Genome scanning for virulence genes in bacteria

描述（由申请人提供）：在过去的五十年里，人类细菌感染的控制和治疗在很大程度上依赖于抗生素的使用。然而，抗生素耐药菌的发病率不断增加，需要开发新的抗生素。对细菌基因的全面了解对于体外生长和体内存活至关重要，这对于确定开发有效抗生素的有希望的靶点至关重要。本项目的目标是利用S. enterica serovar Enterilis（S.作为模式生物。S.沙门氏菌是沙门氏菌中最常见的血清型之一，在美国引起食源性疾病，通过常规污染鸡壳蛋而对公共安全构成独特的关注。我们最近开发了一种称为“转座子序列标签分析”（TSP）的方法，用于在复杂的突变体库中对转座子插入进行定量编目。通过筛选约500个S的库，证明了TSP在细菌基因组功能表征中的可行性。在鸡感染过程中用于减毒的抗肿瘤Tn5突变体为实现这一目标，提出了两个具体目标。我们将使用基于TSP的实验方法来进行全基因组鉴定，这些基因在Specific Aim 1中不适合体外生长，而在Specific Aim 2中小鼠感染期间体内存活所需的毒力基因。对S.在本项目中确定的病原体应提供对这种重要食源性病原体体外和体内存活机制的见解。这些实验方法应该找到广泛的应用到其他不同的细菌物种，这将有助于制定有效的措施，以控制细菌病原体。

DESCRIPTION (provided by applicant): The control and treatment of bacterial infections in humans have been largely dependent on the use of antibiotics for the last five decades. However, the increasing incidence of antibiotic resistant bacteria demands the development of novel antibiotics. Comprehensive knowledge of the bacterial genes that are essential for in vitro growth and in vivo survival is crucial to identify promising targets for the development of effective antibiotics. The goal of this project is to develop experimental approaches for genome-wide identification of bacterial genes that are essential for in vitro growth or in vivo survival using S. enterica serovar Enteritidis (S. enteritidis) as a model organism. S. enteritidis is one of the most common serotypes of Salmonella causing food borne illness in the United States, posing a unique concern for public safety by routinely contaminating chicken shell eggs. We recently developed a method, termed "transposon sequence tag profiling" (TSP), for quantitative cataloguing of transposon insertions in a complex pool of mutants. The feasibility of TSP in functional characterization of a bacterial genome was demonstrated by screening a pool of approximately 500 S. enteritidis Tn5 mutants for attenuation during chicken infection. Two specific aims are proposed to accomplish the goal. We will use experimental approaches based on TSP to conduct genome-wide identification of the essential genes that are not dispensable for in vitro growth in Specific Aim 1 and the virulence genes that are required for in vivo survival during mouse infection in Specific Aim 2. The essential genes and the virulence genes in S. enteritidis that will be identified in this project should provide insights into the mechanisms of both in vitro and in vivo survival of this important food borne pathogen. These experimental approaches should find broad applications to other diverse bacterial species, which will contribute to the development of effective measures to control bacterial pathogens.