印度尼西亚西爪哇省内部登革热实时 PCR 检测方法的评估。

Evaluation of in-house dengue real-time PCR assays in West Java, Indonesia.

机构信息

School of Life Sciences and Technology, Institute of Technology Bandung, Bandung, West Java, Indonesia.

Department of Entomology, National Chung Hsing University, Taichung, Taichung, Taiwan.

出版信息

PeerJ. 2024 Jul 24;12:e17758. doi: 10.7717/peerj.17758. eCollection 2024.

DOI:10.7717/peerj.17758

PMID:39071132

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11283174/

Abstract

Dengue is an infectious disease caused by infection of dengue virus (DENV) transmitted by and . In Indonesia, dengue commonly occurs with an increasing incidence rate annually. It is known that early detection of dengue infection is one of the keys to controlling this disease outbreak. Rapid and accurate early detection to diagnose dengue can be achieved by molecular tests, one of which is through a real-time PCR method. However, real-time PCR assay for dengue developed based on Indonesian DENV sequences has not been available. Therefore, we developed in-house dengue real-time PCR (SYBR- and TaqMan-based) assays and evaluated those assays in routine clinical testing in the community. These assays target the 3' UTR region of the four DENV serotypes and was found to be specific for DENV. The most sensitive assay was the TaqMan assay with the LOD of 482 copy/ml, followed by the SYBR assay with the LOD of 14,398 copy/ml. We recruited dengue suspected patients from three primary health care services in West Java, Indonesia to represent the community testing setting. Dengue infection was examined using the two in-house real-time PCR assays along with NS1, IgM, and IgG rapid diagnostic tests (RDT). In total, as many as 74 clinical specimens of dengue suspected patients were included in this study. Among those patients, 21 were positive for TaqMan assay, 17 were positive for SYBR assay, nine were positive for NS1 test, six were positive for both IgG and IgM tests, and 22 were positive for IgG test only. Compared with our in-house TaqMan assay, the sensitivity of NS1 test, IgM test, and IgG test were 42.86%, 14.29%, and 28.57% respectively. Among these three RDT tests, NS1 showed 100% specificity. Thus, our study confirmed that NS1 test showed high specificity, indicating that a positive result of NS1 can be confidently considered a dengue case. However, NS1, IgM, and IgG tests with RDT are not enough to diagnose a dengue case. We suggest applying the high sensitivity and specificity rRT-PCR test as the gold standard for early detection and antibody test as a follow-up test for rRT-PCR negative cases.

摘要

登革热是一种由登革病毒（DENV）感染引起的传染病，通过和传播。在印度尼西亚，登革热的发病率每年都在增加。众所周知，早期发现登革热感染是控制这种疾病爆发的关键之一。通过分子检测可以快速准确地早期检测到登革热，其中一种方法是实时 PCR 方法。然而，基于印度尼西亚 DENV 序列开发的实时 PCR 检测方法尚未问世。因此，我们开发了内部的登革热实时 PCR（基于 SYBR 和 TaqMan）检测方法，并在社区的常规临床检测中评估了这些方法。这些检测方法针对四种 DENV 血清型的 3'UTR 区域，并且对 DENV 具有特异性。最敏感的检测方法是 TaqMan 检测法，其检测限为 482 拷贝/ml，其次是 SYBR 检测法，检测限为 14398 拷贝/ml。我们从印度尼西亚西爪哇的三个初级保健服务机构招募了登革热疑似患者，以代表社区检测环境。使用两种内部实时 PCR 检测方法以及 NS1、IgM 和 IgG 快速诊断检测（RDT）检测登革热感染。共有 74 例登革热疑似患者的临床标本纳入本研究。在这些患者中，21 例 TaqMan 检测法阳性，17 例 SYBR 检测法阳性，9 例 NS1 检测法阳性，6 例 IgG 和 IgM 检测法均阳性，22 例仅 IgG 检测法阳性。与我们的内部 TaqMan 检测法相比，NS1 检测法、IgM 检测法和 IgG 检测法的灵敏度分别为 42.86%、14.29%和 28.57%。在这三种 RDT 检测中，NS1 检测法的特异性为 100%。因此，我们的研究证实 NS1 检测法具有很高的特异性，表明 NS1 检测阳性可被认为是登革热病例。然而，NS1、IgM 和 IgG 检测法的 RDT 不足以诊断登革热病例。我们建议将高灵敏度和特异性 rRT-PCR 检测作为早期检测的金标准，将抗体检测作为 rRT-PCR 阴性病例的后续检测。