Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Ragama, Sri Lanka.
Department of Medical Microbiology, Faculty of Medicine, University of Kelaniya, Ragama, Sri Lanka.
PLoS One. 2024 Mar 15;19(3):e0295287. doi: 10.1371/journal.pone.0295287. eCollection 2024.
Leptospirosis is the most widespread zoonosis in the world. The disease is more prevalent in tropical regions where the majority of developing countries are located. Leptospirosis is considered a protean manifestation zoonosis with severity of the disease ranging from a mild febrile illness to a severe and life-threatening illness. Clinical symptoms of leptospirosis overlap with other tropical febrile illnesses. Early, rapid, and definitive diagnosis is important for effective patient management. Since Polymerase Chain Reaction (PCR)-based assays are not readily available in most clinical settings, there is a need for an affordable, simple, and rapid diagnostic test. Quantitative PCR (qPCR) and Recombinase Polymerase Amplification (RPA) were implemented at the Faculty of Medicine, University of Kelaniya, and a prospective study to evaluate RPA for diagnosis of acute phase of leptospirosis was conducted. Results indicate that RPA and qPCR were positive in 81% (98/121) of the total positive and acute clinical samples. Of the 81 positive MAT confirmed patients 60 (74%) and 53 (65%) were positive with qPCR and RPA respectively. Retrospective evaluation revealed a high diagnostic accuracy (sensitivity-70% and specificity-87%) of RPA compared to MAT as the reference gold standard. Results further suggest that there is no significant difference between the two assays, qPCR and RPA-SwiftX (P = 0.40). Laboratory procedures for the extraction and detection by qPCR in the laboratory have been optimized to obtain results within 6 hours. However, the RPA-SwiftX method under field conditions took 35 minutes. The RPA-SwiftX method could replace the qPCR which shows similar sensitivity and specificity. Therefore, RPA established under the current study presents a powerful tool for the early and rapid diagnosis of leptospirosis at point-of-care.
钩端螺旋体病是世界上最广泛流行的动物源性传染病。这种疾病在热带地区更为普遍，而热带地区大多数都是发展中国家。钩端螺旋体病被认为是一种表现形式多样的动物源性传染病，其疾病的严重程度从轻度发热疾病到严重且危及生命的疾病不等。钩端螺旋体病的临床症状与其他热带发热疾病重叠。早期、快速和明确的诊断对于有效的患者管理非常重要。由于聚合酶链反应（PCR）为基础的检测在大多数临床环境中不可用，因此需要一种负担得起、简单和快速的诊断测试。定量 PCR（qPCR）和重组酶聚合酶扩增（RPA）在凯拉尼亚大学医学院实施，并且进行了一项前瞻性研究，以评估 RPA 用于诊断钩端螺旋体病的急性期。结果表明，在总共 121 个阳性和急性临床样本中，RPA 和 qPCR 分别有 81%（98/121）为阳性。在 81 例 MAT 确诊的阳性患者中，60 例（74%）和 53 例（65%）分别用 qPCR 和 RPA 检测为阳性。回顾性评估显示，与 MAT 作为参考金标准相比，RPA 的诊断准确性较高（敏感性为 70%，特异性为 87%）。结果进一步表明，两种检测方法，qPCR 和 RPA-SwiftX 之间没有显著差异（P = 0.40）。在实验室中已经优化了 qPCR 的提取和检测实验室程序，以在 6 小时内获得结果。然而，在现场条件下，RPA-SwiftX 方法需要 35 分钟。RPA-SwiftX 方法可以替代 qPCR，其显示出相似的敏感性和特异性。因此，在当前研究中建立的 RPA 为在护理点进行钩端螺旋体病的早期和快速诊断提供了一种强大的工具。
