Yamamoto Norihisa, Hamaguchi Shigeto, Akeda Yukihiro, Santanirand Pitak, Kerdsin Anusak, Seki Masafumi, Ishii Yoshikazu, Paveenkittiporn Wantana, Bonomo Robert A, Oishi Kazunori, Malathum Kumthorn, Tomono Kazunori
Division of Infection Control and Prevention, Osaka University Graduate School of Medicine, Osaka, Japan; Laboratory of Clinical Research on Infectious Diseases, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.
Faculty of Medicine Ramathibodi Hospital, Mahidol University, Bangkok, Thailand.
PLoS One. 2015 Jul 28;10(7):e0133204. doi: 10.1371/journal.pone.0133204. eCollection 2015.
Healthcare-associated infections are a leading cause of morbidity and mortality worldwide. Treatment is increasingly complicated by the escalating incidence of antimicrobial resistance. Among drug-resistant pathogens, carbapenem-resistant Acinetobacter baumannii (CRAb) is of increasing concern because of the limited applicable therapies and its expanding global distribution in developed countries and newly industrialized countries. Therefore, a rapid detection method that can be used even in resource-poor countries is urgently required to control this global public health threat. Conventional techniques, such as bacterial culture and polymerase chain reaction (PCR), are insufficient to combat this threat because they are time-consuming and laborious. In this study, we developed a loop-mediated isothermal amplification (LAMP) method for detecting blaOXA-23-positive CRAb, the most prevalent form of CRAb in Asia, especially in Thailand, and confirmed its efficacy as a surveillance tool in a clinical setting. Clinical samples of sputum and rectal swabs were collected from patients in a hospital in Bangkok and used for LAMP assays. After boiling and centrifugation, the supernatants were used directly in the assay. In parallel, a culture method was used for comparison purposes to evaluate the specificity and sensitivity of LAMP. As a first step, a total of 120 sputum samples were collected. The sensitivity of LAMP was 88.6% (39/44), and its specificity was 92.1% (70/76) using the culture method as the "gold standard". When surveillance samples including sputum and rectal swabs were analyzed with the LAMP assay, its sensitivity was 100.0%. This method enables the direct analysis of clinical specimens and provides results within 40 minutes of sample collection, making it a useful tool for surveillance even in resource-poor countries.
医疗保健相关感染是全球发病和死亡的主要原因。抗菌药物耐药性的不断上升使治疗变得越来越复杂。在耐药病原体中,耐碳青霉烯类鲍曼不动杆菌(CRAb)因其适用治疗方法有限且在发达国家和新兴工业化国家的全球分布不断扩大而日益受到关注。因此,迫切需要一种即使在资源匮乏国家也能使用的快速检测方法来控制这一全球公共卫生威胁。传统技术,如细菌培养和聚合酶链反应(PCR),不足以应对这一威胁,因为它们既耗时又费力。在本研究中,我们开发了一种环介导等温扩增(LAMP)方法来检测blaOXA - 23阳性CRAb,这是亚洲尤其是泰国最常见的CRAb形式,并在临床环境中证实了其作为监测工具的有效性。从曼谷一家医院的患者中收集痰液和直肠拭子的临床样本,并用于LAMP检测。经过煮沸和离心后,上清液直接用于检测。同时,为了评估LAMP的特异性和敏感性,采用培养方法进行比较。第一步,共收集了120份痰液样本。以培养方法作为“金标准”,LAMP的敏感性为88.6%(39/44),特异性为92.1%(70/76)。当用LAMP检测分析包括痰液和直肠拭子在内的监测样本时,其敏感性为100.0%。该方法能够直接分析临床标本,并在样本采集后40分钟内提供结果,使其成为即使在资源匮乏国家也有用的监测工具。