Non-lethal diagnostic methods for koi herpesvirus in koi Cyprinus carpio

Cyprinid herpesvirus 3, also known as koi herpesvirus (KHV), is a viral pathogen responsible for mass mortalities of carp worldwide. In this study, we compared the sensitivity and specificity of ELISA and quantitative PCR (qPCR) methods for the diagnosis of KHV in experimentally infected koi Cyprinus carpio over an 11 mo period. Koi were exposed to KHV at 18 ± 1°C (permissive temperatures for KHV disease) in laboratory-controlled conditions. At 21 d post challenge, the temperature in the system was decreased to <15°C (non-permissive temperature for KHV disease), and fish were monitored for the following 11 mo. At different time points throughout the study, samples of blood and gills were collected from exposed and control koi and subjected to qPCR and ELISA. Survival proportions of 53.3 and 98.8% in exposed and control treatments, respectively, were recorded at the end of the challenge. Traditional receiver-operating characteristic analysis was used to compare the sensitivity of the ELISA and blood and gill qPCR during permissive and non-permissive temperatures. ELISA was superior to qPCR of gills and whole-blood samples in detecting previous exposure to KHV. Similar results were obtained in a second experiment exposing koi to KHV and inducing persistent infection at >30°C (non-permissive temperature for KHV disease). Finally, KHV ELISA specificity was confirmed using cyprinid herpesvirus 1-exposed koi through a period of 3 mo. This study demonstrates that the combination of ELISA and gill qPCR should be recommended in the diagnosis of KHV exposure of suspected carrier-state fish.