Determination of the Interactome between Haemophilus ducreyi and the Human Host.

我们对传染病认识上的一个主要差距是缺乏关于分子相互作用的信息。 在感染病原体和人类宿主之间的网络，这对于细菌来说还没有完成 人类的感染。此应用程序的目的是在文本级别上为 利用RNA在实验感染的人体志愿者中发现主要皮肤病原体杜氏嗜血杆菌(HD) 测序(RNA-seq)和代谢组学。先天性巨结肠症引起下巴，这是一种生殖器溃疡(GU)疾病，容易 艾滋病毒传播，是雅司病流行国家儿童皮肤溃疡(CU)的主要原因。 由于环境水库的原因，使用抗生素根除与HD相关的CU的努力失败了。为了研究 HD生物学，我们建立了一种接种HD gu株35000hp及其衍生物的模型 进入成年志愿者的上臂皮肤。全基因组测序显示大多数铜绿假单胞菌 与35000HP几乎相同，表明我们的模型与GU和CU高度相关。在模型中和在 天然下巴状，HD位于脓肿中，周围有多形核白细胞和 巨噬细胞，并通过逃避吞噬作用留在细胞外。在我们新的初步数据中，我们显示 在HD和寄主以及差异寄主和细菌转录本之间存在相互作用网络 表达与感染部位和受伤部位的代谢变化相关。房屋署主要上调 适应厌氧和金属离子吸收利用相关基因的表达 替代碳源，如抗坏血酸，与“营养毒性”的概念一致。我们的新产品 代谢组学数据显示，抗坏血酸途径在皮损中上调。因此，我们假设 宿主调节其基因转录以吞噬和限制营养物质进入HD，HD调节其 基因转录，以抵消这些宿主防御，宿主基因表达与 受感染部位的代谢组谱，以及参与适应 由宿主反应创造的代谢生态位将是毒力所必需的。我们的具体目标是1) 确定受感染组织中HD和人类宿主之间的代谢组和相互作用组 将宿主转录反应与病变中的代谢变化相关联；2)确定负责的细胞 对于宿主的转录反应采用单细胞rna-seq；3)确定HD基因是否 参与开发宿主生态位是人类毒力所必需的，以及其机制(S) 这些基因对毒力有贡献。这项研究的重要性在于，我们将第一个确定 人类细菌感染部位的相互作用组，定义负责转录的宿主细胞 响应，将宿主响应与病变中的代谢变化相关联，并研究HD如何利用这些 代谢物。我们将提供一个新的洞察力来研究细胞外细菌模型和 人类宿主和揭开面纱的新策略，以控制与HD相关的CU。

A major gap in our understanding of infectious diseases is the lack of information about molecular interaction networks between an infecting pathogen and the human host, which has yet to be accomplished for a bacterial infection in humans. The purpose of this application is to define an interactome on a transcript level for the primary skin pathogen, Haemophilus ducreyi (HD), in experimentally infected human volunteers using RNA sequencing (RNA-seq) and metabolomics. HD causes chancroid, a genital ulcer (GU) disease that facilitates HIV transmission and is as a major cause of cutaneous ulcers (CU) in children in yaws-endemic countries. Efforts to eradicate HD-associated CU with antibiotics failed due to environmental reservoirs. To study the biology of HD, we developed a model in which the HD GU strain 35000HP and its derivatives are inoculated into the skin of the upper arm of adult volunteers. Whole genome sequencing shows that most CU strains are nearly identical to 35000HP, indicating that our model is highly relevant to GU and CU. In the model and in natural chancroid, HD resides in an abscess where it is surrounded by polymorphonuclear leukocytes and macrophages and remains extracellular by evading phagocytosis. In our new preliminary data, we show that an interaction network exists between HD and the host and that differential host and bacterial transcript expression correlates with metabolomic changes at infected vs. wounded sites. HD primarily upregulates the expression of genes involved in adapting to anaerobiosis and uptake and utilization of metal ions and alternative carbon sources, such as ascorbic acid, consistent with the idea of “nutritional virulence.” Our new metabolomics data show that ascorbic acid pathways are upregulated in lesions. Thus, we hypothesize that the host regulates its gene transcription to phagocytize and limit nutrients to HD, that HD regulates its gene transcription to counteract these host defenses, that host gene expression correlates with metabolomic profiles at infected sites, and that bacterial genes that are involved in adaptation to the metabolic niche created by the host response will be required for virulence. Our specific aims are 1) to define the metabolome and the interactome between HD and the human host in infected tissue and to correlate the host transcriptional response to metabolic changes in lesions; 2) to identify the cells responsible for the host transcriptional response using single cell RNA-seq; 3) to determine whether the HD genes involved in exploitation of the host niche are required for virulence in humans and the mechanism(s) by which these genes contribute to virulence. The importance of this study is that we will be the first to determine an interactome at a site of a bacterial human infection, define the host cells responsible for the transcriptional response, correlate the host response to metabolomic changes in lesions, and study how HD exploits these metabolites. We will provide new insights into the interaction between a model extracellular bacterium and the human host and unmask novel strategies to control HD-associated CU.