Detection of Antibiotic Resistance Genes in Bacterial Agents of Hospital-Acquired

描述(由申请人提供)：医院获得性肺炎(HAP)和败血症是危重病人，特别是在医院重症监护病房中发病率和死亡率的主要原因。这些急性疾病是由医院环境中常见的一系列革兰氏阳性和革兰氏阴性细菌感染引起的。疑似HAP或败血症患者的治疗通常是基于推定证据开始的。病原体的存在和身份通常要等到培养结果出来(通常是在获得样本后几天)才能确定，然后进一步推迟完成抗生素敏感性测试。因此，广谱抗生素通常是默认使用的。然而，越来越多的细菌病原体对这些抗生素表现出耐药性，传统的检测方法可能会遗漏临床相关的耐药性，导致治疗失败和增加患者死亡率。因此，尽早确定引起HAP或脓毒症的细菌的身份及其耐药性特征对于做出治疗决定以及修改或降低抗生素的使用至关重要。拟议项目的目的是开发一种分子检测方法，用于同时检测和鉴定微生物和抗生素耐药性的遗传决定因素，包括革兰氏阴性菌对β-内酰胺类和氟喹诺酮类药物的耐药性，以及革兰氏阳性菌对甲氧西林和万古霉素的耐药性。诊断测试将基于研究人员开发的反向线条印迹分析，该方法能够检测和鉴定一系列常见的细菌病原体。与基于培养的方法需要2-4天的延迟相比，它将通过直接从临床样本中获得遗传信息来提供当天的周转，而不需要细菌生长。在第一阶段，将根据从两个医院的重症监护病房获得的疑似HAP或脓毒症患者的呼吸道和血液样本开发该分析。反向线印迹法的结果将与传统的基于培养的ID和药敏试验结果进行比较。在第二阶段，将完成该检测的商业版本，并将在一项前瞻性研究中评估其在预测抗生素耐药表型方面的准确性，从而潜在地改善医院重症监护病房中受影响患者的治疗决策和医疗结果。总体而言，这一新的分子诊断分析将提供一个强有力的工具，通过适当的抗菌剂使用和感染控制措施，努力改善治疗并限制耐药细菌的出现和传播。公共卫生相关性：细菌感染引起的医院获得性肺炎(HAP)和败血症是医院重症监护病房发病率和死亡率的主要原因，许多细菌出现抗生素耐药性是有效治疗的主要威胁。这一应用将导致开发一种快速检测方法，用于同时鉴定引起这些疾病的细菌种类和耐药基因。比现在更早地检测到这种细菌及其耐药性，将使临床医生能够选择更有效的抗生素治疗，提高受影响患者的存活率，并阻止抗药性细菌的传播。

DESCRIPTION (provided by applicant): Hospital-acquired pneumonia (HAP) and sepsis are leading causes of morbidity and mortality in critically ill patients, especially in hospital intensive care units. These acute conditions are caused by infection with a range of Gram-positive and Gram-negative bacteria commonly found in the hospital environment. Treatment of patients with suspected HAP or sepsis is often initiated based on presumptive evidence. The presence and identity of the pathogen are generally not established until culture results are available (usually days after samples are obtained), followed by a further delay for completion of antibiotic susceptibility testing. Therefore broad-spectrum antibiotics are commonly administered by default. However, an increasing proportion of bacterial pathogens exhibit drug resistance to these antibiotics, and conventional testing may miss clinically relevant resistances, resulting in treatment failures and increased patient mortality. Thus, determining as early as possible both the identity of bacteria causing HAP or sepsis and their drug resistance profiles is critical to making treatment decisions, and modifying or de-escalating antibiotic usage. The aim of the proposed project is to develop a molecular test for concurrent detection and identification of microorganisms and genetic determinants of antibiotic resistance, including resistance to ¿-lactams and fluoroquinolones in Gram-negative bacteria, and to methicillin and vancomycin in Gram-positive bacteria. The diagnostic test will be based on a reverse line blot assay developed by investigators, which enables detection and identification of a wide range of common bacterial pathogens. In contrast with the 2-4 day delay required for culture-based methods, it will offer same-day turnaround by obtaining genetic information directly from clinical samples without need for bacterial growth. In Phase I, the assay will be developed based on respiratory and blood samples from patients with suspected HAP or sepsis obtained from intensive care units at two hospital sites. Results of the reverse line blot method will be compared with those of conventional culture-based ID and antibiotic susceptibility testing. In Phase II, a commercial version of the assay will be completed and its accuracy in predicting antibiotic resistant phenotypes, and thereby potentially improving treatment decisions and healthcare outcomes for affected patients in hospital intensive care units, will be evaluated in a prospective study. Overall, this novel molecular diagnostic assay will provide a powerful tool in efforts to improve treatment and limit the emergence and spread of drug resistant bacteria by appropriate antimicrobial use and infection control measures. PUBLIC HEALTH RELEVANCE: Hospital-acquired pneumonia (HAP) and sepsis resulting from bacterial infection are leading causes of morbidity and mortality in hospital intensive care units, and the emergence of antibiotic resistance in many bacteria is a major threat to effective treatment. This application will result in the development of a rapid test for simultaneous identification of bacterial species and drug resistance genes in bacteria causing these conditions. Earlier detection of the bacteria and their antibiotic resistances than is now possible will enable clinicians to select more effective antibiotic therapy, improving the survival rate for affected patients and deterring the spread of drug resistant bacteria.