Evaluation of Chikungunya Diagnostic Assays: Differences in Sensitivity of Serology Assays in Two Independent Outbreaks
Evaluation of Chikungunya Diagnostic Assays: Differences in Sensitivity of Serology Assays in Two Independent Outbreaks
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DOI：
10.1371/journal.pntd.0000753
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发表时间：
2010-07-01
影响因子：
3.8
通讯作者：
Ng, Lee-Ching
中科院分区：
文献类型：
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作者：
Yap, Grace;Pok, Kwoon-Yong;Ng, Lee-Ching
背景：测试了两种内部 MAC-ELISA 检测的灵敏度和特异性，并与市售 CTK 侧流快速检测和 EUROIMMUN IFA 检测抗基孔肯雅病毒 (CHIKV) IgM 检测的性能进行了比较。每个 MAC-ELISA 检测均使用基于全病毒的抗原，该抗原源自新加坡两次基孔肯雅病爆发中涉及的遗传上不同的 CHIKV 毒株（2008 年）； 1 月爆发菌株 E1 糖蛋白 226 氨基酸残基处含有丙氨酸 (CHIKV-A226)，5 月至 9 月爆发菌株在相同残基处含有缬氨酸 (CHIKV-226V)。我们报告了两次疫情之间不同检测方法 IgM 检测效果的差异。还测试了两种 PCR 方案的敏感性。 方法和结果：对于 1 月份爆发的血清，CTK 侧流检测、MAC-ELISA 和 EUROIMMUN IFA 检测的平均检测阈值分别为发烧后 3.75、4.38 和 4.88 天。相比之下，在第二次疫情血清中，使用 CTK 侧流检测的 IgM 检测延迟到发烧后 7 天以上。然而，使用 CHIKV-226V 的 MAC-ELISA 在发烧后 3.96 天检测到第二次爆发血清中的 IgM，这比使用 CHIKV-A226 的相同检测（4.86 天）早了大约一天。两种商业化验的特异性均为 100%，内部 MAC-ELISA 的特异性为 95.6%。对于 PCR 方案的灵敏度测定，基于探针的实时 RT-PCR 方法被发现比基于 SYBR Green 的方法灵敏 10 倍。结论：我们的研究结果表明，使用变体 A226 和 226V 的两种 CHIKV 菌株导致所评估测定的灵敏度发生变化。我们假设观察到的抗原功效差异可能是由于病毒 E1 和 E2 包膜蛋白的氨基酸取代差异所致，尤其是 E1-A226V 取代。该评估证明了在临床和操作环境中应用不同诊断分析之前对其进行评估的重要性。
Background: The sensitivity and specificity of two in-house MAC-ELISA assays were tested and compared with the performance of commercially-available CTK lateral flow rapid test and EUROIMMUN IFA assays for the detection of anti-Chikungunya virus (CHIKV) IgM. Each MAC-ELISA assay used a whole virus-based antigen derived from genetically distinct CHIKV strains involved in two chikungunya disease outbreaks in Singapore (2008); a January outbreak strain with alanine at amino acid residue 226 of the E1 glycoprotein (CHIKV-A226) and a May-to-September outbreak strain that possessed valine at the same residue (CHIKV-226V). We report differences in IgM detection efficacy of different assays between the two outbreaks. The sensitivities of two PCR protocols were also tested.Methods and Findings: For sera from January outbreak, the average detection threshold of CTK lateral flow test, MAC-ELISAs and EUROIMMUN IFA assays was 3.75, 4.38 and 4.88 days post fever onset respectively. In contrast, IgM detection using CTK lateral flow test was delayed to more than 7 days after fever onset in the second outbreak sera. However, MAC-ELISA using CHIKV-226V detected IgM in the second outbreak sera 3.96 days after fever onset, which was approximately one day earlier compared to the same assay using CHIKV-A226 (4.86 days). Specificity was 100% for both commercial assays, and 95.6% for the in-house MAC-ELISAs. For sensitivity determination of the PCR protocols, the probe-based real time RT-PCR method was found to be 10 times more sensitive than one based on SYBR Green.Conclusion: Our findings suggested that the two strains of CHIKV using variants A226 and 226V resulted in variation in sensitivities of the assays evaluated. We postulated that the observed difference in antigen efficacy could be due to the amino acid substitution differences in viral E1 and E2 envelope proteins, especially the E1-A226V substitution. This evaluation demonstrates the importance of appraisal of different diagnostic assays before their application in clinical and operational settings.