Mechanisms of Klebsiella pneumoniae gastrointestinal colonization

肺炎克雷伯菌胃肠道定植机制

• 批准号: 10736879
• 负责人: Muhammad Ammar Zafar
• 金额: $ 55.83万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-14 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10736879
• 关键词: Affect; Animal Model; Antibiotic Resistance; Arginine; Biological; Biological Assay; Bypass; Catabolism; Cell membrane; Cellular Membrane; Data; Development; Diet; Disease; Ethanolamines; Failure; Feces; Gastrointestinal tract structure; Genes; Genetic Screening; Gnotobiotic; Gram-Negative Bacteria; Immune system; Immunocompetent; In Vitro; Infection; Inflammatory; Investigation; Kinetics; Klebsiella pneumoniae; Knock-out; Knowledge; Libraries; Link; Metabolic Pathway; Metabolism; Metagenomics; Modeling; Molecular; Mucous Membrane; Multi-Drug Resistance; Mus; Nosocomial Infections; Nutrient; Nutritional; Operon; Oral; Oropharyngeal; Outcome; Pathway interactions; Patients; Prevention; Process; Regulation; Role; Route; Site; Source; System; burden of illness; colonization resistance; combat; diarrheal disease; digital; drug resistant pathogen; enteric pathogen; epidemiology study; fitness; gastrointestinal; gene product; genomic locus; gut colonization; gut microbiome; gut microbiota; host colonization; in vivo; insight; member; metabolomics; metagenome; metagenomic sequencing; microbiome; microbiome components; microbiota; microorganism; mouse model; mutant; novel; pathogen; pathogenic bacteria; public health relevance; success; transmission process; uptake

Abstract Hospital-acquired infections (HAI) resulting from the transmission of drug-resistant pathogens affect hundreds of millions of patients worldwide. Klebsiella pneumoniae (Kpn), a gram-negative bacterium, is notorious for causing HAI, with many of these infections difficult to treat as Kpn has become multi-drug resistant. Epidemiological studies suggest that gastrointestinal (GI) colonization of Kpn is a major reservoir through which Kpn can cause disease manifestations either in the colonized host or transmit from host to host. This site of Kpn colonization has not been the focus of previous studies as a tractable model of Kpn GI colonization, and host-to-host transmission did not exist. We have recently developed a murine model that allows for the study of Kpn mucosal (oropharynx and GI) colonization, shedding within feces, and transmission through the fecal-oral route. Using an oral route of inoculation and fecal shedding as a marker for GI colonization, we show that Kpn can asymptomatically colonize the GI tract of immunocompetent mice and modifies the host GI microbiota. We premise that specific Kpn genes contribute to its GI colonization, and the products of these genes could serve as novel targets for the prevention of the establishment of GI colonization. More recently, we used our murine model to screen a library of Kpn random transposon mutants (In-seq) to identify the complete set of “GI colonization” genes from a single isolate. A metagenomics sequencing analysis further identified bacterial species and the metabolic pathways affected by Kpn in the GI tract. Herein, we will focus on two sets of pathways identified through In-seq whose products allow Kpn to overcome colonization resistance provided by the resident gut microbiota. Thus, in Aim#1, we will focus on the ethanolamine utilization pathway genes (eut) of Kpn that allow it to utilize ethanolamine (EA), a byproduct of cellular membranes and diet in the gut that can serve as an alternative nutrient source. Unlike many other enteric pathogens that contain a single eut operon, Kpn has two genetically distinct eut operons. We will identify the role of each eut locus in EA metabolism and determine the underlying molecular mechanism through which EA metabolism provides Kpn with a fitness advantage against members of the microbiome. Aim#2 will take a different approach by focusing on the contact-dependent killing machinery of the Kpn (Type 6 secretion system [T6SS]) in overcoming colonization resistance provided by the resident microbiota. We will focus on the unique regulatory mechanism that modulates the expression of Kpn T6SS in the GI tract and provide it with a selective and competitive advantage against the resident gut microbiota. Results from these studies would provide us with a fundamental understanding of the molecular mechanisms involved in the establishment of GI colonization by an incoming pathogen. These studies will also lay the groundwork for developing potential strategies to reduce the Kpn disease burden.

抽象的 耐药病原体传播导致的医院获得性感染 (HAI) 影响数百人 肺炎克雷伯菌 (Kpn) 是一种革兰氏阴性细菌，因感染而臭名昭著。 导致 HAI，其中许多感染难以治疗，因为 Kpn 已产生多重耐药性。 流行病学研究表明，Kpn 的胃肠道 (GI) 定植是主要的储存库 Kpn 可以在定植宿主中引起疾病​​表现，或者在宿主之间传播。 作为 Kpn GI 定植的易处理模型，Kpn 定植的研究并不是以前研究的焦点，并且 我们最近开发了一种可以进行研究的小鼠模型。 Kpn 粘膜（口咽和胃肠道）定植、粪便内脱落以及通过粪口传播 我们使用口服途径接种和粪便排出作为胃肠道定植的标志。 Kpn 可以无症状地定植于免疫活性小鼠的胃肠道并改变宿主胃肠道 我们假设特定的 Kpn 基因有助于其胃肠道定植，以及这些基因的产物。 最近，我们使用了可以作为预防胃肠道定植的新目标。 我们的小鼠模型用于筛选 Kpn 随机转座子突变体库 (In-seq)，以识别完整的一组 来自单个分离株的“胃肠道定植”基因进一步鉴定出细菌。 胃肠道中受 Kpn 影响的物种和代谢途径在此，我们将重点关注两组途径。 通过 In-seq 鉴定，其产品使 Kpn 能够克服居民提供的定植抵抗力 因此，在目标#1 中，我们将重点关注 Kpn 的乙醇胺利用途径基因 (eut)。 让它利用乙醇胺 (EA)，乙醇胺是肠道内细胞膜和饮食的副产品，可以作为 与许多其他含有单一 eut 操纵子的肠道病原体不同，Kpn 有两个。 我们将鉴定每个 eut 位点在 EA 代谢中的作用并确定 EA 代谢通过其潜在分子机制为 Kpn 提供健身优势 Aim#2 的成员将采取不同的方法，重点关注接触依赖性杀伤。 Kpn（6 型分泌系统 [T6SS]）的机制克服了由 我们将重点关注调节表达的独特调节机制。 Kpn T6SS 在胃肠道中的表达，并为其提供针对常驻肠道的选择性和竞争优势 这些研究的结果将为我们提供对分子的基本了解。 这些研究还将涉及传入病原体在胃肠道定植的建立机制。 为制定减少 Kpn 疾病负担的潜在策略奠定基础。