Schwarzer Anne, Ziegler Ute, Fertey Jasmin, Kreuz Markus, Vahlenkamp Thomas W, Groschup Martin H, Ulbert Sebastian
Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Friedrich-Loeffler Institut, Südufer 10, 17493, Greifswald-Insel Riems, Germany.
Fraunhofer Institute for Cell Therapy and Immunology, Department of Infection Research and Diagnostics, Perlickstraße 1, 04103, Leipzig, Germany.
Sci Rep. 2025 Aug 6;15(1):28752. doi: 10.1038/s41598-025-14448-4.
West Nile virus (WNV), Usutu virus (USUV) and tick-borne encephalitis virus (TBEV) are worldwide endemic zoonotic orthoflaviviruses, often co-circulating in the same areas. Serological studies in animals, mostly birds and horses, are important means to monitor the spread of these viruses and the infection risks for humans. However, cross-reactive antibodies to these structurally similar flaviviruses frequently impact serological differentiation in enzyme-linked immunosorbent assays (ELISAs), hence time-consuming virus neutralization tests (VNTs) have to be employed in laboratories with high biosafety level. This study presents ELISAs using recombinant flavivirus E proteins with point mutations in the conserved fusion loop domain (Equad proteins) for differentiating IgY or IgG antibodies against WNV, USUV or TBEV in ducks, geese, chickens and horses. Panels of 169 duck and goose sera, 101 chicken sera and 136 horse sera were tested in Equad ELISAs, which resulted in high sensitivity and specificity, further improved by a pre-absorption step for the differentiation of WNV and USUV antibodies. Equad ELISAs for poultry and horse sera enable the reliable differentiation of WNV, USUV and TBEV specific antibodies without the need for VNTs, which has important implications for conducting seroprevalence studies as well as for veterinary routine diagnosis.
西尼罗河病毒（WNV）、乌苏图病毒（USUV）和蜱传脑炎病毒（TBEV）是全球地方性人畜共患的正黄病毒，常于同一地区共同传播。对动物（主要是鸟类和马匹）进行的血清学研究是监测这些病毒传播及人类感染风险的重要手段。然而，针对这些结构相似的黄病毒的交叉反应性抗体常常影响酶联免疫吸附测定（ELISA）中的血清学鉴别，因此在具有高生物安全水平的实验室中必须采用耗时的病毒中和试验（VNT）。本研究展示了利用在保守融合环结构域具有点突变的重组黄病毒E蛋白（Equad蛋白）的ELISA，用于鉴别鸭、鹅、鸡和马体内针对WNV、USUV或TBEV的IgY或IgG抗体。用Equad ELISA检测了169份鸭和鹅血清、101份鸡血清以及136份马血清样本，结果显示其具有高灵敏度和特异性，通过预吸收步骤进一步提高了对WNV和USUV抗体的鉴别能力。针对家禽和马血清的Equad ELISA能够可靠地区分WNV、USUV和TBEV特异性抗体，无需进行VNT，这对开展血清流行率研究以及兽医常规诊断具有重要意义。
