ACTINOBACILLUS ACTINOMYCETEMCOMITANS VIRULENCE

描述(改编自《调查者摘要》)：革兰氏阴性 细菌，放线菌伴生放线杆菌，一直是 深入研究，因为它被认为是牙周的主要病原体 疾病。这一物种潜在致病的一些特征是 筛选出的菌株侵入牙龈组织并产生 白毒素水平相对较高。一些特征较少的毒力 潜在的包括多种细胞抑制因子和细胞毒性因子。的关注点 这种应用是一种新的毒素，称为细胞致死性膨胀毒素(CDT)， 首席调查员最近在A. 伴生放线菌。在初步实验中，克隆了CDT基因座 在一株非致病性大肠埃希氏菌中， 获得了核苷酸序列。重组细胞提取物引起的 中国仓鼠卵巢(CHO)细胞、人卵巢上皮细胞的形态变化 CDT特有的细胞样系和牙周膜成纤维细胞。这个 这项研究的主要目的是启动对 完整的遗传和生化特征。 伴生放线菌。提出了三个具体目标。目标1是确定 并对可能的CDT基因座的基因和基因产物进行了鉴定。目标2 目的是确定CDT的细胞抑制和杀伤作用的动力学。 培养的细胞，目标3是检查CDT的遗传组织 轨迹。帧移位突变体将被构建并用于检测 每个CDT基因对扩张和杀伤活动的贡献。这个 细胞膨胀和细胞毒性的动力学将用CHO来表征 细胞作为模型系统。Northern blotting和引物延伸将用于 确定组成CDT基因座的三个基因是否被组织成一个 多顺反子操纵子。CDT代表了一种以前未知的势 伴生放线菌毒力因子。这部小说的表达方式 毒素可能在局限性青少年的发育中有重要意义 牙周炎(LJP)以及其他类型的细菌疾病，因为它 扩展了细菌的毒力潜力。意义重大 这种毒素的鉴定和描述已经取得了进展 轨迹，现在重要的是扩大我们对它的表达和 生物活动。CDT基因克隆和测序的可行性 在开发工具方面提供了优势，用于研究 这种毒素在口腔疾病中具有致病性。

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.