Investigating the role of prophage encoded gene products on mycobacterial gene expression and intrinsic antibiotic resistance

项目摘要 这项研究的目的是了解原噬菌体是如何调节分枝杆菌的 抗生素耐药性和改变基因表达。耐药性感染引起的 由临床上重要的分枝杆菌引起的疾病仍然是一个重大的公共卫生负担。 结核分枝杆菌是一种新出现的病原体在囊性纤维化患者的治疗 成功率只有45%，被认为是最耐药的分枝杆菌之一。 耐药分离株通常表现出内在抗生素耐药基因的表达增加， 使得药物治疗具有挑战性。M.肠球菌分离株通常是溶原菌，这意味着它们的 基因组携带一个或多个前噬菌体，整合的病毒基因组，有可能 调节内在的抗生素耐药性。原噬菌体改变基因表达的机制 和分枝杆菌的抗生素耐药性还不清楚，这是这项赠款的重点。 转录调节因子whiB 7的表达在应激反应中增加，例如亚应激反应。 抗生素的抑制浓度和巨噬细胞的细胞内环境， 在分枝杆菌内在抗生素耐药性和巨噬细胞存活中起关键作用 （八）、我们的实验室首次证明原噬菌体有助于内在抗生素 耐药性和whiB 7表达增加，这反过来又积极调节大的 一组内在抗生素抗性基因（8-10）。我们发现了一组新的前噬菌体 在M. P.属于MabR（7，11）簇的龟类， 确定MabR原噬菌体McProf增加M.龟类对 氨基糖苷类抗生素是M.鹅/龟感染（9）。我们 发现携带McProf的分枝杆菌具有增强的whiB 7反应，特别是当 被第二个噬菌体重复感染尚不清楚McProf如何与 第二个原噬菌体驱动whiB 7表达和内在抗性的变化。我们 假设McProf基因组表达基因检测应激并调节 通过诱导whiB 7在分枝杆菌中产生抗生素耐药性。了解 在分枝杆菌溶原菌中whiB 7调节的机制，我们提出表征 在携带McProf的溶原菌中增强whiB 7反应的细胞应激 原噬菌体，并鉴定和表征McProf基因，其上调whiB 7以响应 应力

PROJECT SUMMARY The goal of the proposed research is to understand how prophages regulate mycobacterial antibiotic resistance and alter gene expression in mycobacteria. Drug-resistant infections caused by clinically important mycobacteria continue to be a significant public health burden. Mycobacterium abscessus is an emerging pathogen in cystic fibrosis patients with a treatment success rate of only 45%, and is considered one of the most drug-resistant mycobacteria. Resistant isolates commonly display increased expression of intrinsic antibiotic resistance genes, making drug treatment challenging. M. abscessus isolates are typically lysogens, meaning their genomes carry one or more prophages, integrated viral genomes, that have the potential to regulate intrinsic antibiotic resistance. The mechanism by which prophage alter gene expression and antibiotic resistance in mycobacteria is not yet understood and is the focus of this grant. Expression of the transcriptional regulator whiB7 increases in response to stresses, such as sub- inhibitory concentrations of antibiotics and the intracellular environment of macrophages and plays a critical role in mycobacterial intrinsic antibiotic resistance and survival in macrophages (8). Our lab showed for the first time that prophage contribute to intrinsic antibiotic resistance and increased expression of whiB7, which in turn positively regulates a large set of intrinsic antibiotic resistance genes (8-10). We identified a novel group of prophages found in the genomes of M. abscessus and M. chelonae that belong to cluster MabR (7, 11) and determined that the MabR prophage, McProf, increases M. chelonae resistance to aminoglycosides, cornerstone antibiotic treatments for M. abscessus/chelonae infections (9). We found that mycobacteria carrying McProf have an enhanced whiB7 response particularly when superinfected by a second phage. It is not understood how the prophage McProf interacts with the second prophage to drive changes in whiB7 expression and intrinsic resistance. We hypothesize that genes expressed from the McProf genome detect stress and regulate antibiotic resistance in mycobacteria through the induction of whiB7. To understand the mechanisms of whiB7 regulation in mycobacterial lysogens, we propose characterizing the cellular stresses that enhance the whiB7 response in bacterial lysogens carrying the McProf prophage and identify and characterize McProf genes that upregulate whiB7 in response to stress.