Genome-wide identification of virulence genes in Acinetobacter baumannii in vivo

描述（由申请人提供）：鲍曼不动杆菌是一种革兰氏阴性菌，已成为一种重要的机会性病原体，在美国构成重大的公共卫生风险。在临床上，鲍曼不动杆菌正在成为一种主要的医院病原体，特别是在专门从事呼吸护理、创伤和烧伤的重症监护病房。尽管有三个鲍曼不动杆菌菌株的完整基因组序列，但大多数对哺乳动物宿主发病至关重要的基因的身份尚不清楚。我们打算从近4000个细菌基因中找出哪些是哺乳动物宿主生存所必需的，并在实验诱导的菌血症中证明每个基因的必要性。这一结果将在理解鲍曼不动杆菌感染的病理生物学和制定治疗这些感染的策略或预防其发生的疫苗方面迈出重要一步。我们的长期目标是系统地建立鲍曼不动杆菌感染发病机制的分子模型。此应用程序的目的是识别所有基因

DESCRIPTION (provided by applicant): Acinetobacter baumannii, a Gram-negative bacterium, has emerged as an important opportunistic pathogen that poses a significant public health risk in the United States. Clinically, A. baumannii is emerging as a leading nosocomial pathogen, particularly in intensive care units specializing in respiratory care, trauma and burns. Despite having the complete genome sequences for three strains of A. baumannii, the identity of most genes that are essential for pathogenesis in a mammalian host is not known. We propose to identify which of the nearly 4000 bacterial genes are essential for survival in a mammalian host and to demonstrate the necessity for each gene during experimentally induced bacteremia. The result will represent a major step forward in understanding the pathobiology of A. baumannii infections and for devising strategies to treat these infections or vaccinate against their occurrence. Our long-term goal is to systematically establish a molecular model of pathogenesis for A. baumannii infection. The objective of this application is to identify all genes essential to establish infection within a mammalian host. Based on a successful pilot experiment with 109,000 mutant of A. baumannii ATCC17978 in a murine model of bacteremia, our central hypothesis is that genes of A. baumannii essential for development of bacteremia can be determined by combining global transposon mutagenesis, a murine model of infection, and high throughput sequencing. Our rationale for conducting this work is to establish a framework that will direct detailed mechanistic investigations of A. baumannii pathogenesis in compromised patients. Using a saturating transposon library of A. baumannii mutants, the animal model of bacteremia, and high- throughput DNA sequencing, we will accomplish the following specific aims: 1) identify virulence genes essential for A. baumannii to colonize and disseminate within a mammalian host; and 2) confirm A. baumannii virulence genes that contribute most significantly to colonization and dissemination in the murine model using defined mutations. This contribution will be significant because, in these comprehensive and unbiased experiments, we will determine for the first time the complete A. baumannii virulence gene set for two strains during a mammalian infection. This proposal should be considered innovative as it applies transposon-directed insertion-site sequencing (TraDIS) for genome-wide identification of A. baumannii virulence genes using a forward-genetic screen in a relevant animal model of infection.