Detection of infectious pancreatic necrosis virus (IPNV) from asymptomatic redbanded seabream, Pagrus auriga Valenciennes, and common seabream, Pagrus pagrus (L.), using a non-destructive procedure.
A non-destructive procedure based on nested RT-PCR and dot-blot hybridization has been developed for the detection of asymptomatic IPNV-carrier fish. The pair of primers designed for RT-PCR amplified a 599-bp fragment of the pVP2 region within the polyprotein gene, resulting in the detection of IPNV genotype III.1. The use of a nested RT-PCR allowed the amplification of IPNV genotypes III.1 and I.2. In addition, a 191-bp probe was designed for hybridization studies used in combination with the nested RT-PCR. The application of the nested RT-PCR to analyse blood samples from asymptomatic redbanded seabream, Pagrus auriga, and common seabream, P. pagrus, specimens showed a 53.1% and 77.8% prevalence of IPNV-carriers, respectively. The combination of nested RT-PCR and dot-blot hybridization increased the detection rates up to 100% for redbanded seabream and 94.4% for common seabream. Therefore, the protocol described in this study is highly sensitive and specific for the detection of IPNV in asymptomatic carrier fish, and, in addition, the results demonstrate the carrier state in two newly cultured sparid species in southern Spain.
Lopez-Jimena
B
B
Departamento de Microbiologia, Facultad de Ciencias, Universidad de Malaga, 29071 Malaga, Spain.
Garcia-Rosado
E
E
Infante
C
C
Cano
I
I
Manchado
M
M
Castro
D
D
Borrego
J J
JJ
Alonso
M C
MC
D000818:Animals; D018175:Birnaviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D005393:Fish Diseases / Q000175:diagnosis* / Q000821:virology; D018061:Infectious pancreatic necrosis virus / Q000302:isolation & purification*; D025202:Molecular Diagnostic Techniques / Q000379:methods / Q000662:veterinary*; D009693:Nucleic Acid Hybridization / Q000379:methods; D020133:Reverse Transcriptase Polymerase Chain Reaction; D021541:Sea Bream; D012680:Sensitivity and Specificity