P. gingivalis genes essential for tobacco smoke survival

牙龈卟啉单胞菌基因对烟草烟雾生存至关重要

• 批准号: 10642944
• 负责人: David A Scott
• 金额: $ 37.17万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10642944
• 关键词: Abscess; Adult; Alveolar Bone Loss; Anaerobic Bacteria; Award; Bacteria; Basic Science; Biological Assay; Biological Process; Blood Vessels; Cells; Chronic; Cigarette; Clinical; Communicable Diseases; Complex; Consumption; Data; Databases; Developed Countries; Diabetes Mellitus; Disease; Dose; Economic Burden; Environment; Epidemiology; Epithelial Cells; Epithelium; Essential Genes; Exhibits; Exposure to; Financial cost; Funding Mechanisms; Future; Gene Deletion; Genes; Genome; Gingiva; Growth; Health; Homologous Gene; Immune; Immune response; In Vitro; Infection; Inflammation; Invaded; Libraries; Lung diseases; Modeling; Mouth Diseases; Mus; National Institute of General Medical Sciences; New Zealand; Nicotine; Oral; Oral cavity; Osteitis; Patients; Peptide Hydrolases; Periodontal Diseases; Periodontitis; Persons; Physiology; Population; Porphyromonas gingivalis; Predisposition; Premature Birth; Prevention; Refractory; Research; Resistance; Resources; Scandinavia; Smoke; Smoker; Smoking; Standardization; Stress; Subcutaneous abscess; System; Testing; Tissues; Tobacco; Tobacco smoke; Tobacco smoking behavior; Tooth structure; Toxin; Treatment Protocols; Variant; Vascular Diseases; Virulence; Work; biological adaptation to stress; cigarette smoke; cigarette smoke-induced; fitness; immune resistance; in vivo; infection rate; insight; mouse model; mutant; non-smoker; novel; novel vaccines; oral cavity epithelium; pathogen; periodontopathogen; success; therapeutic target; tobacco regulation; tobacco smoke exposure; trait; transcriptome; transcriptome sequencing; translational potential; transposon sequencing

PROJECT SUMMARY/ABSTRACT Periodontitis is a highly prevalent disease associated with several debilitating systemic conditions, including vascular and lung diseases, diabetes mellitus, and pre-term birth. The most recent epidemiological evidence suggests that tobacco smoking accounts for the majority of destructive periodontal disease cases in developed nations. Smoking enhances infection rates and enriches numbers of a key periodontal pathogen, Porphyromonas gingivalis. However, the mechanisms underlying this phenomenon are in need of elucidation. In the first iteration of award, facilitated by a two year NIGMS-funding mechanism, we generated an 80,000 colony transposon sequencing library for P. gingivalis strains ATCC 33277 and exploited this resource to: (i) Successfully identify 256 genes that are putatively essential for surviving tobacco-induced stress; (ii) Delineate a “core stress genome” commonly required for tobacco-survival, epithelial colonization and subcutaneous abscess formation; and (iii) Validated several genes as conditionally essential for P. gingivalis to survival of cigarette smoke- induced stress in vitro by generating single gene deletion mutants that exhibit either reduced absolute fitness in competition assays with the parental P. gingivalis strain in smoked medium; (iv) We have also generated a novel murine model of tobacco-exacerbated periodontitis. We now plan to build on these successes via three specific aims, as follow: Aim 1: Further elucidate P. gingival CSE-related survival strategies by generating mutants in single genes that are both CSE-essential (TnSeq data) ;and tobacco-regulated (RNAseq and qPCR); yet not compromised in their ability to colonize oral epithelial cells (TIGK model) or immune resistance (murine abscess model). Aim 2: Determine in vivo relevance of putatively CSE-essential genes using a murine smoke-exposure chamber model in a chronic Baker model of P. gingivalis-induced periodontitis. Aim 3: Establish the broad applicability of data gathered in Aims 1 and 2 by confirming biological function and distribution of CSE essential genes in multiple P. gingivalis strains. Long term advances are likely to include (a) A better understanding of how P. gingivalis thrives in a tobacco-toxin rich environment; (b) Future therapeutic targeting of essential genes to control P. gingivalis infection in smokers (CSE-essentiality data) and in general (multiple disease-relevant conditions, core stress genome data); (c) Alternate treatment regimens for smokers based on mechanistic insight into smoke-induced and/or exacerbated periodontal diseases;and (d) The establishment of an essential gene database for tobacco- enhanced pathogens that will facilitate the identification of common bacterial strategies for surviving tobacco smoke exposure, thus, broadening the significance of the research beyond the oral cavity.

项目概要/摘要 牙周炎是一种高度流行的疾病，与多种使人衰弱的全身性疾病相关， 包括血管和肺部疾病、糖尿病和早产。最新的流行病学。 有证据表明，吸烟是造成破坏性牙周病的主要原因 发达国家吸烟会增加感染率并增加关键牙周病原体的数量， 然而，这种现象背后的机制需要阐明。 在为期两年的 NIGMS 资助机制的推动下，第一次奖励，我们产生了 80,000 个菌落 牙龈卟啉单胞菌菌株 ATCC 33277 的转座子测序文库，并利用该资源： (i) 成功鉴定出 256 个被认为对于摆脱烟草引起的应激至关重要的基因； (ii) 描绘烟草生存、上皮定植通常所需的“核心应激基因组” 和皮下脓肿形成； (iii) 验证了几个基因对于牙龈卟啉单胞菌在香烟烟雾中的生存是有条件必需的- 通过产生单基因缺失突变体在体外诱导应激，这些突变体表现出降低的 在烟熏培养基中与亲本牙龈卟啉单胞菌菌株的竞争测定中的绝对适应性； （iv）我们还建立了一种新型的烟草加剧牙周炎的小鼠模型。 我们现在计划通过以下三个具体目标在这些成功的基础上再接再厉： 目标 1：通过在单基因中产生突变体进一步阐明牙龈卟啉单胞菌 CSE 相关的生存策略 既是 CSE 必需的（TnSeq 数据）；又是烟草监管的（RNAseq 和 qPCR）；但在这方面并未受到影响； 它们定殖口腔上皮细胞（TIGK 模型）或免疫抵抗（鼠脓肿模型）的能力。 目标 2：使用小鼠烟雾暴露确定假定的 CSE 必需基因的体内相关性 牙龈卟啉单胞菌引起的牙周炎的慢性贝克模型中的腔室模型。 目标 3：通过确认生物功能，建立目标 1 和 2 中收集的数据的广泛适用性 以及 CSE 必需基因在多种牙龈卟啉单胞菌菌株中的分布。 长期进展可能包括 (a) 更好地了解牙龈卟啉单胞菌如何在环境中茁壮成长 (b) 未来治疗牙龈卟啉单胞菌的关键基因 吸烟者感染（CSE-必要性数据）和一般情况（多种疾病相关病症、核心应激 (c) 基于对烟雾诱发机制的了解，为吸烟者提供替代治疗方案 (d) 建立烟草重要基因数据库 增强的病原体将有助于识别烟草存活的常见细菌策略 因此，烟雾暴露将研究的意义扩大到口腔之外。