Virulence Gene Discovery in Outbreak-Associated Pneumococci

描述（由申请人提供）：由肺炎链球菌（肺炎球菌）引起的传染病暴发在20世纪初的美国很常见。今天，在美国，肺炎球菌暴发发生在宿主人群中，具有可识别的危险因素。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动态变化，将扩大高毒力，与疫情相关的菌株的生态位。导致肺炎球菌爆发的细菌毒力因素尚不清楚。在本提案中，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的荚膜血清型12F肺炎球菌的候选毒力基因，并在其他国家出现高毒力特征。我们将使用多位点序列分型来表征12F菌株的克隆多样性，并将使用抑制性减法杂交（Aim 1）来鉴定代表性菌株的附属基因。我们假设辅助基因可能包括导致12F肺炎球菌高毒力特征的毒力基因。我们将筛选从鼻咽定植和侵袭性疾病中分离出来的肺炎球菌的各种集合，使用多重PCR发现辅助基因，以突出那些在侵袭性疾病中显著过度代表的基因（目的2）。这一建议建立了未来的实验研究，将解决在这里发现的候选毒力基因的功能。公共卫生相关性：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，在其他国家出现的这种血清型具有更高的潜在毒力。揭示这些菌株的进化起源和任何与其毒力潜力相关的新基因将提供有关促进暴发的细菌因素的基本知识，并可能最终揭示细菌发病的新机制、监测的新标记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.