Characterization of a Kingella kingae leukotoxin

描述（由申请人提供）：Kingella kingae是一种挑剔的革兰氏阴性球芽孢杆菌属奈瑟菌科，是人类口咽菌群的正常居民。这种细菌属于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，在世界范围内报道了越来越多的侵袭性国王卡氏菌感染，这表明该生物是儿童和成人患者心脏和骨骼感染的重要原因。尽管关于金氏杆菌感染的临床和诊断方面的信息不断涌现，但这种病原体的毒力机制在很大程度上仍然未知。我们的研究结果表明，王氏K.产生一种与细胞膜相关的强效蛋白性白毒素（Ktx）。这是第一个证据表明，在感染过程中，王氏白毒素可能是逃避宿主免疫反应的重要毒力因子。本研究的目的是识别和表征Ktx。在拟议的研究中，我们将：-研究Ktx在不同临床分离株中的分布；-王氏金龟基因组序列；-从K. kingae中纯化Ktx并进行蛋白鉴定；-使用随机和定点诱变方法鉴定ktx基因并产生ktx突变体；-表征Ktx对不同人体细胞的毒性。鉴定国王K.毒力因子将是必不可少的了解病原体如何引起疾病。本研究结果对金氏杆菌感染的防治具有重要意义。

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.