Research note: Improved virus isolation strategies enhance the detection rate for avian leukosis virus eradication in a local chicken breed in China.

The detection rate of avian leukosis virus subgroup K (ALV-K) has been increasing annually in Chinese local breeds, its poor replicative capacity limits the success of virus isolation. To improve ALV-K eradication, this study assessed the detection limits of the magnetic-particle chemiluminescent assay and ELISA kits using a standardized virus stock. Additionally, we also determined the impact of serial passage during virus isolation on the positive detection rate. Using a standardized ALV-K stock, the magnetic-particle chemiluminescent kit detected positive signals at a 256-fold dilution, whereas the most sensitive ELISA kit reached its limit at a 64-fold dilution. When applied to 13 plasma samples from a local chicken flock, the magnetic-particle chemiluminescent kit at passage 0 yielded a detection rate of 76.9% (10/13), compared to only 15.4% (2/13) by ELISA. After a single cell passage, the chemiluminescent method achieved 100% positivity (13/13), while ELISA required two passages to reach 61.5% (8/13). Quantitative real-time PCR (qPCR) confirmed the presence of ALV-K in at least 9 plasma samples, corroborating the chemiluminescent results. These results demonstrate that the magnetic-particle chemiluminescent assay exhibits significantly higher sensitivity than ELISA and that cell passaging markedly enhances the positive rate of virus isolation. By integrating a highly sensitive chemiluminescent assay with an optimized viral isolation protocol, this study provides a novel strategy for facilitating ALV-K eradication in local Chinese chicken flocks.