Characterization of a Kingella kingae leukotoxin

描述(申请人提供)：Kingella kingae是奈瑟亚科的一种挑剔的革兰氏阴性球菌，是人类口咽部菌群的正常居住者。该细菌属于Hacek组，可引起感染性心内膜炎。最近，由于分离和鉴定技术的改进，世界各地关于侵袭性国王克雷伯菌感染的报道越来越多，这表明该菌是儿童和成人心脏和骨骼感染的重要原因。尽管关于K.kingae感染的临床和诊断方面的信息正在涌现，但这种病原体的毒力机制在很大程度上仍然不清楚。我们的结果表明，Kingae产生一种与细胞膜相关的有效蛋白白毒素(KTX)。这是国王克雷伯氏菌白毒素的首次证据，它可能是在感染期间逃避宿主免疫反应的一个重要毒力因子。本研究的目的是对KTX进行鉴定和表征。在拟议的研究中，我们将：-研究KTX在不同K.kingae临床分离株中的分布；-测序Kkingae基因组；-从Kkingae中提纯KTX，然后进行蛋白质鉴定；-鉴定KTX基因并使用随机和定点突变方法产生KTX突变体；-表征KTX对不同人类细胞的毒性。确定国王克雷伯氏菌的毒力因子对于了解病原菌如何致病至关重要。本研究结果对王氏克雷伯氏菌感染的防治具有一定的指导意义。 与公共卫生相关：Kingella kingae是一种人类口腔细菌，可导致心脏和骨骼系统感染。我们建议对能够破坏人类白细胞的Kingae毒素进行表征。本研究结果对王氏克雷伯氏菌感染的防治具有一定的指导意义。

DESCRIPTION (provided by applicant): Kingella kingae is a fastidious gram-negative coccobacillus of the Neisseriaceae family and is a normal inhabitant of the human oropharyngeal flora. This bacterium belongs to HACEK group and can cause infective endocarditis. Recently, as the result of improved isolation and identification techniques, an increasing number on invasive K. kingae infections have been reported throughout the world, suggesting that the organism is an important cause of heart and skeletal infections in pediatric and adult patients. Despite the emerging body of information on the clinical and diagnostic aspects of K.kingae infections, the virulence mechanisms of this pathogen remain largely unknown. Our results suggest that K. kingae produces a potent proteinaceious leukotoxin (Ktx) that is associated with cell membranes. This is the first evidence of a K. kingae leukotoxin that may be an important virulence factor for evasion of the host immune response during infection. The goal of this research is identify and characterize Ktx. In the proposed research we will: - Study distribution of Ktx in different K. kingae clinical isolates; - Sequence K. kingae genome; - Purify Ktx from K. kingae followed by the protein identification; - Identify ktx gene and generate ktx mutants using random and site-directed mutagenesis approaches; - Characterize Ktx toxicity to different human cells. Identification of K. kingae virulence factors will be essential to understanding how the pathogen causes disease. The results obtained in this study will have significance for the prevention and treatment of K. kingae infections. PUBLIC HEALTH RELEVANCE: Kingella kingae is a human oral bacterium that can cause infections of heart and skeletal system. We propose to characterize K. kingae toxin that can destroy human white blood cells. The results obtained in this study will have significance for the prevention and treatment of K. kingae infections.