Bloodstream infection detection directly on whole blood

 描述(由申请人提供)：败血症是对感染的全身炎症反应，每年在美国影响大约100万人，并导致高达50%的严重感染患者死亡。快速和适当的抗菌药物使用对于脓毒症的治疗至关重要。在感染性休克患者中，治疗每延迟一小时，存活率就会降低8%。血培养是金标准的诊断方法，然而，它需要一天以上的时间才能得出结果。因此，患者接受广谱抗生素治疗，或可能在最初几个小时内不接受治疗。分子检测是一种很有吸引力的解决方案，然而，目前可用的检测是基于细菌基因组DNA的聚合酶链式反应扩增和复杂的检测策略。这些技术容易受到DNA污染，缺乏足够的灵敏度，而且价格昂贵。在这里，我们建议开发一种新的平台，用于快速、廉价地鉴定全血中的微生物，而不需要血液培养。新的平台将基于单分子扫描(SM-SCAN)，这是一种新的分子检测方法，在这种方法中，无需PCR扩增即可检测到与目标核酸的探针杂交(专利正在申请中)。SM-Scanning使用直接且廉价的检测策略。初步结果表明，SM-Scan可以检测到5个大肠杆菌细胞。在这项提议中，我们将通过开发一种能够在不到1.5小时内以极高的灵敏度检测血液样本中的微生物的方法来展示SM-SCAN的特性。目的1致力于建立一种快速检测5ml全血标本中细菌存在的高灵敏度方法。目的2建立一种能够同时检测细菌和真菌的多重检测方法，并对其性能进行测定。

 DESCRIPTION (provided by applicant): Sepsis is a whole-body inflammatory response to infection that affects approximately 1 million people each year in the United States and kills up to 50 percent of severely affected patients. Rapid and appropriate antimicrobial administration is critical for the management of sepsis. Each hour of treatment delay is associated with an 8% decrease of survival in septic shock patients. Blood culture is the gold standard diagnostic method, however, it takes more than one day to provide results. Therefore, patients receive broad-spectrum antibiotics or may not receive treatment in the initial hours. Molecular detection is an attractive solution, however, currently available tests are based on PCR amplification of bacterial genomic DNA followed by complex detection strategies. These techniques suffer from DNA contamination, lack sufficient sensitivity and are expensive. Here, we propose the development of a novel platform for rapid and inexpensive identification of microorganisms in whole blood without blood culture. The new platform will be based on Single Molecule Scanning (SM-Scanning), a novel molecular detection method in which probe hybridization to a target nucleic acid is detected without PCR amplification (Patent Pending). SM-Scanning uses a straight forward and inexpensive detection strategy. Preliminary results demonstrate that SM-Scanning can detect 5 Escherichia coli cells. In this proposal, we will demonstrate the properties of SM-Scanning by developing an assay capable of detecting microorganisms in blood samples in less than 1.5 hours with extremely high sensitivity. Aim 1 focuses on developing a rapid test capable of detecting the presence of bacteria in 5 ml whole blood samples with high sensitivity. Aim 2 focuses on developing and determining the performance of a multiplex assay capable of detecting bacteria and fungi in a single assay.