Small RNAs of Bartonella bacilliformis; the agent of Carrion's disease in humans

杆状巴尔通体的小RNA；

基本信息

批准号：
9227738
负责人：
Michael F Minnick
金额：
$ 22.7万
依托单位：
UNIVERSITY OF MONTANA
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：
2016-12-01 至 2018-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9227738
关键词：
Acute Address Altitude Andean Area Arthropod Vectors Arthropods Bacteremia Bacteria Bartonella Bartonella Infections Bartonella bacilliformis Biological Process Biology Bite Blood Cells Chemistry Childhood Chronic Colombia Complex Complication Contracts Cues Disease Disease Outbreaks EMSA Ecuador Endothelial Cells Environment Erythrocytes Etiology Exposure to Fatality rate Fever Genes Genetic Transcription Goals Gram-Negative Bacteria Growth Hematocrit procedure Hemolytic Anemia Human Icterus Incidence Infection Insect Vectors Intervention Left Lesion Life Light Lutzomyia genus Mediating Messenger RNA Metabolic Pathway Metabolism Midgut Modeling Molecular Myalgia Natural History Organism Oroya Fever Parasites Pathogenesis Pathogenicity Persons Peru Phagocytosis Phase Play Population Populations at Risk Post-Transcriptional Regulation Prevalence Proteins Reactive Oxygen Species Regulation Reporting Research Risk Role Sand Flies Skin Small RNA South America Surface Symptoms Syndrome Temperature Transcript Translations Untranslated RNA Vascular Endothelial Cell Verruga Peruana Virulence Virulent antimicrobial axenic culture base biological adaptation to stress climate change cost differential expression environmental change extracellular genetic manipulation genetic strain high risk mutant neglected tropical diseases overexpression pathogen pathogenic bacteria response secondary infection skin lesion transcriptome transcriptome sequencing transcriptomics tumor vector

项目摘要

Bartonella bacilliformis (Bb) is a highly virulent, sand fly-borne, Gram-negative bacterium that causes Carrión’s disease in humans. Carrión’s disease is emerging in endemic regions of the Andes and expanding into historically non-endemic areas of Ecuador, Colombia and Peru. The at-risk population is currently ~1.7 million people in a 56,000 square-mile area. Incidence rates of 12.7/100 person years have been reported in endemic regions. Bb infections can be life-threatening, with fatality rates of 40-88%, if left untreated. Pediatric populations are especially at high-risk. In non-endemic regions, the disease often manifests as an acute illness with an ~80% reduction in erythrocyte hematocrit (Oroya fever), whereas in endemic regions, angiomatous skin lesions (verruga peruana) and chronic bacteremia prevail, effectively creating a human reservoir. Little is known about Bb’s molecular pathogenesis and relationship with its phlebotomine sand fly vector, Lutzomyia verrucarum. We hypothesize that small, noncoding RNAs (sRNAs) play key roles in optimizing Bb’s ability to survive and replicate in human cells and sand flies via transcriptional and post-transcriptional regulation. Bb’s natural history offers an exceptional model to examine regulatory roles of sRNAs, as the pathogen is frequently subjected to dramatic shifts in temperature, pH, and disparate chemistries of the extracellular and intracellular niches of the sand fly vector and human host, respectively. Our long-term goal is to elucidate how Bb regulates the adaptive responses necessary for infection, with overarching objectives of: a) generating information that can be applied towards the eventual eradication of Carrión's disease and b) providing a model of sRNA-mediated regulation of survival genes (e.g., stress response, metabolism, virulence, etc.) in Bartonella and other arthropod-borne, intracellular pathogenic bacteria. The hypothesis will be addressed by three specific aims. In aim 1, we will characterize Bb’s baseline transcriptome (sRNAs and mRNAs) during axenic growth using RNA-Seq. Differential transcription as a function of relevant environmental cues will also be explored. Second, we will compare transcriptomes of Bb grown axenically to those infecting human vascular endothelial cells and erythrocytes. These results will allow us to identify infection-specific transcripts produced in the intracellular niche. In aim 2, we will characterize the transcriptome expressed during infection of sand flies. From these results, we can identify infection-specific RNAs produced during extracellular growth in the insect vector. In aim 3, we will analyze functions of the two dominant, infection-specific sRNAs identified above; one produced in host cells and a second in sand flies. First, we will generate respective sRNA mutant and overexpression strains by genetic manipulation. Second, strains will be analyzed for growth and survival during infection, and their transcriptomes analyzed to identify potential targets. The mRNA targets of the dominant sRNAs will be verified by EMSA and the results used to formulate the respective sRNA-mediated regulatory networks. These results will define the functions for two dominant, “infection-specific” sRNAs of Bb.

Bartonella bacilliformis（BB）是一种高毒，沙蝇，革兰氏阴性细菌，会导致Carrión 人类疾病。 Carrión的疾病正在安第斯山脉的地方性地区出现，并扩展到历史上，厄瓜多尔，哥伦比亚和秘鲁的非内态地区。高危人口目前约170万在56,000平方英里的地区。报告的发病率是12.7/100人年的内部分子。地区。如果未治疗，BB感染可能会威胁生命，死亡率为40-88％。小儿人口尤其是高风险。在非末端地区，该疾病通常表现为急性疾病红细胞血细胞比容降低约80％皮肤病变（Verruga Peruana）和慢性细菌占上风，有效地产生了人类储层。几乎没有关于BB的分子发病机理以及与其流血化的砂飞载体的关系，Lutzomyia Verrucarum。我们假设小型的非编码RNA（SRNA）在优化BB的能力方面发挥了关键作用通过转录和转录后调节在人类细胞和沙蝇中生存和复制。 BB的自然历史提供了一个特殊的模型来检查SRNA的调节作用，因为病原体经常是在细胞外和细胞内的温度，pH和不同的化学体中发生急剧变化沙蝇载体和人类寄主的壁ni。我们的长期目标是阐明BB 调节感染所需的自适应反应，并具有：a）生成的总体目标可以应用于消除Carrión病的事件和b）提供模型的信息 Bartonella中SRNA介导的生存基因的调节（例如，应激反应，代谢，病毒等）和其他节肢动物传播的细胞内致病细菌。该假设将由三个具体目标。在AIM 1中，我们将在Axenic期间表征BB的基线转录组（SRNA和mRNA）使用RNA-Seq的生长。差分转录作为相关环境提示的函数也将是探索。其次，我们将在轴承中比较BB的转录组与感染的人血管的转录组内皮细胞和红细胞。这些结果将使我们能够识别产生的特定感染的成绩单在AIM 2中，我们将表征在砂过程中表达的转录组苍蝇。从这些结果中，我们可以确定在细胞外生长过程中产生的特异性RNA 在AIM 3中，我们将分析确定的两个主要感染特异性SRNA的功能多于;一个在宿主细胞中产生，第二秒在沙蝇中产生。首先，我们将产生相对的SRNA突变体和通过基因操纵而过表达的菌株。其次，将分析菌株的生长和生存在感染期间及其转录组进行了分析，以鉴定潜在靶标。 mRNA目标 EMSA将验证主要的SRNA，并用于制定相应的SRNA介导的结果监管网络。这些结果将定义BB的两个主要“感染特异性” SRNA的功能。