Natural outbreak of viral encephalopathy and retinopathy in juvenile sea bass, Dicentrarchus labrax: study by nested reverse transcriptase-polymerase chain reaction.
In order to improve the sensitivity of the diagnosis of viral encephalopathy and retinopathy (VER) in sea bass, a nested reverse transcriptase-polymerase chain reaction (RT-PCR) detection method was developed. The reverse transcription step and the first stage PCR were performed using outer primers specific for the coat protein gene, whereas a new primer set was used as inner primers for the second stage PCR. Fish were collected just before, during and after a VER outbreak occurring in a mediterranean fish farm. For each time point, ten different fish were analysed individually by nested RT-PCR, single step PCR and virus cultivation. The results showed that the frequency of positive samples was always higher using the nested RT-PCR assay. In particular, it was possible to detect nodavirus specific signals 1 month before the appearance of the first mortalities, but only by nested RT-PCR. Altogether these results showed that the sensitivity of nodavirus detection is greatly improved using a nested RT-PCR method. In particular, it was possible to monitor the presence of viral genome in asymptomatic carrier fish using this method.
Thiery
R
R
CNEVA Brest, Laboratoire de Pathologie des Animaux Aquatiques, Technopôle Brest-Iroise, Plouzané, France. rthiery@ifremer.fr
Raymond
J C
JC
Castric
J
J
D000818:Animals; D001492:Bass / Q000821:virology*; D001921:Brain / Q000821:virology; D004668:Encephalitis Viruses / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D004671:Encephalitis, Arbovirus / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D005393:Fish Diseases / Q000175:diagnosis / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012160:Retina / Q000821:virology; D012164:Retinal Diseases / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity