Metabolic Discrimination of Unknown Bacterial Pathogens

描述（由申请人提供）：范德比尔特大学和阿伯丁试验场的美国陆军埃奇伍德化学和生物中心（ECBC）之间的合作将开发一种采用多相传感的广谱活性检测技术，以提供生化武器、未知药物或其他威胁的生物功能特征。这些特征将与先进的算法一起使用，以区分作用于一组靶细胞系的不同药物。所提出的方法是非常通用和普遍的，因为我们正在测量毒素的生物影响，而不仅仅是它们的存在。这将解决一个至关重要的和尚未满足的需求，诊断工具能够识别未知或重新设计的威胁代理的行动机制，无视现有的，代理特定的传感器的准确检测。将创建诊断工具，以确定对炭疽、蓖麻毒素、葡萄球菌肠毒素B（SE B）和肉毒梭菌毒素有反应的细胞系中发生的代谢和信号传导途径的关键变化的特征。毒素与细胞的相互作用导致大量的代谢和信号传导事件，因为毒素以尚未完全理解的特异性和非特异性方式破坏正常的细胞功能。为了提供一种手段来表征未知毒素的影响，所提出的诊断系统将在一系列时间和体积尺度上监测特定代谢和信号传导途径的关键参数。 新开发的终点代谢率的孔板方案将与信号事件的商业荧光测定相结合。此外，相同的代谢和信号事件将被捕获为动态生物特征，使用修改的细胞传感器，同时监测多个分析物的时间尺度分钟。 这种方法将按比例缩小，以实现动态分辨率的数量级秒的微型纳米生理仪。这种方法应该在发现新药和意外和/或不希望的生理活性;环境和工业毒理学;代谢组学和信号传导;以及筛选病原菌和毒素的对策，治疗和预防中找到广泛的应用。

DESCRIPTION (provided by applicant): A collaboration between Vanderbilt University and the U.S. Army Edgewood Chemical and Biological Center (ECBC) at the Aberdeen Proving Grounds will develop a wide-spectrum, activity-detection technology that employs multiphasic sensing, in order to provide a biofunctional signature of a CBW agent, unknown drug, or other threat. The signatures will be used with advanced algorithms to discriminate between different agents acting on a set of target cell lines. The proposed approach is extraordinarily versatile and general, because we are measuring the biological impact of the toxins, rather than simply their presence. This will address a critically important and as-yet-unmet need for diagnostic tools capable of identifying the mechanism of action of unknown or reengineered threat agents that defy accurate detection by existing, agent-specific sensors. The diagnostic tools will be created to establish signatures of key changes in metabolic and signaling pathways that occur in cell lines responsive to Anthrax, Ricin, Staphylococcal Enterotoxin B (SEB), and Clostridium Botulinum toxins. The interaction of the toxins with cells leads to a multitude of metabolic and signaling events, as toxins disrupt normal cellular functions in specific and non-specific ways that are not yet fully understood. To provide a means to characterize the effects of unknown toxins, the proposed diagnostic system will monitor key parameters of specific metabolic and signaling pathways over a spectrum of time- and volume-scales. Newly developed well-plate protocols for end-point metabolic rates will be coupled with commercial fluorescence assays for signaling events. Additionally, the same metabolic and signaling events will be captured as dynamic biosignatures using a modified Cytosensor that simultaneously monitors multiple analytes on the time scale of minutes. This approach will be scaled down to achieve dynamic resolution on the order of seconds in a microfabricated NanoPhysiometer. This approach should find wide application in the discovery of new drugs and unexpected and/or undesired physiological activity; environmental and industrial toxicology; metabonomics and signaling; and screening of countermeasures, therapies, and prophalaxis for pathogenic bacteria and toxins.