ACTINOBACILLUS ACTINOMYCETEMCOMITANS VIRULENCE

伴放线放线杆菌毒力

• 批准号: 6379841
• 负责人: JOSEPH M. DIRIENZO
• 金额: $ 20.6万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-01 至 2003-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6379841
• 关键词: Actinobacillus actinomycetemcomitans; CHO cells; affinity chromatography; bacterial cytopathogenic effect; bacterial genetics; bacterial toxins; cytotoxicity; frameshift mutation; genetic mapping; genetic transcription; molecular cloning; northern blottings; oral bacteria; polymerase chain reaction; virulence

DESCRIPTION (adapted from the Investigator's abstract): The gram-negative bacterium, Actinobacillus actinomycetemcomitans, has been the subject of intensive study because it is considered to be a major pathogen of periodontal diseases. Some of the hallmarks of the pathogenic potential of this species are the ability of select strains to invade gingival tissues and to produce relatively high levels of leukotoxin. A number of less-characterized virulence potentials include a variety of cytostatic and cytotoxic factors. The focus of this application is a novel toxin, known as cytolethal distending toxin (CDT), which the Principal Investigator has recently discovered in A. actinomycetemcomitans. In preliminary experiments, the cdt locus was cloned in a non-enteropathogenic strain of Escherichia coli and the complete nucleotide sequence obtained. Extracts of the recombinant cells caused morphological changes in Chinese hamster ovary (CHO) cells, human epithelial cell-like lines and periodontal ligament fibroblasts characteristic of CDT. The major objective of this study is to initiate an investigation towards the complete genetic and biochemical characterization of the CDT of A. actinomycetemcomitans. Three specific aims are proposed. Aim 1 is to identify and characterize the genes and gene products of the putative cdt locus. Aim 2 is to determine the kinetics of cytostatic and cytolytic effects of CDT on cultured cells, and Aim 3 is to examine the genetic organization of the cdt locus. Frame shift mutants will be constructed and employed to examine the contribution of each of the cdt genes to distension and killing activities. The kinetics of cell distension and cytotoxicity will be characterized using CHO cells as a model system. Northern blotting and primer extension will be used to determine if the three genes that comprise the cdt locus are organized into a polycistronic operon. The CDT represents a previously unknown potential virulence factor for A. actinomycetemcomitans. The expression of this novel toxin may have important implications in the development of localized juvenile periodontitis (LJP), as well as in other types of bacterial disease, because it extends the repertoire of virulence potentials of the bacterium. Significant advances have been made in the identification and description of this toxin locus and it is now important to extend our understanding of its expression and biological activities. The availability of the cloned and sequenced cdt genes provides an advantage in the development of tools for the study of the role of this toxin in the pathogenicity of oral disease.

描述（改编自研究者摘要）：革兰氏阴性 放线菌伴放线杆菌（Actinobacillusactinomycetemcomitans）是一种细菌， 因为它被认为是牙周炎的主要病原体， 疾病该物种致病潜力的一些特征是 选择菌株侵入牙龈组织并产生 白细胞毒素水平相对较高一些不太明确的毒力 潜在的包括多种细胞生长抑制因子和细胞毒性因子。的焦点 本申请是一种新的毒素，称为细胞致死膨胀毒素（CDT）， 这是首席研究员最近在A. 伴放线菌在初步实验中，克隆了cdt基因座 在非肠致病性大肠杆菌菌株中， 获得的核苷酸序列。重组细胞的提取物引起 中国仓鼠卵巢（CHO）细胞、人上皮细胞 CDT的细胞样细胞系和牙周膜成纤维细胞特征。的 本研究的主要目的是对 完整的遗传和生化特性的CDT的A。 伴放线菌提出了三个具体目标。目标1：识别 并表征推定的CDT基因座的基因和基因产物。目的2 是为了确定CDT的细胞抑制和细胞溶解作用的动力学， 目的3是检测CDT的遗传结构 基因座将构建移码突变体并用于检查 每个CDT基因对扩张和杀伤活性的贡献。的 细胞膨胀和细胞毒性的动力学将使用CHO 细胞作为一个模型。北方印迹和引物延伸将用于 确定构成CDT基因座的三个基因是否被组织成 多顺反子操纵子CDT代表了一种以前未知的潜力 毒力因子A.伴放线菌这部小说的表达方式 毒素可能对局部青少年的发育有重要影响， 牙周炎（LJP），以及其他类型的细菌性疾病，因为它 扩展了细菌的毒力潜力库。显著 在鉴定和描述这种毒素方面已经取得了进展 现在重要的是扩大我们对它的表达的理解， 生物活动。cdt基因克隆和测序的可用性 提供了一个优势，在工具的发展，为研究的作用， 这种毒素在口腔疾病的致病性。