Detection of spring viremia of carp virus isolates by hybridization with non-radioactive probes and amplification by polymerase chain reaction

For detection of spring viremia of carp virus (SVCV) DNA probes have been c onstructed using the reverse transcription-polymerase chain reaction (RT-PC R) amplification technique and cDNA cloning in plasmid and phage vectors. T he specific primers for amplification of SVCV M and G genes were chosen and synthesized. Studies were carried out to establish the sensitivity and spe cificity of viral RNA detection in infected cell culture and pathogenic mat erial from fish by the use of non-radioactive probes and RT-PCR. The effici ency of amplification with primers, complementary to the genome of the refe rence Fijan strain, was estimated in RT-PCR experiments with two SVCV strai ns. Under the same conditions, the quantity of PCR products amplified from the M2 strain was less than that from the ZL4 strain, which implies that th e latter is more similar to the reference European SVCV Fijan isolate. Usin g DNA probes and dot-blot hybridization, SVCV was tested in samples taken f rom different organs of artificially infected carp with clinical signs of a cute disease. The virus could be detected most reliably in fish brain. In m ost cases the hybridization signal was positive with samples having a viral titer of not less than 10(5) TCID50/g. (C) 1999 Elsevier Science B.V. All rights reserved.