Antibiotic resistance among hypermutator carbapenem resistant Klebsiella pneumoniae

超突变碳青霉烯类耐药肺炎克雷伯菌的抗生素耐药性

基本信息

批准号：
10532461
负责人：
M. Hong Thi NGUYEN
金额：
$ 26.02万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-06-21 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10532461
关键词：
Antibiotic Resistance Antibiotic Therapy Antibiotics Bacteria Biological Ceftazidime Chronic Clinical Clinical Microbiology DNA DNA biosynthesis Data Defect Development Enterobacteriaceae Infections Exhibits Future Gastrointestinal tract structure Gene Mutation Genes Genetic Recombination Genomics Horizontal Gene Transfer Human In Vitro Infection Intravenous Investigation Klebsiella pneumoniae Laboratories Link Literature Mediating Meropenem Microbiology Mismatch Repair Molecular Mus Mutation Organ Organ Transplantation Outcome Patients Phenotype Plasmids Population Prospective Studies Public Health Resistance Resistance development Role Sampling Sepsis Solid Stress Survivors Testing Tissues United States Validation carbapenem resistance carbapenem-resistant Enterobacteriaceae carbapenemase chronic infection colistin resistance effective therapy emerging antibiotic resistance environmental change experimental study fitness gastrointestinal gene repair genome sequencing gut colonization improved improved outcome innovation insight molecular sequence database mortality mouse model mutant novel organ transplant recipient pathogen promoter recurrent infection repository resistance gene resistant Klebsiella pneumoniae response screening stem success tool transmission process treatment strategy whole genome

项目摘要

Project Summary Carbapenem resistant Enterobacteriaceae (CRE) are major public health threats. CR-Klebsiella pneumoniae (CRKP) are the most common CRE globally. Treatment of CRKP and other CRE infections with new antibiotics like ceftazidime-avibactam (CZA) and meropenem-vaborbactam (MVB) has improved survival, but recurrent infections are common and emergent resistance is problematic. CRE infections are usually due to strains that colonize the GI tract. Among other bacteria, it is now apparent that GI colonization is caused by a population of closely related, but genetically distinct strains. These strains can exhibit a range of antibiotic resistance and biologic attributes, which are often not appreciated by studying single colonies from microbiologic cultures. In studies of natural bacterial populations, ~1 to 5% of isolates exhibit high spontaneous mutation rates, which may confer selective advantages under environmental stress and increase diversity within the population. Such hypermutation (HM) strains are most prevalent in humans during chronic colonization and recurrent infections, and they most often stem from mutations to DNA mismatch repair (MMR) genes like mutS, mutL or mutH. There are few studies of HM among CRKP or CRE. We believe that HM is under-recognized in these bacteria because studies have not assessed long-term GI colonization, recurrent infections, or populations of strains from clinical samples. In this project, we hypothesize that 1) HM CRKP can be recovered from patients with chronic GI colonization and persistent/recurrent infections; 2) MMR and other gene mutations promote HM in vitro, and CZA and MVB resistance in vitro and within infected organs; and 3) these mutations promote transmission and receipt of antibiotic resistance gene (ARG)-bearing plasmids by CRKP in vitro and during GI colonization. In pilot screening studies, we recovered HM CRKP from ~30% of patients with chronic GI colonization, or persistent or recurrent infections. MMR gene mutations were identified in most HM CRKP strains. One of these mutations (MutH V76G) was proven to contribute to HM, MVB and CZA resistance, and enhanced transfer and acceptance of plasmids containing ARGs in vitro, and to CRKP tissue burdens and emergence of MVB resistance within infected organs of intravenously (IV)-infected mice. In aim 1 of this proposal, we will continue to screen clinical CRKP isolates for HM phenotype. We will perform whole genome sequencing on HM isolates, and create isogenic mutant strains to determine if certain mutations contribute to HM. In aim 2, we will evaluate the role of HM mutations in emergence of CZA, MVB and colistin resistance in vitro and within mouse organs following IV infection. Then, we will determine impact of HM mutations on transfer of ARG-bearing plasmids in vitro and during mouse GI colonization. This project will mark the first systematic investigations of CRE clinical isolates for HM phenotypes. If successful, experiments will provide new understanding of HM and its crucial roles in regulating CRKP antibiotic resistance. Isogenic HM strains created here will also be powerful tools for assessing resistance barriers for novel antibiotics and new targets of antibiotic discovery.

项目概要碳青霉烯类耐药肠杆菌（CRE）是主要的公共卫生威胁。 CR-肺炎克雷伯菌 (CRKP) 是全球最常见的 CRE。用新型抗生素治疗 CRKP 和其他 CRE 感染头孢他啶-阿维巴坦 (CZA) 和美罗培南-伐硼巴坦 (MVB) 等药物可提高生存率，但复发率较高感染很常见，出现的耐药性是个问题。 CRE 感染通常是由以下菌株引起的定植于胃肠道。在其他细菌中，现在很明显，胃肠道定植是由一群细菌引起的密切相关但遗传上不同的菌株。这些菌株可以表现出一系列抗生素耐药性生物学属性，通过研究微生物培养物中的单个菌落通常无法理解这些属性。在对自然细菌种群的研究表明，约 1% 至 5% 的分离株表现出较高的自发突变率，这可能在环境压力下赋予选择性优势并增加人口的多样性。这样的超突变（HM）菌株在慢性定植和反复感染期间在人类中最为普遍，它们通常源于 DNA 错配修复 (MMR) 基因（如 mutS、mutL 或 mutH）的突变。那里关于CRKP或CRE中HM的研究很少。我们认为 HM 在这些细菌中未被充分认识，因为研究尚未评估长期胃肠道定植、反复感染或临床菌株种群样品。在这个项目中，我们假设 1) HM CRKP 可以从慢性胃肠道患者中恢复定植和持续/复发感染； 2）MMR和其他基因突变在体外促进HM，以及CZA 体外和受感染器官内的 MVB 耐药性； 3）这些突变促进传播和接收在体外和胃肠道定植过程中通过 CRKP 产生抗生素抗性基因 (ARG) 携带质粒。试点中筛选研究中，我们从约 30% 患有慢性胃肠道定植、或持续或反复感染。在大多数 HM CRKP 菌株中发现了 MMR 基因突变。这些突变之一 (MutH V76G) 被证明有助于抵抗 HM、MVB 和 CZA，并增强转移和接受能力体外含有 ARG 的质粒，以及 CRKP 组织负荷和 MVB 抗性的出现静脉注射（IV）感染小鼠的感染器官。在该提案的目标 1 中，我们将继续筛选临床 CRKP 分离株具有 HM 表型。我们将对 HM 分离株进行全基因组测序，并创建等基因突变株以确定某些突变是否导致 HM。在目标 2 中，我们将评估体外和 IV 后小鼠器官内出现 CZA、MVB 和粘菌素耐药性的 HM 突变感染。然后，我们将确定 HM 突变对体外携带 ARG 的质粒转移的影响，以及在小鼠胃肠道定植期间。该项目将标志着 CRE 临床分离株的首次系统研究对于 HM 表型。如果成功，实验将提供对 HM 及其关键作用的新认识调节 CRKP 抗生素耐药性。这里创建的同基因 HM 菌株也将成为评估的强大工具新型抗生素的耐药性障碍和抗生素发现的新靶点。