Community behavior of Yersinia pseudotuberculosis within microcolonies

小菌落内假结核耶尔森菌的群落行为

• 批准号: 9088649
• 负责人: Kim Davis
• 金额: $ 16.2万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9088649
• 关键词: Antibiotic Resistance; Antibiotics; Antimicrobial susceptibility; Automobile Driving; Bacteria; Bacterial Genes; Bacterial Infections; Behavior; Cells; Communities; Development; Devices; Diffuse; Disease; Disease Progression; Distal; Environment; Flow Cytometry; Food Contamination; Gene Expression; Gene Expression Regulation; Generations; Genes; Genetic Transcription; Goals; Growth; Heterogeneity; Image; Immune response; Individual; Infection; Inflammatory; Ingestion; Intestines; Lead; Link; Medical; Metabolic; Metabolism; Microfluidic Microchips; Microfluidics; Modeling; Nitric Oxide; Oral; Pasteurella pseudotuberculosis; Pathway interactions; Peripheral; Population; Population Dynamics; Population Heterogeneity; Predisposition; Process; Proteins; Recruitment Activity; Reporter; Research; Role; Signal Transduction; Site; Stress; Systemic infection; Testing; Therapeutic; Time; Tissues; Treatment outcome; Type III Secretion System Pathway; Variant; Virulence; Virulence Factors; antimicrobial; base; biological adaptation to stress; constriction; design; enteric pathogen; extracellular; feeding; fluorescence imaging; in vivo; lymph nodes; macrophage; member; microorganism; mouse model; neutrophil; novel; novel therapeutics; pathogen; programs; public health relevance; rapid growth; response; transcriptome sequencing

 DESCRIPTION (provided by applicant): It has been known for decades that bacterial populations are heterogeneous, however, it has been extremely difficult to develop therapeutics that effectively eliminate all members of bacterial populations. Much of this difficulty lies in th fact that individual bacteria are replicating at different rates within a population, and this lead to differences in antimicrobial susceptibility. In remains unclear what drives differences in bacteria growth rates during infection, and whether the host immune response can slow the growth of subpopulations of bacteria. Recent advances in fluorescence imaging have enabled the study of heterogeneity within bacterial populations at the single cell level. Using fluorescent transcriptional reporters, we have shown that Y. pseudotuberculosis forms multiple subpopulations within a single site of bacterial replication, based on the response of individual cells to host-derived stresses. The proximity of individual bacteria to host cells drove gene expression changes, resulting in a small population of bacteria around the periphery of replicating clusters (called microcolonies) that specifically responds to host-derived stress. The impact of host stress on individual bacteria remains unclear; does this lead to the generation of a slow growing population on the periphery of microcolonies? The studies described in this proposal will utilize fluorescent reporter constructs to visualize bacterial responses to host stress over the course of infection, and determine the effects of host-derived stresses on bacterial growth rates. We hypothesize that individual bacteria around the periphery of microcolonies simultaneously respond to multiple host stresses, which results in a slower rate of bacterial replication within this subpopulation. To test this hypothesis, we will: 1) utilize stable and unstable fluorescent proteins to determine whether the stresses imparted by the host are transient, or maintained over the course of infection; 2) develop an oral feeding model of Y. pseudotuberculosis intestinal infection, to determine if spatial regulation of gene expression within intestinal tissues impacts the ability of bacteria to disseminate to deep tissues; and 3) develop a microfluidics model to live image bacterial replication during stress responses, to determine what happens to individual bacteria as they respond to single stresses or multiple stresses simultaneously. Although these studies will focus on Y. pseudotuberculosis growth, the models developed within this proposal can easily be applied to further the study of other enteric pathogens, and additional medically-relevant bacterial pathogens, to better understand the mechanisms driving the formation of slow growing bacterial populations. By gaining a better understanding of this process, we hope to provide pertinent information for the development of novel therapeutics to eliminate all members of bacterial populations, and thus clear infection.

 描述（由适用提供）：几十年来，细菌种群是异质的，但是，开发有效消除所有细菌种群的理论非常困难。这些困难的大部分在于，个体细菌在人群中以不同的速度复制，这导致抗菌敏感性差异。仍然不清楚是什么驱动了感染过程中细菌生长率差异的差异，以及宿主免疫增强响应是否会减慢细菌亚群的生长。荧光成像的最新进展已使单个细胞水平细菌种群中的异质性研究。使用荧光转录记者，我们表明Y.假结核病在细菌复制的单个部位内形成了多个亚群，这是基于单个细胞对宿主衍生应力的响应的反应。单个细菌与宿主细胞的接近性驱动基因表达变化，从而导致在复制簇（称为微卵石）周围周围的细菌群，这些细菌特异性地响应了宿主衍生的胁迫。宿主压力对单个细菌的影响尚不清楚。这是否导致微菌落周围的人群增长缓慢？该提案中描述的研究将利用荧光报告基因构建体来可视化细菌反应，以在感染过程中对宿主压力的反应，并确定宿主衍生的压力对细菌生长速率的影响。 我们假设微菌落周围的单个细菌只是对多种宿主应力做出反应，从而导致该亚群中细菌复制速度较慢。为了检验这一假设，我们将：1）利用稳定且不稳定的荧光蛋白来确定宿主施加的应力是短暂的还是在感染过程中保持的； 2）开发Y. pseudotuberculosis肠道感染的口服喂养模型，以确定肠道组织中基因表达的空间调节是否会影响细菌散发到深层组织的能力； 3）开发一个微流体模型，以在应力反应过程中实现细菌复制，以确定单个细菌对单个应力或多种应力的反应时会发生什么。尽管这些研究将集中在Y.假结核病的生长上，但在该提案中开发的模型可以轻松地用于进一步研究其他肠道病原体，以及其他与医学上相关的细菌病原体，以更好地了解促进生长慢细菌种群的形成的机制。通过更好地了解这一过程，我们希望为开发新疗法提供相关信息，以消除所有细菌种群的成员，从而清除感染。