Defining the molecular inventory of staphylococcal tissue abscesses and its effects on the host-pathogen interface

摘要： 多药耐药细菌感染，特别是由金黄色葡萄球菌引起的感染，是公认的 作为21世纪最大的威胁之一。血液感染是最严重的葡萄球菌 尽管我们采用了最好和最新的治疗方法，但疾病的表现往往是致命的。器官脓肿是 金黄色葡萄球菌是金黄色葡萄球菌全身性感染的主要贡献者，也是入侵病原体的初始宿主。 脓肿的形成本身遵循不同的发育阶段，由宿主和细菌积极促进，并且 最终为金黄色葡萄球菌创造了一个有利的利基。虽然过去的研究已经产生了关于 脓肿的结构，我们缺乏关于脓肿的分子组成的信息，特别是在 不同的脓肿阶段。这种对脓肿形成过程中分子事件的有限了解是 尤其令人担忧，因为它阻碍了有针对性地设计抗葡萄球菌策略的有意义的尝试。 我们的初步数据显示，脓肿形成的特点是宿主广泛移位 在一种被称为营养免疫的过程中，过渡金属接近脓肿。因此，在体内 成像显示，脓肿内的细菌缺乏锌和铁。因为可用金属水平可以 作为入侵病原体的生物标志物，我们假设波动的元素分布 与脓肿形成相关的细菌活动。沿着这些思路，我们发现了锌饥饿的起点 随后与不同的免疫细胞群接触。然而，除了这些发现之外， 与金属重新定位有关的年表和因素，金黄色葡萄球菌对这些刺激的检测，以及相应的 细菌的反应完全没有被探索过。因此，我们计划在这项提案中解决这些问题。 目前设计对葡萄球菌组织发育的有意义的调查的一个障碍 脓肿是一种显著程度的脓肿异质性，可能是不同发育阶段的结果 同一器官中的个别病变。为了解释组织脓肿的非同步性质，我们有 确定了一组可用于不同脓肿阶段的潜在蛋白质标志物。这些蛋白质将作为 分子时钟，这样我们就可以在整个发育过程中跟踪单个脓肿的进展。 基于这些标记，我们将创建活体记者并表征发育中的分子清单 脓肿，重点是元素和蛋白质组成的变化。在这里，我们将关联不同的 活体成像方式，包括3D生物发光成像、核磁共振和成像质谱仪， 微液萃取表面分析的先进蛋白质组学。一旦我们确定了脓肿是如何 脓肿形成不同阶段微环境的变化，我们将进行转录组分析 以评估环境刺激如何影响葡萄球菌的病理生理，以及在 转，脓肿发展。结合起来，拟议的实验将检查宿主病原体发生的事件 这是一项重要的国际合作，并为抗击葡萄球菌感染的新的、有针对性的治疗战略铺平了道路。

Summary: Multidrug-resistant bacterial infections, particularly those caused by Staphylococcus aureus, are recognized as one of the greatest threats of the 21st century. Bloodstream infections are the most severe staphylococcal disease manifestation and are often fatal despite our best and most current therapies. Organ abscesses are primary contributors to S. aureus systemic infections and serve as initial reservoirs for the invading pathogen. Abscess formation itself follows distinct developmental stages, is actively facilitated by host and bacterium, and ultimately creates an advantageous niche for S. aureus. While past studies have generated an overview of abscess architecture, we lack information on the molecular composition of abscesses, particularly in the context of different abscess stages. This limited knowledge of the molecular events during abscess formation is especially alarming, for it hinders meaningful attempts at targeted design of anti-staphylococcal strategies. Our preliminary data show that abscess formation is characterized by the host’s extensive relocation of transition metals in proximity to the abscess in a process known as nutritional immunity. Consequently, in vivo imaging reveals that bacteria within the abscess are starved for zinc and iron. Since available metal levels can serve as biomarkers for invading pathogens, we hypothesize that fluctuating elemental distributions orchestrate bacterial activities associated with abscess formation. Along these lines, we showed that zinc starvation primes S. aureus for subsequent contact with different immune cell populations. Beyond these findings, however, the chronology and factors involved in metal relocation, detection of these stimuli by S. aureus, and corresponding bacterial responses are entirely unexplored. We thus plan to address these questions in this proposal. One current barrier to the design of meaningful investigations into the development of staphylococcal tissue abscesses is a significant degree of abscess heterogeneity, likely a result of different developmental stages of individual lesions in the same organ. To account for the non-synchronous nature of tissue abscesses, we have identified a group of potential proteinaceous markers for different abscess stages. These proteins will serve as molecular clocks so we can follow the progression of individual abscesses through the developmental process. Based on these markers, we will create in vivo reporters and characterize the molecular inventory of developing abscesses, focusing on changes in elemental and proteinaceous compositions. Here, we will correlate various in vivo imaging modalities, including 3D-bioluminiscent imaging, MRI, and imaging mass spectrometry, with advanced proteomics via micro Liquid Extraction Surface Analysis. Once we have established how the abscess microenvironment changes during different phases of abscess formation, we will perform transcriptome analysis of bacterial subpopulations to assess how environmental stimuli affect staphylococcal pathophysiology and, in turn, abscess development. Combined, the proposed experiments will examine the events at the host-pathogen interface and pave the way for novel and targeted treatment strategies to combat staphylococcal infections.