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）和败血症是危重患者发病和死亡的主要原因，特别是在医院重症监护病房。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。通常在获得培养结果（通常在获得样本数天后）之后才能确定病原体的存在和身份，然后进一步延迟完成抗生素敏感性试验。因此，通常默认使用广谱抗生素。然而，越来越多的细菌病原体对这些抗生素表现出耐药性，常规检测可能会遗漏临床相关的耐药性，导致治疗失败和患者死亡率增加。因此，尽早确定引起HAP或败血症的细菌及其耐药谱对于制定治疗决策、调整或减少抗生素使用至关重要。拟议项目的目的是开发一种分子测试，用于同时检测和鉴定抗生素耐药性的微生物和遗传决定因素，包括革兰氏阴性细菌对内酰胺和氟喹诺酮类药物的耐药性，以及革兰氏阳性细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行印迹试验，该试验能够检测和鉴定广泛的常见细菌病原体。与基于培养的方法所需的2-4天延迟相比，它将通过直接从临床样本中获取遗传信息而无需细菌生长而提供当日周转。在第一阶段，该检测方法将基于从两家医院重症监护病房获得的疑似HAP或败血症患者的呼吸和血液样本。将反线印迹法的结果与传统的基于培养的ID和抗生素药敏试验结果进行比较。在二期试验中，将完成该试验的商业版本，并在一项前瞻性研究中评估其预测抗生素耐药表型的准确性，从而潜在地改善医院重症监护病房受影响患者的治疗决策和医疗结果。总之，这种新的分子诊断方法将通过适当的抗菌药物使用和感染控制措施，为改善治疗和限制耐药细菌的出现和传播提供有力的工具。公共卫生相关性：由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和死亡的主要原因，许多细菌出现抗生素耐药性是有效治疗的主要威胁。这一应用将导致一种快速测试的发展，用于同时鉴定细菌种类和引起这些疾病的细菌中的耐药基因。比现在更早地发现细菌及其抗生素耐药性，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，并阻止耐药细菌的传播。

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.