Tamba Marco, Caminiti Antonino, Prosperi Alice, Desprès Philippe, Lelli Davide, Galletti Giorgio, Moreno Ana, Paternoster Giulia, Santi Annalisa, Licata Elio, Lecollinet Sylvie, Gelmini Luca, Rugna Gianluca, Procopio Anna, Lavazza Antonio
Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna. Via Antonio Bianchi, 7. 25126 Brescia, Italy.
Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna. Via Antonio Bianchi, 7. 25126 Brescia, Italy.
J Virol Methods. 2017 Oct;248:202-206. doi: 10.1016/j.jviromet.2017.07.010. Epub 2017 Jul 27.
West Nile virus (WNV) and Usutu virus (USUV), genus Flavivirus, are members of the Japanese encephalitis virus antigenic complex, and are maintained primarily in an enzootic cycle between mosquitoes and birds. WNV is zoonotic, and poses a threat to public health, especially in relation to blood transfusion. Serosurveillance of wild birds is suitable for early detection of WNV circulation, although concerns remain to be addressed as regards i) the type of test used, whether ELISA, virus neutralization test (VNT), plaque reduction neutralization test (PRNT), ii) the reagents (antigens, revealing antibodies), iii) the different bird species involved, and iv) potential cross-reactions with other Flaviviruses, such as USUV. The authors developed an indirect IgG ELISA with pan-avian specificity using EDIII protein as antigen and a monoclonal antibody (mAb 1A3) with broad reactivity for avian IgG. A total of 140 serum samples were collected from juvenile European magpies (Pica pica) in areas where both WNV and USUV were co-circulating. The samples were then tested using this in-house ELISA and VNT in parallel. Estimation of test accuracy was performed using different Bayesian two latent class models. At a cut-off set at an optical density percentage (OD%) of 15, the ELISA showed a posterior median of diagnostic sensitivity (DSe) of 88% (95%PCI: 73-99%) and a diagnostic specificity (DSp) of 86% (95%PCI: 68-99%). At this cut-off, ELISA and VNT (cut-off 1/10) performances were comparable: DSe=91% (95%PCI: 79-99%), and DSp=77% (95%PCI: 59-98%). With the cut-off increased to 30 OD%, the ELISA DSe dropped to 78% (95%PCI: 52-99%), and the DSp rose to 94% (95%PCI: 83-100%). In field conditions, the cut-off that yields the best accuracy for the ELISA appears to correspond to 15 OD%. In areas where other Flaviviruses are circulating, however, it might be appropriate to raise the cut-off to 30 OD% in order to achieve higher specificity and reduce the detection of seropositive birds infected by other Flaviviruses, such as USUV.
西尼罗河病毒（WNV）和乌苏图病毒（USUV）属于黄病毒属，是日本脑炎病毒抗原复合物的成员，主要在蚊子和鸟类之间的动物疫源性循环中传播。WNV是人畜共患病原体，对公众健康构成威胁，尤其是在输血方面。野生鸟类的血清学监测适用于早期发现WNV的传播，不过仍有一些问题需要解决，涉及：i）所使用的检测类型，无论是酶联免疫吸附测定（ELISA）、病毒中和试验（VNT）、蚀斑减少中和试验（PRNT）；ii）试剂（抗原、显色抗体）；iii）所涉及的不同鸟类物种；iv）与其他黄病毒（如USUV）的潜在交叉反应。作者开发了一种以E结构域III（EDIII）蛋白为抗原、对鸟类IgG具有广泛反应性的单克隆抗体（mAb 1A3）的具有泛鸟类特异性的间接IgG ELISA。在WNV和USUV共同传播的地区，从幼年欧洲喜鹊（Pica pica）身上采集了总共140份血清样本。然后使用这种内部ELISA和VNT对样本进行平行检测。使用不同的贝叶斯双潜在类别模型进行检测准确性评估。在设定光密度百分比（OD%）为15的临界值时，ELISA显示诊断敏感性（DSe）的后验中位数为88%（95%可信区间：73 - 99%），诊断特异性（DSp）为86%（95%可信区间：68 - 99%）。在此临界值下，ELISA和VNT（临界值1/10）的表现相当：DSe = 91%（95%可信区间：79 - 99%），DSp = 77%（95%可信区间：59 - 98%）。当临界值提高到30 OD%时，ELISA的DSe降至78%（95%可信区间：52 - 99%），DSp升至94%（95%可信区间：83 - 100%）。在野外条件下，ELISA产生最佳准确性的临界值似乎对应于15 OD%。然而，在其他黄病毒传播的地区，可能适合将临界值提高到30 OD%，以实现更高的特异性并减少对感染其他黄病毒（如USUV）的血清阳性鸟类的检测。
