Yun Tao, Chen Haipeng, Yu Bin, Zhang Cun, Chen Liu, Ni Zheng, Hua Jionggang, Ye Weicheng
Institute of Animal Husbandry and Veterinary Sciences, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China.
College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Tai'an 271018, China.
J Virol Methods. 2015 Aug;220:55-9. doi: 10.1016/j.jviromet.2015.04.012. Epub 2015 Apr 20.
A novel duck reovirus (N-DRV) disease emerged in China in 2000 and it has become an epidemic genotype. A test for detection of virus-specific antibodies in serum samples would be useful for epidemiological investigations. Currently, Currently, serological assays for N-DRV diagnosis are not available. A test for detection of virus-specific antibodies in serum samples would be useful for epidemiological investigations. In this study, a highly sensitive and specific indirect enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to N-DRV was developed. The outer capsid (σC) of N-DRV was cloned and expressed in Escherichia coli as a coating antigen. The antigen concentration and serum dilution were optimized using a checkerboard titration. Furthermore, the specificity of σC-ELISA assay was confirmed by cross checking with other duck viral pathogens. In comparison with the western blot, the sensitivity and specificity of the σC-ELISA was 92.6% and 88.9%, respectively, and agreement of two tests was excellent with κ value of 0.786 (p < 0.05). A serological survey was performed using the assay on serum samples from different age and species of duck flocks in the Zhejiang and Jiangsu Province, China. The seropositive rate of the 1209 serum samples was 57.7%. In conclusion, the developed σC-ELISA assay is a very specific and sensitive test that will be useful for large-scale serological survey in N-DRV infection and monitoring antibodies titers against N-DRV.
2000年，一种新型鸭呼肠孤病毒（N-DRV）疾病在中国出现，并已成为一种流行基因型。检测血清样本中病毒特异性抗体的试验对于流行病学调查将是有用的。目前，尚无用于N-DRV诊断的血清学检测方法。检测血清样本中病毒特异性抗体的试验对于流行病学调查将是有用的。在本研究中，开发了一种用于检测N-DRV抗体的高度敏感和特异的间接酶联免疫吸附测定（ELISA）。将N-DRV的外衣壳（σC）克隆并在大肠杆菌中表达作为包被抗原。使用棋盘滴定法优化抗原浓度和血清稀释度。此外，通过与其他鸭病毒病原体交叉检测证实了σC-ELISA试验的特异性。与western blot相比，σC-ELISA的敏感性和特异性分别为92.6%和88.9%，两种检测方法的一致性极佳，κ值为0.786（p<0.05）。使用该检测方法对中国浙江省和江苏省不同年龄和品种鸭群的血清样本进行了血清学调查。1209份血清样本的血清阳性率为57.7%。总之，所开发的σC-ELISA检测方法是一种非常特异和敏感的检测方法，将有助于大规模血清学调查N-DRV感染情况并监测针对N-DRV的抗体滴度。
