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Vibrio cholerae-chitin interactions and their role in cholera transmission and evolution

霍乱弧菌-几丁质相互作用及其在霍乱传播和进化中的作用

基本信息

批准号：
9094029
负责人：
Ankur Dalia
金额：
$ 16.1万
依托单位：
TRUSTEES OF INDIANA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-05-13 至 2018-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9094029
关键词：
Address Affect Architecture Area Bacteria Biological Assay Biology Carbohydrates Carbon Catabolism Cells Cessation of life Chitin Chitinase Chitosan Cholera Clinical Competence Crustacea Cyclic AMP Receptor Protein DNA Data Disease Disease Outbreaks Environment Evolution Exhibits Fluorescence Fluorescence Microscopy Fluorescence-Activated Cell Sorting Food Contamination Gene Expression Profile Gene Mutation Genes Genetic Determinism Genetic Screening Genome Glucosamine Goals Growth Heterogeneity Horizontal Gene Transfer Human Infection Laboratories Life Cycle Stages Link Metabolism Methods Microbe Microbial Biofilms Modeling Molecular Mutagenesis Nature Nitrogen Nutrient Oligosaccharides Outcome Pathway interactions Pattern Phenotype Physiological Physiology Polymers Polysaccharides Population Population Heterogeneity Prevention strategy Probiotics Process Regulation Reporter Reporter Genes Research Research Project Grants Role Signal Transduction Source Surface System Testing Therapeutic Intervention Vibrio Vibrio cholerae Virulence Virulence Factors Work Zooplankton antimicrobial base cell transformation combat contaminated water drinking water extracellular feeding fitness genetic approach homologous recombination microbial microorganism mutant novel novel strategies pathogen prevent public health relevance research study response sensor histidine kinase transcriptome transcriptome sequencing transmission process uptake waterborne

项目摘要

DESCRIPTION (provided by applicant): To successfully survive and transmit to their host, facultative pathogenic microorganisms must acquire nutrients in the diverse niches they occupy. Additionally, these nutrients can serve as signaling compounds to alter the physiology of these microbes. Thus, understanding the nutrients used and sensed by pathogenic microbes throughout their life cycle can be exploited for developing novel probiotics, antimicrobials, and preventative strategies. The research proposed here will uncover the molecular mechanisms underlying the interaction of Vibrio cholerae with the metabolite chitin and their role in the survival and evolution of this pathogen in its environmental reservoir and transmission to its human host. Vibrio cholerae is the causative agent of the diarrheal disease cholera and is annually responsible for 3-5 million infections and >100,000 deaths worldwide. This facultative pathogen is a common resident of the aquatic environment and causes disease if ingested in contaminated food or water. Much about the metabolism of this pathogen in the aquatic environment and the human host, however, remain unclear. A major carbon and nitrogen source for Vibrio species in the aquatic environment is the polysaccharide chitin, which is the primary constituent of the shells of crustacean zooplankton. Chitin is a polymer composed of β1,4- linked N-acetyl glucosamine (GlcNAc) residues, and is the second-most abundant polysaccharide in nature with >1011 tons being produced annually in the aquatic environment. Cholera infections are seasonal in endemic areas and are closely associated with seasonal blooms of zooplankton. In aquatic microcosm experiments in the laboratory, it has been shown that chitin enhances V. cholerae growth and biofilm formation and, therefore, likely enhances the waterborne transmission of this pathogen. Many studies have demonstrated the importance of biofilm formation on the virulence of V. cholerae using bacteria grown in rich media. Relatively few studies, however, have assessed the virulence of biofilms grown on chitin, the most physiologically relevant nutrient for this pathogen in the aquatic environment. In addition to its role as a nutrient, chitin also induces a physiological state in V. cholerae known as natural competence, where bacteria can take up DNA from the extracellular environment. This DNA can then be catabolized or integrated into the genome by homologous recombination. This latter process is known as natural transformation and is shared by diverse microbial species. Evolutionarily, natural transformation provides microorganisms a mechanism for acquiring genes and mutations that enhance their fitness. In this manner, seasonal blooms of zooplankton may promote rapid evolution of V. cholerae in endemic areas. The goals of this research project are to characterize the mechanisms and outcomes of V. cholerae-chitin interactions and their roles in the waterborne transmission and evolution of this pathogen. We have developed three aims to address these goals. In Aim 1, we propose to use unbiased high-throughput genetic screens (Tn-seq), whole transcriptome analysis (RNA-seq), and cutting-edge genetic approaches to uncover novel factors required for the formation of chitin biofilms and their corresponding regulation. In preliminary work we have characterized the genes required for uptake of chitin degradation products (chitin oligosaccharides, GlcNAc, and chitosan oligosaccharides). Thus, in Aim 2, we propose to use defined mutant strains and gene reporter fusions to assess how the uptake of distinct chitin degradation products affects the spatial architecture of a chitin biofilm and the virulence of chitin-grown V. cholerae. In Aim 3, we propose to assess the mechanisms underlying chitin-induced natural transformation in V. cholerae. Specifically, we have previously shown that in diverse naturally competent microbial species, there is phenotypic heterogeneity among competent bacteria during natural transformation. To address the mechanisms underlying this heterogeneity and their role in this conserved and critical evolutionary process we propose to use fluorescence-activated cell sorting (FACS), RNA-seq, fluorescent gene reporters, and complementary molecular methods to identify genes and pathways that correlate with successful natural transformation. These aims will provide a deeper understanding of how V. cholerae degrades, utilizes, and responds to chitin in the aquatic environment, and may uncover novel targets and strategies to prevent seasonal outbreaks of cholera in endemic areas.