Virulence Gene Discovery in Outbreak-Associated Pneumococci

描述（由申请人提供）：世纪早期，由肺炎链球菌（肺炎球菌）引起的传染病爆发在美国很常见。今天，美国的肺炎球菌疫情发生在具有可识别风险因素的宿主人群中。然而，存在严重的担忧，即最近在美国引入结合疫苗所带来的肺炎球菌种群的动态变化，将扩大高毒力爆发相关菌株的生态位。使肺炎球菌易于引起暴发的细菌毒力因子尚不清楚。在这项提案中，我们将使用分子流行病学方法来发现在美国引起真正爆发并在其他国家出现高毒力特征的荚膜血清型12F肺炎球菌中的候选毒力基因。我们将使用多位点序列分型来表征12F菌株的克隆多样性，并且我们将使用抑制性消减杂交（Aim 1）来鉴定代表性菌株中的辅助基因。我们推测，辅助基因可能包括毒力基因，有助于12 F肺炎球菌的高毒力特性。我们将使用多重PCR筛选从明确定义的宿主人群中的鼻咽定植和侵袭性疾病中分离的肺炎球菌的不同集合，以发现辅助基因，以突出那些在侵袭性疾病中显著过度表达的基因（目的2）。这项建议建立了未来的实验研究，将解决这里发现的候选毒力基因的功能。自1989年以来，荚膜血清型12F肺炎球菌在美国至少引起了四次侵袭性疾病的爆发。此外，该血清型在其他国家出现的毒力潜力增加。揭示这些菌株的进化起源以及与其毒力潜力相关的任何新基因将提供有关促进爆发的细菌因素的基本知识，并可能最终揭示细菌发病机制的新机制，新的监测标志物和疾病预防的新机会。

DESCRIPTION (provided by applicant): Outbreaks of infectious disease caused by the bacterium Streptococcus pneumoniae, the pneumococcus, were common in the United States early in the 20th century. Today, pneumococcal outbreaks in the U.S. occur in host populations with identifiable risk factors. However, there exist serious concerns that the changing dynamics of pneumococcal populations, brought about by the recent introduction of a conjugate vaccine in the U.S., will broaden the niche of hypervirulent, outbreak-associated strains. The bacterial virulence factors that predispose pneumococci to cause outbreaks are unknown. In this proposal, we will use a molecular epidemiological approach to discover candidate virulence genes in capsular serotype 12F pneumococci that have caused genuine outbreaks in the U.S. and have emerged with hypervirulent characteristics in other countries. We will characterize the clonal diversity of 12F strains using multilocus sequence typing, and we will identify accessory genes in representative strains using suppressive subtractive hybridization (Aim 1). We hypothesize that accessory genes may include virulence genes that contribute to the hypervirulent characteristics of 12F pneumococci. We will screen diverse collections of pneumococci, which were isolated from nasopharyngeal colonization and invasive disease in well-defined host populations, for the discovered accessory genes using multiplex PCR, in order to highlight those genes that are significantly over-represented from invasive disease (Aim 2). This proposal sets up future experimental studies that will address the function of candidate virulence genes that are discovered here. PUBLIC HEALTH RELEVANCE: Capsular serotype 12F pneumococci have caused at least four outbreaks of invasive disease in the United States since 1989. In addition, this serotype has emerged with increased virulence potential in other countries. Revealing the evolutionary origin of these strains and any novel genes linked to their virulence potential will provide basic knowledge about the bacterial factors that promote outbreaks, and may ultimately reveal new mechanisms of bacterial pathogenesis, new markers for surveillance, and new opportunities for disease prevention.