Although transmission stu&es showed that an infect~ous agent was involved in an eplzootic mortality in hatchery-reared fingerling and yearling lake trout Salvelinus namaycush, routine diagnostic examinations repeatedly f a~l e d to detect infectious or paras~tic agents. Histological examinatlon consistently showed epithehal hyperplasia inside the mouth, on the snout, and over the body. Electron microscopy revealed a putative virus associated with the hyperplaslc tissue; however, the virus has yet to be isolated in cell culture. Morphologically, the virus resembled a herpesvirus, but its taxonomic grouping has yet to b e firmly established. In laboratory studies, the infection could be transmitted by cohabitation with diseased fish, by exposure to water from tanks holding diseased fish, and by bath challenge with filtrates (0.45 pm and 0.22 pm) of hyperplastic epidermal tissue. As judged from laboratory challenges and hatchery observations, the disease d~d not appear to affect brook trout Salvelinus fonhnalis, brown trout S a l n~o trutta, rainbow trout Salnio gairclneri, or Atlantic salmon Salmo salar.

A reverse transcriptase polymerase chain reaction (RT-PCR) assay was developed for the detection and identification of aquatic birnaviruses. The four sets of primers (PrA, PrB, PrC, and PrD) that we used are specific for regions of cDNA coded by genome segment A of aquatic birnaviruses. PrA identifies a large fragment (1,180 bp) within the pVP2-coding region, and PrB identifies a 524-bp fragment within the sequence amplified by PrA. Primer set PrC frames a genome fragment (339 bp) within the NS-VP3-coding region, and PrD identifies a 174-bp sequence within the fragment identified by PrC. PrB and PrD amplified cDNAs from all nine recognized serotypes of aquatic birnavirus serogroup A as well as the N1 isolate that may represent a 10th serotype. These results indicate that these three primer sequences are highly conserved and can be used in PCR assays for group identification of these viruses. PrA routinely produced amplification products from eight serotypes but exhibited variable results with one serotype, and primer PrC identified 6 of the 11 virus isolates tested. The qualitative sensitivity of the RT-PCR assay was evaluated by comparison of the results with those of cell culture isolation assays. With the exception of one sample, the RT-PCR assay with primer PrD was as accurate as cell culture isolation for detecting virus in kidney and spleen tissues from naturally infected, asymptomatic carrier fish. These results indicate that the RT-PCR assay can be a rapid and reliable substitute for cell culture methods for the detection of aquatic birnaviruses.

The response of mouse L cells to infection with wild-type (wt) and temperature-sensitive (ts) mutants of vesicular stomatitis virus was monitored by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to delineate the synthesis of host cell and viral proteins. Experiments utilized transcriptase mutants of complementation group I (ts114 and ts13), a group IV mutant (ts44) that is restricted in total RNA synthesis (RNA-) but not in primary transcription, and a group II mutant (ts52) variably restricted in RNA synthesis (RNA+). L cells infected with ts mutants at permissive temperature exhibited the wt response of progressive inhibition of host cell protein synthesis accompanied by accumulation of all five viral proteins. Mutant ts44 (IV) also switched off cell protein synthesis at restrictive temperature and accumulated all five viral proteins, but with disproportionate ratios of N and G proteins. At restrictive temperature, cells infected with group I ts mutants failed to accumulate any viral protein and did not exhibit significant reduction in host cell protein synthesis. These data suggest that vesicular stomatitis virus inhibits cell protein synthesis at a stage of viral infection after transcription and possibly translation but preceding replication of progeny viral RNA.

in wild non-salmonid fish (Stephens, Newman,

A protocol for experimental challenge with infectious pancreatic necrosis (IPN) virus was defined with brook trout Salvelinus fontinalis as the model species. Fish were exposed by immersion for 5 h in water containing IPN virus at a concentration of 105 plaque‐forming units per milliliter. We propose the protocol as a standard challenge based on our studies of exposure methods and host response to virulent virus. Immersion challenge induced higher and more consistent mortality than did challenge by hyperosmotic infiltration. Challenge virus should be sequentially transferred no more than five times in cell culture because further transfers reduced the virulence of the virus. At 12°C, mortalities due to primary infection occurred 6–12 d after immersion challenge and were highest (≥70%) in fish 27–56 d old. Susceptibility to lethal infection was enhanced by nutritional stress.