Mechanisms and Circumvention of Daptomycin Resistance in Streptococcus mitis

摘要 草绿色组链球菌（VGS），尤其是缓症链球菌（Streptococcus mitis），是多种疾病的关键病原体， 侵袭性血管内感染，包括：i）血小板减少症患者的“突破性菌血症”和“中毒性休克” 癌症患者;和ii）感染性心内膜炎（IE）。鉴于青霉素耐药性和其他 β-内酰胺MIC在S.轻症菌株中，严重感染所占比例相对或完全 β-内酰胺类抗生素耐药（R）菌株令人不安。此外，在这种情况下，使用替代疗法的临床结果 方案（例如，万古霉素）一直令人失望，可能与高患病率有关， 万古霉素在这些菌株中的“耐受性”。这促使人们使用新的杀菌剂，如达托霉素 (DAP)严重的S。β-内酰胺耐药菌株的缓解综合征。令人震惊的是， 由DAP治疗诱导的快速、持久和高水平的DAP-R显著降低了对这种治疗的热情。 方法。此外，S.轻症组比所有其他VGS组高2-10倍。的 报告的侵袭性S.出现DAP-R的缓解感染是有限的，这是由于 到目前为止，DAP在此类感染中的使用相对较少。然而，S.缓症β-内酰胺-R 加上VANC治疗在这些综合征中的不一致结果实际上确保了DAP使用的增加， 导致DAP-RS。缓解感染。此外，DAP使用率高的医疗中心最近证实， 在肠球菌中有大量MIC“蠕动”。了解DAP-R在体内出现的机制 S.因此，缓解症加上避免其演变的策略具有重要的临床意义。 我们的初步数据表明，在S。缓和症可能是多因素的， 表型和基因型水平。最有趣的是，我们现在已经展示了两种形式的令人信服的证据， “DAP过度积累”，其中给定链球菌链中的单个细胞可以过度捕获DAP， 要么死亡（“利他性自杀”），要么抵抗DAP杀伤，以保护剩余的细胞群免受 DAP暴露和致死性。这是革兰氏阳性细胞中DAP-R的一种明显独特的机制。 病原体 在这项建议中，我们将使用战略荧光显微镜，流式细胞术与多维 生理询问，单细胞分选加上基因分型，以揭示DAP-R S.缓蚀剂可以抵抗DAP曝光。最后，我们将使用两个充分表征的血管内模型， 感染，离体（腔室模型）和体内（实验兔IE），以确定DAP方案， 避免DAP-R的出现，提高S.缓解。总之，这些研究将揭示 临床策略，以防止出现的DAP-R在S.可能还有其他革兰氏阳性病原体。

ABSTRACT Viridans group streptococci (VGS), especially Streptococcus mitis, are pivotal pathogens in a variety of invasive endovascular infections,including: i) “breakthrough bacteremias” and “toxic shock” in neutropenic cancer patients; and ii) infective endocarditis (IE). Given world-wide trends in penicillin-resistance and other β-lactam MIC “creeps” amongst S. mitis strains, the proportion of serious infections caused by relatively or fully β-lactam-resistant-(R) strains is disturbing. Moreover, clinical outcomes in such cases utilizing alternate regimens (e.g., vancomycin) have been disappointing, presumably related to the high prevalence of vancomycin “tolerance” in such strains. This has prompted use of newer bactericidal agents, like daptomycin (DAP) for severe S. mitis syndromes in strains with β-lactam-resistance. Alarmingly, recent recognition of rapid, durable and high-level DAP-R induced by DAP therapy has significantly reduced enthusiasm for such approaches. In addition, DAP MICs in the S. mitis group are 2-10-fold higher than all other VGS groups. The number of reported clinical cases of invasive S. mitis infections in which DAP-R has emerged is limited, due to the relatively infrequent use of DAP in such infections to-date. However, progressive rise in S. mitis β-lactam-R plus the inconsistent outcomes of VANC therapy in such syndromes virtually assures increased DAP use, leading to DAP-R S. mitis infections. Moreover, medical centers with high DAP usage have recently confirmed substantial MIC “creeps” amongst enterococci. Understanding mechanism(s) of emergence of DAP-R in S. mitis, plus strategies to circumvent its evolution are, thus, of great clinical significance. Our Preliminary Data showed that DAP-R outcomes in S. mitis are likely to be multifactorial on both phenotypic and genotypic levels. Most interestingly, we have now shown compelling evidence of two forms of “DAP hyperaccumulation” in which individual cells in a given streptococcal chain can hyper-capture DAP and either die (“altruistic suicide”) or resist DAP killing, in order to protect the remainder of the cell population from DAP exposures and lethality. This is an apparently unique mechanism of DAP-R amongst gram-positive pathogens. In this proposal, we will use strategic fluorescence microscopy, flow cytometry with multidimensional physiologic interrogations, and single cell sorting plus genotyping to divulge mechanisms by which DAP-R S. mitis can resist DAP exposures. Finally, we will use two well-characterized models of endovascular infections, ex vivo (chamber model) and in vivo (experimental rabbit IE), to define DAP regimens to both circumvent emergence of DAP-R and enhance clearance of S. mitis. In summary, these studies will divulge clinical strategies to forestall emergence of DAP-R in S. mitis and perhaps other Gram-positive pathogens.