Metabolome-Based Biomarkers and Neutraceuticals in Clostridium Difficile Infection

描述(申请人提供)：艰难梭菌是美国医院感染的主要原因，超过耐药葡萄球菌和肠球菌。艰难梭菌感染的高流行率是医院和长期护理机构面临的一个主要健康问题，这是由于常见的患者风险因素；特别是高龄和共病、抗菌药的使用以及艰难梭菌感染的困难。 孢子消毒。新出现的高毒力艰难梭菌株也促进了CDI的升级，导致疾病严重程度和死亡率增加。此外，35%的患者将经历复发的CDI，这是一个重要的临床问题，往往导致较差的临床结果。尽管CDI的早期诊断对优化临床治疗和改善预后至关重要，但准确预测CDI复发的诊断方法尚不存在，复发的危险因素仍然难以捉摸。CDI复发的频率很高，加上没有得到及时和有效治疗的患者的临床结果很差，这突显了识别准确的疾病复发生物标志物的必要性，这些标志物可以开发成新的诊断方法。这是我们研究的主要目标。尽管已知抗菌素对肠道保护性微生物区系的破坏与感染者症状的发展密切相关，但我们对CDI易感性和复发的理解仍有很大差距。在这一修订的应用中，我们证明了将下一代DNA测序与全球代谢组学整合到微生物生态系统网络中，为发现复发CDI的新诊断和治疗方案提供了一个分析框架。我们的多组学方法的意义在于在继续发展为复发疾病的原发CDI患者中识别候选的16S rRNA和生化生物标记物(误分率为12%)。这些多组学研究在CDI患者中产生了以下新的观察结果：“当使用基于分子的检测来诊断CDI时，发现了一个主要的潜在缺陷。当使用不能区分CDI和无症状艰难梭菌携带者的敏感的核酸扩增测试时，误诊率为54%。”基于我们将γ-氨基丁酸及其合成前体确定为CDI复发的候选生物标志物，服用GABA受体A激动剂唑吡坦的住院患者发生CDI的风险增加了4.88倍。我们的目标1的目标是在200名患者中验证CDI复发的候选16S rRNA生物标志物，并将这些发现与临床元数据相关联。对于目标2，我们期望在相同的200名患者队列中验证CDI复发的候选代谢生物标记物，并进行多组学分析。我们预计，多组生物标志物--单独或联合使用--将准确预测CDI复发。考虑到完全缺乏复发的CDI诊断方法，我们的目标是开发一个或多个生物标志物作为金标准方法，以验证其他能够区分CDI与无症状定植的成本效益分析，以及识别有复发CDI风险的患者。这种类型的诊断代表着优先的临床需求和重大的生物医学市场机会。我们还预计，新采用的核酸扩增试验大大高估了CDI的发病率。我们的初步数据表明，由于明显的误诊，这类患者可能正在接受完全不适当的治疗选择，例如儿童粪便微生物区系移植。

DESCRIPTION (provided by applicant): Clostridium difficile is the leading cause of nosocomial infection in the U.S., outpacing both antibiotic-resistant staphylococcus and enterococcus. The high prevalence of Clostridium difficile infection (CDI) represents a major health problem for hospitals and long-term care facilities alike due to common patient risk factors; notably advanced age and comorbidity, use of antimicrobials and difficulty in C. difficile spore disinfection. Newly-emerged hypervirulent C. difficile strains have also contributed to CDI escalation resulting in increased disease severity and mortality. Furthermore, 35% of patients will experience recurrent CDI, a significant clinical issue that often results in poor clinical outcome. Despite the fact that early diagnosis of CDI is crucial for optimal clinical management and improved prognosis, diagnostic assays that accurately predict CDI recurrence do not exist and risk factors for recurrent CDI remain elusive. The high frequency of CDI recurrence, coupled with poor clinical outcomes for cases not promptly and effectively treated, underscores the need to identify accurate biomarkers of disease recurrence that can be developed into new diagnostic assays. This is the major goal of our study. Although antimicrobial disruption of the protective gut microflora is known to strongly correlate with the development of symptoms in infected individuals, there is still a major gap in our understanding of CDI susceptibility and recurrence. In this revised application, we demonstrate that integration of next generation DNA sequencing with global metabolomics into microbial ecosystem networks provides an analytical framework for the discovery of new diagnostic and treatment options for recurrent CDI. The significance of our multi-omics approach is the identification of candidate 16S rRNA and biochemical biomarkers in primary CDI patients who go on to develop recurrent disease (misclassification rate of 12%). These multi-omic studies led to the following novel observations in CDI patients: "Identification of a major potential deficiency when using molecular-based testing to diagnose CDI. A 54% misdiagnosis rate was indicated when using sensitive nucleic acid amplification tests that cannot discriminate CDI from asymptomatic C. difficile carriage. " Based on our identification of γ-aminobutyric acid (GABA) and precursors of GABA synthesis as candidate biomarkers of CDI recurrence, a 4.88-fold higher CDI risk association was identified in hospitalized patients prescribed Zolpidem (Ambien), a GABA receptor A agonist. Our goal for Aim 1 is to validate candidate 16S rRNA biomarkers of CDI recurrence in a larger 200 patient based cohort and correlate these findings with clinical metadata. For Aim 2, we expect to validate candidate metabolomic biomarkers of CDI recurrence in the same 200 patient cohort and perform multi-omic analysis. We expect that the multi-omic biomarkers - alone or in combination - will accurately predict CDI recurrence. Considering the complete lack of recurrent CDI diagnostic assays, we aim to develop one or more biomarkers for use as a gold standard method that will validate other cost effective assays enabling differentiation of CDI from asymptomatic colonization, as well as identification of patients at risk for recurrent CDI. This type of diagnostic represents a priority clinical need and a major biomedical market opportunity. We also anticipate that newly adopted nucleic acid amplification testing is greatly overestimating CDI incidence. Our preliminary data indicate that such patients are likely receiving wholly inappropriate treatment options resulting from their apparent misdiagnosis e.g. fecal microbiota transplantation in children.