Role of metal transport in Streptococcus sanguinis virulence and competitiveness

 描述(申请人提供)：感染性心内膜炎是一种严重的心脏感染，死亡率超过20%。这种疾病被认为是当细菌或其他微生物进入血液，附着在先前受损的心脏瓣膜上并繁殖时发生的。口腔或绿色链球菌是这种疾病的主要原因，在这一组中，血链球菌尤其重要。心内膜炎的预防主要依赖于在牙科手术前对高危患者进行抗生素预防，这可能会导致菌血症。然而，现在人们认识到，大多数心内膜炎病例是由于日常活动导致菌血症，而牙科抗生素预防对此没有任何保护作用。毒力决定因素的鉴定可以为新的预防措施提供新的目标，这些措施能够为高危人群提供持续的保护，而不会选择口腔中的抗生素耐药性。为了实现这一目标，我们在动物模型中对血链球菌突变株进行了多次筛选，以确定心内膜炎毒力的丧失。这些研究已经确定了一种脂蛋白，SSAB，它对心内膜炎的毒力是唯一重要的，也是一个有希望的治疗或预防的靶点。SSAB和它所属的家族中的其他成员一样，在锰(也许还有铁)的吸收、耐氧和毒力方面发挥作用。有许多机制可以促进耐氧性和毒力，但目前还不清楚哪些机制在链球菌中使用或最重要。我们在SSAB的潜在功能方面取得了一些最新进展，这些功能不仅对血链球菌来说是新的，而且对任何链球菌都是新的，包括表征了对有氧生长至关重要的依赖于锰的核糖核苷酸还原酶，表征了SSAB突变体中保持的超氧化物歧化酶活性的生物学相关性，确定了模拟兔心内膜炎模型结果的生理相关的体外生长条件，观察了SSAB蛋白的金属依赖加工，以及发现了SSAB在耐酸性中的重要作用。我们建议通过研究SSAB的调节和功能来扩展这些新的发现。这将为我们最近的发现所产生的与金属运输相关的多个假说提供任何链球菌的第一次测试。鉴于我们知道SSAB突变体的主要缺陷是代谢，我们研究的一个重要部分将是对在严格控制的条件下生长的O2和酸应激细胞进行前所未有的代谢组学分析。随后将对选定的菌株进行体外和体内口腔定植和心内膜炎毒力模型的测试，以测试我们发现的生物学相关性。这项研究的结果有望是 直接适用于一些已知需要锰的其他重要病原体和口腔定殖者。此外，这项研究有望找到控制口腔健康和预防感染性心内膜炎的新方法。

 DESCRIPTION (provided by applicant): Infective endocarditis is a serious infection of the heart with mortality rates in excess of 20%. This disease is thought to occur when bacteria or other microbes gain access to the blood, attach to previously damaged heart valves, and multiply. The oral or viridans streptococci are a leading cause of this illness, and among this group, Streptococcus sanguinis is especially important. Prevention of endocarditis relies primarily on antibiotic prophylaxis for at-risk patients prior to dental procedures that are likelyto result in bacteremia. However, it is now recognized that most cases of endocarditis result from daily activities that lead to bacteremia, for which dental antibiotic prophylaxis provides no protection. The identification of virulence determinants could provide new targets for novel prophylactic measures capable of providing continuous protection for persons at risk without selecting for antibiotic resistance in the mouth. Toward this goal, we have performed multiple screens of S. sanguinis mutants for loss of endocarditis virulence in an animal model. These studies have identified a lipoprotein, SsaB, that is uniquely important for endocarditis virulence and is also a promising target for therapy or prevention. SsaB, like other members of the family to which it belongs, functions in Mn (and perhaps Fe) uptake, O2 tolerance, and virulence. There are a number of mechanisms by which Mn could contribute to O2 tolerance and virulence, but it is not clear exactly which mechanisms are employed or are most important in streptococci. We have made a number of recent advances in relation to SsaB's potential functions that are novel not just for S. sanguinis, but for any Streptococcus, including characterization of an Mn-dependent ribonucleotide reductase that is essential for aerobic growth, characterization of the biological relevance of superoxide dismutase activity remaining in an ssaB mutant, identifying physiologically relevant in vitro growth conditions that mimic findings from the rabbit endocarditis model, the observation of metal-dependent processing of the SsaB protein, and the finding of an important role for SsaB in acid tolerance. We propose to expand upon these novel findings by examining SsaB regulation and function. This will provide the first test in any Streptococcus of multiple hypotheses generated by our recent findings that are related to metal transport. Given that we know that the primary defect of an ssaB mutant is metabolic, an important part of our study will be an unprecedented metabolomic analysis of O 2 and acid-stressed cells grown under carefully controlled conditions. This will be followed by testing of selected strains in in vitro and in vivo models of oral colonization and endocarditis virulence to test the biological relevance of our findings. The results of this study promise to be directly applicable to a number of other important pathogens and oral colonizers that are known to require Mn. Moreover, this study promises to identify novel approaches to control oral health and prevent infective endocarditis.