Impact of commensal Corynebacterium species on pathogen colonization and microbiota composition

 描述（由申请人提供）：迫切需要进行功能研究，以阐明人体微生物组中不同细菌种属之间的相互作用机制，以了解它们在该微生物群落中的作用。研究表明，非白喉棒状杆菌物种在构建健康的鼻腔和皮肤微生物群方面发挥着重要作用。这些通常无害的抗生素涉及影响金黄色葡萄球菌和肺炎链球菌的定植，这些病原体也经常定植在鼻/皮肤部位。我们的数据表明，一些棒状杆菌物种水解人类皮肤表面脂质的类似物，释放具有抗肺炎球菌活性的游离脂肪酸（FFA）。因此，我们假设棒状杆菌属。通过产生从宿主上皮表面脂质释放具有抗菌活性的FFA的脂肪酶来改变它们的栖息地，并且这些脂质底物的水平影响皮肤微生物群的组成，减少病原体定殖。该提案的总体目标是获得关于棒状杆菌属细菌是如何在大肠杆菌中生长的遗传和机理知识。从宿主表面脂质释放这些抗菌FFA，并确定改变皮肤脂质组成是否影响微生物群组成。这项研究的基本原理是，揭示这些机制将创造机会，以确定特定的菌株，基因和/或棒状杆菌产生的化合物与潜在的治疗用途。我们将使用基于培养的方法和人类样本的组合来识别相关基因，以及它们在培养物和上皮表面上表达的影响。 本建议的具体目的是（1）确定棒状杆菌属如何。从宿主脂质中释放抗菌游离脂肪酸，（2）鉴定和表征控制需要脂质和不依赖脂质的棒状杆菌属中细胞外脂肪酶活性的调节机制。和（3）确定局部施用的皮肤表面脂质类似物对三种不同人类皮肤部位（干性、湿性和油性）的微生物群的影响。为了实现这些目标，我们将使用遗传学、生物化学和生物信息学方法（目标1和2）的组合，沿着基于16 S rRNA基因的方法（目标3）。这项工作的意义将是提供基本的机械信息，这是迫切需要的，以确定具体菌株的棒状杆菌属。以及棒状杆菌产生的化合物，其具有作为替代的、非抗生素疗法的潜力，以预防病原体如S. pneumoniae和S.金黄色。我们策略中的三个主要创新是（1）使用共培养细菌的功能分子研究来阐明不同细菌物种在人类微生物组中的作用，（2）选择一个允许直接在人类宿主中进行非侵入性研究的系统，以及（3）将我们的发现转化为测试人类皮肤表面的简单干预。我们的长期目标是开发新的可持续方法来管理鼻腔和皮肤微生物群的组成，通过排除或控制病原体来预防疾病。

 DESCRIPTION (provided by applicant): Functional studies to elucidate mechanistic interactions between different bacterial species within the human microbiome are critically needed to understand their role in this microbial community. Research suggests that non-diphtheriae Corynebacterium species play a significant role in structuring healthy nasal and skin microbiota. These generally harmless commensals are implicated in affecting colonization by Staphylococcus aureus and Streptococcus pneumoniae, pathogens that also frequently colonize nasal/skin sites. Our data indicate that some Corynebacterium species hydrolyze analogs of human skin-surface lipids releasing free fatty acids (FFAs) with anti-pneumococcal activity. Thus, we hypothesize that Corynebacterium spp. modify their habitat by producing lipases that release FFAs with antibacterial activity from host epithelial-surface lipids and that the level of these lipid substrates impacts the composition of the skin microbiota, diminishing pathogen colonization. The overall objective of this proposal is to gain genetic and mechanistic knowledge of how commensal Corynebacterium spp. release these antibacterial FFAs from host surface lipids and to determine if altering skin lipid composition affects microbiota composition. The rationale for this study is that uncovering these mechanisms will create opportunities to identify specific strains, genes and/or Corynebacterium-produced compounds with potential for therapeutic use. We will use a combination of culture-based approaches and human samples to identify relevant genes, and the impact of their expression, in culture and on epithelial surfaces. The Specific Aims of this proposal are (1) to determine how Corynebacterium spp. release antibacterial free fatty acids from host lipids, (2) to identify and characterize the regulatory mechanisms that control activity of extracellular lipases in lipid-requiring and lipid-independent Corynebacterium spp. and (3) to determine the impact of a topically applied skin-surface-lipid analog on the microbiota of three different human skin sites (dry, moist and oily). To accomplish these, we will use a combination of genetic, biochemical and bioinformatic approaches (Aims 1 and 2), along with a 16S rRNA gene-based approach (Aim 3). The significance of this work will be to provide the foundational mechanistic information that is critically needed to identify specifc strains of Corynebacterium spp. and Corynebacterium-produced compounds with potential as alternative, non-antibiotic therapies to prevent infections by pathogens, such as S. pneumoniae and S. aureus. The three main innovations in our strategy are (1) using functional molecular studies of bacteria in coculture to elucidate the role of different bacterial species within the human microbiome, (2) choosing a system that allows for non-invasive studies directly in the human host, and (3) translating our findings to test a simple intervention on human skin surfaces. Our long-term goal is to develop new and sustainable ways to manage the composition of nasal and skin microbiota to prevent disease by excluding or controlling pathogens.