhibitory concentration (MIC) on solid medium. We found that low-level resistance emerged reproducibly as loss-of-function mutations in RshA (MAB_3542c), an anti-sigma factor that negatively regulates activity of SigH, which orchestrates a response to oxidative stress in mycobacteria. Since oxidative stress is generated in response to many antibiotics, we went on to show that deletion of <i>rshA</i> confers low to moderate resistance-by measure of MIC-to a dozen agents recommended or evaluated for the treatment of Mab pulmonary infections. Interestingly, this moderate resistance was associated with a wide range of drug tolerance, up to 1,000-fold increased survival of a Δ<i>rshA</i> Mab mutant upon exposure to several β-lactams and DNA gyrase inhibitors. Consistent with the putative involvement of the SigH regulon, we showed that addition of the transcription inhibitor rifabutin (RBT) abrogated the high-tolerance phenotype of Δ<i>rshA</i> to representatives of the β-lactam and DNA gyrase inhibitor classes. In a survey of 10,000 whole Mab genome sequences, we identified several loss-of-function mutations in <i>rshA</i> as well as non-synonymous polymorphisms in two cysteine residues critical for interactions with SigH. Thus, the multidrug multiform resistance phenotype we have uncovered may not only constitute a step toward canonical resistance acquisition during treatment but also contribute directly to treatment failure.</p>\",\"PeriodicalId\":8152,\"journal\":{\"name\":\"Antimicrobial Agents and Chemotherapy\",\"volume\":\" \",\"pages\":\"e0105124\"},\"PeriodicalIF\":4.1000,\"publicationDate\":\"2024-12-05\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11619451/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Antimicrobial Agents and Chemotherapy\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1128/aac.01051-24\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2024/10/29 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q2\",\"JCRName\":\"MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Antimicrobial Agents and Chemotherapy","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1128/aac.01051-24","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/10/29 0:00:00","PubModel":"Epub","JCR":"Q2","JCRName":"MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
非典型途径中的低水平耐药性可能是临床上获得高水平靶向耐药性突变的垫脚石。为了捕捉脓肿分枝杆菌(Mab)的这些中间步骤,我们在固体培养基上用最低抑菌浓度(MIC)两倍的莫西沙星进行了经典的突变体选择实验。我们发现,RshA(MAB_3542c)中的功能缺失突变可重复出现低水平抗性,RshA 是一种反σ因子,可负向调节 SigH 的活性,而 SigH 可协调分枝杆菌对氧化应激的反应。由于氧化应激是对许多抗生素的反应,我们继续研究发现,根据 MIC 指标,rshA 的缺失会产生低度到中度的抗药性,而这些抗药性是推荐或评估用于治疗分枝杆菌肺部感染的十几种药物的抗药性。有趣的是,这种中度耐药性与广泛的药物耐受性有关,当暴露于几种β-内酰胺类药物和DNA回旋酶抑制剂时,ΔrshA Mab突变体的存活率可提高1000倍。与推测的 SigH 调节子的参与相一致,我们发现转录抑制剂利福布丁(RBT)的加入可消除 ΔrshA 对β-内酰胺类和 DNA 回旋酶抑制剂的高耐受性表型。在对10,000个Mab全基因组序列的调查中,我们发现了rshA中的几个功能缺失突变,以及与SigH相互作用至关重要的两个半胱氨酸残基的非同义多态性。因此,我们发现的多药多型耐药性表型不仅可能是治疗过程中获得典型耐药性的一个步骤,还可能直接导致治疗失败。
Multidrug tolerance conferred by loss-of-function mutations in anti-sigma factor RshA of Mycobacterium abscessus.
Low-level drug resistance in noncanonical pathways can constitute steppingstones toward acquisition of high-level on-target resistance mutations in the clinic. To capture these intermediate steps in Mycobacterium abscessus (Mab), we performed classic mutant selection experiments with moxifloxacin at twofold its minimum inhibitory concentration (MIC) on solid medium. We found that low-level resistance emerged reproducibly as loss-of-function mutations in RshA (MAB_3542c), an anti-sigma factor that negatively regulates activity of SigH, which orchestrates a response to oxidative stress in mycobacteria. Since oxidative stress is generated in response to many antibiotics, we went on to show that deletion of rshA confers low to moderate resistance-by measure of MIC-to a dozen agents recommended or evaluated for the treatment of Mab pulmonary infections. Interestingly, this moderate resistance was associated with a wide range of drug tolerance, up to 1,000-fold increased survival of a ΔrshA Mab mutant upon exposure to several β-lactams and DNA gyrase inhibitors. Consistent with the putative involvement of the SigH regulon, we showed that addition of the transcription inhibitor rifabutin (RBT) abrogated the high-tolerance phenotype of ΔrshA to representatives of the β-lactam and DNA gyrase inhibitor classes. In a survey of 10,000 whole Mab genome sequences, we identified several loss-of-function mutations in rshA as well as non-synonymous polymorphisms in two cysteine residues critical for interactions with SigH. Thus, the multidrug multiform resistance phenotype we have uncovered may not only constitute a step toward canonical resistance acquisition during treatment but also contribute directly to treatment failure.
期刊介绍:
Antimicrobial Agents and Chemotherapy (AAC) features interdisciplinary studies that build our understanding of the underlying mechanisms and therapeutic applications of antimicrobial and antiparasitic agents and chemotherapy.