Volatile Metabolite-Based Detection of Clostridium difficile Infection

 描述(申请人提供)：艰难梭菌感染(CDI)是全球医院内腹泻和死亡的主要原因，在过去十年中发病率和严重性急剧上升，与耐甲氧西林金黄色葡萄球菌不相上下，甚至超过耐甲氧西林金黄色葡萄球菌，成为美国最常见的与医疗保健相关的感染。减少CDI在医院内传播的一个关键障碍是延迟及时识别CDI病例，随后推迟启动适当的抗菌治疗，并实施接触预防和隔离措施，以减少CDI的水平传播。尽管以实验室为基础的快速CDI检测取得了进展，但临床症状和粪便样本出现之间仍有2-5天的延迟，样本收集和可操作的检测结果之间还有0.5-3.5天的延迟。由于预期会延迟获得诊断结果，许多供应商凭经验开出CDI抗菌治疗处方，导致最终CDI检测为阴性的患者频繁和不必要地接受治疗。显然，对能够缩短临床症状出现和CDI准确识别之间的间隔的诊断方法的需求尚未得到满足。为了解决这一未得到满足的需求，我们提出了一种基于检测改变的CDI微生物组的挥发性代谢组的CDI诊断的新方法。艰难梭菌在抗生素破坏的肠道微生物群中繁衍生息，并发出一种独特的气味，可以通过人类和狗的嗅觉检测到。在一项先导性研究中，我们使用分析化学技术评估了疑似CDI患者的粪便样本的挥发性代谢组，并发现了CDI患者和非CDI患者的粪便挥发性代谢组的主要差异。我们提出两个并行执行的目标，以检验这一假设，即改变的CDI肠道微生物组具有独特的挥发性代谢产物谱，不同于其他抗生素相关性腹泻患者的谱，可用于在医院环境中识别CDI患者。使用热解吸GC-MS/MS来捕获和鉴定这些痕量挥发性代谢物，并使用有监督的学习方法-支持向量机、微阵列分析的预测分析和随机森林-来分析这个高维数据矩阵，我们将解开CDI的复杂卷曲，(1)识别CDI住院患者与其他抗生素相关腹泻患者的粪便样本的特征挥发性代谢物特征，以及(2)确定CDI患者住院环境中与其他抗生素相关腹泻患者相比的环境空气样本中的CDI挥发性代谢物特征。这项研究的最终目标是推导和验证CDI在粪便样本和患者环境中的挥发性代谢特征，为一种新型的CDI检测奠定基础，该检测可以与用于床边识别CDI的护理点式气体传感器系统相结合，减少诊断的延误，并减少这种常见的、高度病态的和危及生命的感染的医院传播。

 DESCRIPTION (provided by applicant): Clostridium difficile infection (CDI) is a major cause of nosocomial diarrhea and mortality worldwide, increasing dramatically in incidence and severity over the past decade and rivaling or exceeding methicillin-resistant Staphylococcus aureus as the most common healthcare-associated infection in the US. A critical barrier to reducing the nosocomial spread of CDI is the delay in timely identification of CDI cases, with subsequent delays in initiation of appropriate antimicrobial therapy and implementation of contact precautions and isolation measures to reduce horizontal CDI transmission. Despite advances in rapid laboratory-based CDI testing, there are still delays of 2-5 days between the onset of clinical symptoms and stool sampling and another 0.5-3.5 days between sample collection and actionable test results. Anticipating a delay in obtaining diagnostic results, many providers prescribe CDI antimicrobial therapy empirically, resulting in frequent and unnecessary treatment of patients who ultimately test negative for CDI. There is clearly an unmet need for diagnostic methods that can reduce this interval between onset of clinical symptoms and accurate identification of CDI. To address this unmet need, we propose a novel approach to CDI diagnosis based on detection of the volatile metabolome of the altered CDI microbiome. C. difficile flourishes in the antibiotic- disrupted intestinal microbiome and emits a distinctive odo detectable by human and canine olfaction. In a pilot study, we assessed the volatile metabolome of stool specimens from patients with suspected CDI using analytical chemistry techniques and discerned major differences in the stool volatome of patients with and without CDI. We propose two aims executed in parallel, to test the hypothesis that the altered CDI intestinal microbiome has a unique volatile metabolite profile, distinct from the profile of patiens with other causes of antibiotic-associated diarrhea, which can be used to identify patients with CDI in the hospital setting. Using thermal desorption GC-MS/MS to capture and identify these trace volatile metabolites and supervised learning methods - support vector machines, prediction analysis for microarray analysis, and random forests - to analyze this high-dimensional data matrix, we will deconvolute the complex volatome of CDI, (1) identifying the characteristic volatile metabolite profile of stool samples from hospitalized patients with CDI, compared to patients with other antibiotic-associated diarrhea, and (2) defining the CDI volatile metabolite profile in ambient air samples from the inpatient environment of patients with CDI, compared to patients with other antibiotic-associated diarrhea. The ultimate objective of this research is to derive and validate the volatile metabolic signature of CDI in stool samples and in the patient's environment, laying the groundwork for a novel CDI assay that can be coupled to a point-of-care gas sensor system for the bedside identification of CDI, reducing delays in diagnosis and decreasing nosocomial transmission of this common, highly morbid, and life- threatening infection.