The impact of hypoxia on Staphylococcus aureus metabolism and virulence during osteomyelitis

项目摘要/摘要 金黄色葡萄球菌是一种重要的人类病原体，可引起危及生命的感染。 在各种宿主组织中。骨髓炎是一种侵袭性和衰弱的骨骼感染，是最常见的 葡萄球菌病的常见表现。事实上，骨髓炎约占全球 在美国，每1000名儿科医院的入院者，并使高达25%的开业人数复杂化 骨折。由于普遍存在的抗菌素耐药性，骨感染是出了名的难以治疗。 以及病原体诱导的骨重建，这限制了抗生素对感染灶的渗透。S. 到目前为止，金黄色葡萄球菌是肌肉骨骼感染的最常见原因，但 人们对葡萄球菌在体内存活并最终破坏骨骼知之甚少。的首要目标是 这项建议是为了了解金黄色葡萄球菌如何调节其毒力和代谢程序以在 骨髓炎期间的骨骼。 像大多数哺乳动物组织一样，骨骼天生就是低氧的。此外，维护骨骼健康 需要常驻成骨细胞和骨吸收破骨细胞持续的骨转换。这些 反过来，骨骼细胞需要以高葡萄糖摄取率为特征的特殊新陈代谢，这是 预计将限制入侵病原体可用的碳源。为了更好地理解S。 金黄色杆菌在这种低氧和代谢独特的环境中茁壮成长，我们创造了一个强大的小鼠模型 能够精确量化细菌负荷和骨转换的骨髓炎。通过应用 转座子测序(TnSeq)到这个骨髓炎模型，我们识别了&gt；200个葡萄球菌基因 对骨骼中的生存很重要。重要的是，细菌对缺氧的反应被发现是至关重要的。 骨髓炎期间存活的决定因素，因为低氧生长不仅决定了能量的产生 葡萄球菌使用的策略，但也增加了依赖于群体的毒力因子的产生， 参与骨质破坏。根据这些初步数据，我们假设金黄色葡萄球菌在 骨的促进是由(A)对低氧反应的法定人数调节的毒力因子的表达和(B)特异性的 营养利用计划，使骨骼在独特的新陈代谢环境中生长。建议数 AIMS将检验这一假说，以确定(I)低氧生长触发增加的机制 葡萄球菌的毒力，(Ii)支持细菌在骨骼中生长的代谢途径，以及(Iii) 宿主低氧信号通路在骨髓炎抗菌免疫和骨重建中的作用。 这些研究的完成将阐明侵袭性感染期间微生物的生存策略，确定 低氧对细菌发病机制的影响，并有助于满足新的骨髓炎的迫切需要 通过定义假定的抗菌剂和抗毒力靶标进行治疗。

PROJECT SUMMARY / ABSTRACT Staphylococcus aureus is an important human pathogen, capable of causing life-threatening infections in a variety of host tissues. Osteomyelitis, an invasive and debilitating infection of bone, is one of the most common manifestations of staphylococcal disease. Indeed, osteomyelitis accounts for approximately 2-3 of every 1,000 admissions to pediatric hospitals in the United States, and complicates up to 25% of open fractures. Bone infections are notoriously refractory to treatment due to widespread antimicrobial resistance and pathogen-induced bone remodeling, which limits penetration of antibiotics into the infectious focus. S. aureus is by far the most common cause of musculoskeletal infection, yet the mechanisms by which staphylococci survive within and ultimately destroy bone are poorly understood. The overarching objective of this proposal is to understand how S. aureus regulates its virulence and metabolic programs to survive within bone during osteomyelitis. Bone, like most mammalian tissues, is inherently hypoxic. Moreover, maintenance of skeletal health requires constant bone turnover by resident bone-forming osteoblasts and bone-resorbing osteoclasts. These skeletal cells, in turn, require a specialized metabolism characterized by high rates of glucose uptake, which is expected to limit the carbon sources available to invading pathogens. In order to better understand how S. aureus thrives within this hypoxic and metabolically unique environment, we created a powerful murine model of osteomyelitis capable of precise quantification of both bacterial burdens and bone turnover. By applying transposon sequencing (TnSeq) to this osteomyelitis model, we identified >200 staphylococcal genes important for survival in bone. Importantly, bacterial responses to hypoxia were found to be critical determinants of survival during osteomyelitis, as hypoxic growth not only dictates the energy production strategies used by staphylococci, but also augments the production of quorum-dependent virulence factors that participate in bone destruction. Based on these preliminary data, we hypothesize that S. aureus survival in bone is facilitated by (a) quorum-regulated virulence factor expression in response to hypoxia, and (b) specific nutrient utilization programs that enable growth in the unique metabolic environment of bone. The proposed Aims will test this hypothesis to determine (i) the mechanism by which hypoxic growth triggers increased staphylococcal virulence, (ii) the metabolic pathways that support bacterial growth in bone, and (iii) the role of host hypoxic signaling pathways in antibacterial immunity and bone remodeling during osteomyelitis. Completion of these studies will elucidate microbial survival strategies during invasive infection, determine the impact of hypoxia on bacterial pathogenesis, and help to meet a critical need for new osteomyelitis therapeutics by defining putative antimicrobial and anti-virulence targets.