Novel Methodology for Rapid Antibiotic Susceptibiility Testing in S. aureus

描述（由申请人提供）：细菌对抗生素的耐药性的出现仍然是一个重大的公共卫生挑战。导致抗生素耐药性上升的因素包括广谱抗生素的广泛和不当处方以及患者不遵守抗生素治疗方案。卫生界的应对措施是限制抗生素使用，并在可能的情况下开有针对性的窄谱抗菌药物。同时，目前抗菌药物的开发越来越向窄谱药物集中。为了有效使用抗菌药物治疗，必须开发新的抗生素敏感性检测方法，使医生能够在尽可能接近初始检查的情况下开出这些窄谱抗生素。在这里，我们建议研究一种新的机制，这将是诊断的基础，以满足这一重要的公共卫生需求。在这个R21应用程序中，波士顿大学弗劳恩霍夫制造创新中心的Alexis Sauer-Budge博士和布里格姆妇女医院和哈佛医学院的Jean Lee博士合作探索了一种假设，即对细菌的机械应力会破坏细胞壁并迅速启动细胞壁修复生物合成途径。在针对细胞壁生物合成的抗生素存在的情况下，敏感菌株将无法从损伤中恢复并死亡，而耐药菌株将恢复。因此，当在荧光染料染色的情况下施加机械应力时，可以通过荧光显微镜快速确定细菌的敏感性。我们建议通过建立流动池和光学仪器，改变实验条件来优化抗生素敏感性检测方案，并量化不同菌株和细菌生长阶段的差异来研究这一假设。我们建议使用金黄色葡萄球菌的模型系统，包括甲氧西林敏感菌株和甲氧西林耐药菌株。如果成功，我们预计这种方法将适用于广泛的细菌和抗生素的各种作用机制。公共卫生相关性：多重耐药细菌感染的日益流行是一个日益严重的公共卫生问题。为了对抗这一趋势并使医生充分能够开出适当的抗菌药物治疗，必须开发新的抗生素敏感性快速诊断方法。在这里，我们建议通过开发一种以金黄色葡萄球菌为模型系统的快速检测抗生素敏感性的新技术来解决抗生素敏感性诊断方面的空白。

DESCRIPTION (provided by applicant): The emergence of bacterial resistance to antibiotics continues to be a significant public health challenge. Factors contributing to the rise in antibiotic resistance include widespread and inappropriate prescription of broad spectrum antibiotics and patient non-compliance to antibiotic regiments. The healthcare community is responding by limiting antibiotics and prescribing targeted narrow-spectrum antimicrobial therapies when possible. Meanwhile, current antimicrobial development is focused more and more on narrow-spectrum drugs. To enable the effective use of antimicrobial therapy, new methods for detection of antibiotic susceptibility must be developed that enable physicians to prescribe these narrow spectrum antibiotics as close to initial examination as possible. Here, we propose to investigate a novel mechanism that would be the basis for a diagnostic to meet this important public health need. In this R21 application, Dr. Alexis Sauer-Budge at the Fraunhofer Center for Manufacturing Innovation at Boston University and Dr. Jean Lee of Brigham and Women's Hospital and Harvard Medical School have teamed up to explore the hypothesis that mechanical stress on bacteria will damage the cell wall and rapidly initiate cell wall repair biosynthesis pathways. In the presence of antibiotics targeted at cell wall biosynthesis, susceptible strains will be unable to recover from the damage and will die, whereas resistant strains will recover. The susceptibility of the bacteria can therefore be determined rapidly via fluorescence microscopy when the mechanical stress is applied in the presence of fluorescent dyes which stain the damaged bacteria. We propose to investigate this hypothesis by building a flow cell and optical apparatus, varying experimental conditions to optimize the protocol for antibiotic susceptibility detection, and quantify the variability due to different strains and growth phases of the bacteria. We propose to use the model system of Staphylococcus aureus, both methicillin-susceptible and methicillin- resistant strains. If successful, we anticipate this methodology will be applicable to a wide range of bacteria and antibiotics of various mechanisms of action. PUBLIC HEALTH RELEVANCE: The increasing prevalence of multi-drug resistant bacterial infections is a growing public health problem. To combat the trend and to fully enable physicians to prescribe appropriate antimicrobial therapy, new rapid diagnostic methods for antibiotic susceptibility must be developed. Here, we propose to address the gap in antibiotic susceptibility diagnostics by developing a novel technique for rapid detection of antibiotic susceptibility using Staphylococcus aureus as our model system.