Intracellular persistence factors in urinary tract infections

尿路感染的细胞内持久性因素

• 批准号: 535296762
• 负责人: Professor Dr. Christoph Ernst
• 金额: --
• 依托单位: Institut für Medizinische Mikrobiologie, Immunologie und Hygiene
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/535296762?language=en
• 关键词: Intracellular; persistence; factors; urinary; tract

Persistent infections are often caused by bacterial pathogens that display in vitro antibiotic susceptibility, illustrating the intrinsic limitations of antibiotic therapy in the complex environment of the host. While variable patient factors contribute to antibiotic efficacy, fundamental principles of antibiotic persistence have been established. In vitro, antibiotic persistence is observed under ideal growth conditions in dormant subpopulations that tolerate antibiotics due to inactivity of antibiotic targets. In vivo, intracellular infection stages provide protection against many components of the immune system, while host pathogen interactions can promote antibiotic persistence. In my postdoctoral studies, my colleagues and I studied the adaptive evolution of multidrug- resistant Klebsiella pneumoniae and discovered global evolution of persistence in urinary tract infections, which involved an intrinsic ability of K. pneumoniae to persist in LAMP1 positive vacuoles of bladder epithelial cells and tolerate lethal concentrations of antibiotics. A follow-up investigation of intracellular antibiotic persistence in K. pneumoniae found that dormant, multidrug-tolerant bacteria occurred at higher frequency in the intracellular environment compared to the extracellular environment, in line with an increasing body of evidence linking host pathogen interactions to antibiotic tolerance. To target intracellular bacterial infections, we developed a high-throughput screening assay of intracellular survival and conducted a compound screen that led to the discovery of a host-targeting compound that induces intracellular antimicrobial activity in bladder cells and displays in vivo efficacy in a mouse model of latent UTI. I propose to further investigate the intrinsic ability of K. pneumoniae to persist intracellularly in LAMP1 positive vacuoles of bladder epithelial cells to gain a better understanding of the molecular basis of persistent urinary tract infections. Intracellular persistence factors will be identified in an arrayed transposon mutant library. Validation experiments and phenotypic characterization of genes with the strongest impact on intracellular survival will lead to a more detailed mechanistic investigation that focuses on one of the identified persistence factors. Protein interaction experiments will help to elucidate a pathway of intracellular persistence which could lead to the discovery of host pathogen interactions that promote intracellular survival. Moreover, the impact of a mutation that disrupts intracellular survival will be investigated in a mouse model of urinary tract infection which will provide an opportunity to study the role of intracellular infection stages in causing persistent infections.

持续性感染通常由在体外显示抗生素敏感性的细菌病原体引起，这说明了抗生素治疗在宿主复杂环境中的内在局限性。虽然可变的患者因素有助于抗生素疗效，但抗生素持久性的基本原则已经确立。在体外，在理想的生长条件下，由于抗生素靶标的失活，在耐受抗生素的休眠亚群中观察到抗生素持久性。在体内，细胞内感染阶段提供针对免疫系统的许多组分的保护，而宿主病原体相互作用可促进抗生素持久性。在我的博士后研究中，我和我的同事研究了多重耐药肺炎克雷伯菌的适应性进化，并发现了尿路感染持续性的全球进化，这涉及到K的内在能力。pneumoniae）在膀胱上皮细胞的LAMP1阳性空泡中持续存在，并耐受致死浓度的抗生素。对克雷伯氏菌胞内抗生素持久性的跟踪研究。pneumoniae发现，与细胞外环境相比，休眠的多药耐受性细菌在细胞内环境中以更高的频率出现，这与越来越多的将宿主病原体相互作用与抗生素耐受性联系起来的证据一致。为了靶向细胞内细菌感染，我们开发了一种细胞内存活的高通量筛选测定，并进行了化合物筛选，发现了一种宿主靶向化合物，该化合物在膀胱细胞中诱导细胞内抗微生物活性，并在潜伏性UTI小鼠模型中显示体内疗效。我建议进一步研究K的内在能力。本发明的目的在于使肺炎链球菌在膀胱上皮细胞的LAMP1阳性空泡中持续存在于细胞内，以更好地理解持续性尿路感染的分子基础。将在阵列转座子突变体文库中鉴定细胞内持久性因子。验证实验和细胞内生存的影响最大的基因的表型表征将导致一个更详细的机制调查，重点是确定的持久性因素之一。蛋白质相互作用实验将有助于阐明细胞内持久性的途径，这可能导致发现宿主病原体相互作用，促进细胞内的生存。此外，将在尿路感染的小鼠模型中研究破坏细胞内存活的突变的影响，这将提供研究细胞内感染阶段在引起持续性感染中的作用的机会。