Obiettivo
The proposed research concerns the application of advanced biotechnology for solving a major fish health legislative problem concerning the recent finding of an important viral pathogen, VHS virus, in the marine environment and investigative scientific research on host pathogen relationships of marine VHS at the molecular level. Specific objectives are:
I. To determine the distribution of VHS virus in selected stocks of wild fish species in the Eastern Atlantic, North Sea and Baltic Sea.
II. To assess by experimental infection the virulence of marine isolates of VHS virus for aquaculture fish species of commercial significance including studies of disease pathogenesis.
III. To identify genetic and/or antigenic differences between marine VHSV isolates and other VHSV isolates.
IV. To identify genetic and/or antigenic differences between virulent and non-virulent VHSV isolates.
STATE OF PROGRESS
For the first two years the results of 17 research vessel cruises are reported. A total of 46,046 fish were sampled (5,634 fish samples remaining to be tested in 1999) comprising 43 species. Numbers of virus isolations were 107 from 12 species namely herring 40; sprat 37; dab 7; cod 7; Norway pout 7; whiting 2; plaice 2; flounder 1; blue whiting 1; rockling 1; lesser argentine 1; poor cod 1. Isolates were from the south Baltic Sea, Skagerrak, Kattegat, south Norwegian coast and the Atlantic, north and west of Scotland. Four isolates were made from cod skin showing lesions, 1 from normal cod skin and the rest were made from internal organs of asymptomatic fish. Using all types of skin lesion material from cod for virus isolation it was shown no correlation existed
Virulence testing of the marine isolates against rainbow trout: By bathing 32 isolates of European marine origin were tested in combination in 12 trials with either no mortality or very little mortality and therefore little evidence of virulence. By IP route 44 isolates, tested in pools, of European and Pacific origin were tested and found to be significantly virulent, mean mortalities of 14-44% for European, and 37-57% for Pacific isolates were recorded. By testing 20 European marine VHSV isolates singly by the IP route some isolates where found to be virulent whereas others were not. Mean mortalities from 0% to 42% were recorded.
Virulence testing of the marine isolates against turbot: The virulence of 36 marine isolates, divided into 11 groups, was tested in turbot by bath challenge. Total cumulative mortality ranged between 27% and 93% indicating that some marine isolates have high virulence while others have low virulence against turbot.
Virulence testing of the marine isolates against salmon: Thirty four marine isolates were divided into 11 groups and the virulence of each group tested by bath challenge of salmon in freshwater. Total cumulative mortalities varied from 0% in most groups up to 7.5% for one group. This indicates that marine VHSV isolates are avirulent or of very low virulence for salmon in freshwater.
The RNase protection assay (RPA) methodology was used to compare 24 strains of European marine origin. Two reference strains were used the virulent freshwater isolate 07-71 and an isolate from farmed turbot. The reference was the first 500 PB of the N gene. The isolates could be grouped into 4 using the turbot isolate as reference. 2 groups were from the Baltic, Skagerrak, and Kattegat area and 2 from the North Sea. One of the Baltic groups showed similarity to the other reference strain, the virulent 07-71.
Monoclonal antibodies that recognise different marine VHSV isolates have been produced. Unfortunately none of these discriminate marine isolates from other VHSV isolates.
DNA sequence analysis has been carried out on the genetic material from a total of thirty-one viral isolates. DNA sequence analysis has been carried out on five genes (G, N, M1, M2 and NV) from twenty-two viral isolates and two genes (G and N) from the remaining nine isolates. Sequence data was collected from these genes and sequence alignments were carried out. The alignments have shown that the viral isolates are very closely related at both nucleotide and protein level but sequence differences have been identified which may be specific to viruses from different geographical locations.
ACHIEVEMENTS
From 17 fishery research vessel cruises 46,046 fish comprising 43 species were sampled and 107 isolates of virus were made from 12 species.
Identification of an area of readily detectable prevalence in the Baltic, Skagerrak and Kattegat. The Atlantic, west and north of Scotland include new areas for the occurrence of VHSV.
All isolates tested up to now have low virulence to rainbow trout by bath challenge. Challenge by IP route showed many have virulence potential. Isolates show variable virulence to turbot by bath challenge and are avirulent, or of very low virulence, to salmon in freshwater.
RNase method has potential for rapid comparison of isolates and shows promise as a means of grouping isolates.
DNA sequence analysis has been carried out on the genetic material from a total of thirty-one viral isolates. Sequence data was collected from these genes and sequence alignments were carried out.
DESCRIPTION OF THE WORK
a) Collection of virus: Surveys by research vessels of Norway, Denmark and UK with emphasis on cod, haddock, sprat and herring will establish the distribution of the virus in these species in the North Sea, the Baltic Sea and adjacent areas. The surveys will result in a collection of isolates of the virus for use in laboratory studies.
b) Determination of virulence: Virulence of marine VHSV isolates and existing VHSV isolates from fresh water sources for aquaculture fish species of relevance to the EU will be tested: Rainbow trout, Atlantic salmon and other available species will be tested, e.g. turbot, halibut and cod.
c) Genetic/antigenic analyses and production of marker reagents including:
Sequencing of viral genomes
Comparative sequence analyses
Production of genetic probes in the form of oligonucleotides
Production of monoclonal antibodies.
Campo scientifico (EuroSciVoc)
CORDIS classifica i progetti con EuroSciVoc, una tassonomia multilingue dei campi scientifici, attraverso un processo semi-automatico basato su tecniche NLP. Cfr.: Il Vocabolario Scientifico Europeo.
CORDIS classifica i progetti con EuroSciVoc, una tassonomia multilingue dei campi scientifici, attraverso un processo semi-automatico basato su tecniche NLP. Cfr.: Il Vocabolario Scientifico Europeo.
- scienze naturali scienze biologiche microbiologia virologia
- scienze naturali scienze biologiche genetica DNA
- scienze naturali scienze biologiche biochimica biomolecole proteine
- scienze naturali scienze biologiche genetica genomi genomi virali
- scienze naturali scienze biologiche zoologia ittiologia
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Coordinatore
AB11 9DB Aberdeen
Regno Unito
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