Eighteen years of vaccination against viral haemorrhagic septicaemia in France.

Viral haemorrhagic septicaemia (VHS) has been considered for many years to be a major cause of loss in the French trout industry. The high prevalence of VHS in certain geographic areas made a control strategy based on control policy unfeasible. This provided the impetus for immunoprophylaxis development that resulted in 3 successive types of vaccines: inactivated, live attenuated and recombinant vaccines. When delivered by intraperitoneal injection, the 2 propiolactone-inactivated VHS virus was immunogenic and/or protective for trout all of sizes, but it was not suitable for the practical immunization of alevin, the trout life stage that is the most sensitive to VHS. A carp cell-passed, attenuated variant of the VHS virus was effective after both immersion or injection delivery and met the practical requirements of juvenile vaccination. However, this vaccine was discarded because it retained some virulence that discouraged the launching of its commercialization. Then came the era of genetically engineered vaccines. The recombinant glycoprotein of VHSV produced in Escherichia coli or in Saccharomyces cerevisiae failed to protect fish whatever the route of delivery. A recombinant baculovirus vaccine was found to be immunogenic and protective against VHS, but only when delivered by injection. Due to its cost and route of delivery, the latter vaccine was not licensed. Simultaneously, the sudden occurrence of another rhabdovirosis, infectious haematopoietic necrosis (IHN), in France, rendered vaccination against VHS questionable. Indeed, no cross-protection between these 2 rhabdoviroses exists. If vaccination is still believed to be an effective control method for VHS, it should be based in the future upon an autoreplicative vaccine.(ABSTRACT TRUNCATED AT 250 WORDS)

A recombinant viral haemorrhagic septicaemia virus glycoprotein expressed in insect cells induces protective immunity in rainbow trout.

Viral haemorrhagic septicaemia (VHS) is a fish rhabdovirus infection of world-wide importance. Control policies have been established but the disease still causes heavy losses in fish farming. The development of a recombinant subunit vaccine was initiated to produce a safe and effective vaccine to protect fish against VHS. The VHS virus (VHSV) glycoprotein, which induces neutralizing antibodies in rainbow trout, was chosen for expression in insect cells using a baculovirus vector. The M(r) of the recombinant protein estimated by SDS-PAGE was slightly lower than that of the native viral protein. The recombinant protein displayed different degrees of glycosylation and was recognized in ELISA by neutralizing antibodies. It was transported to the plasma membrane of insect cells where its ability to induce membrane fusion was preserved. The efficacy of the recombinant protein as a vaccine was compared with those of an inactivated and an attenuated vaccine. When injected intraperitoneally into rainbow trout, the baculovirus-encoded protein was shown (i) to induce the synthesis of VHSV-neutralizing antibodies and (ii) to confer protection against virus challenge. Immunization performed by immersion failed. This is the first report of a recombinant vaccine that protects fish against VHSV.

Cloning and sequencing the messenger RNA of the N gene of viral haemorrhagic septicaemia virus.

The mRNA transcribed from the N gene of viral haemorrhagic septicaemia virus (VHSV) of salmonids has been cloned in Escherichia coli and expressed. Fusion proteins were recognized by monoclonal antibody directed against the N protein from the viral particle. A 1212 bp long open reading frame (ORF) coding for 404 amino acids with a calculated Mr of 44590 was deduced from the nucleotide sequence. The ORF was preceded by a 93 bp segment including in position 42 the AACAC pentanucleotide which is presumed to be the start signal for transcription by analogy with other rhabdoviral mRNAs. The upstream 41 bp region could correspond to the covalently linked positive polarity leader RNA as also found on the N mRNA from infectious haematopoietic necrosis virus (IHNV). This may be a characteristic of fish lyssaviruses. The AAACC sequence, which is part of the leader, was not found. Amino acids 44 to 359 from IHNV and 45 to 360 from VHSV are 45.3% homologous. A strong homology which could reflect functional importance was also found for potential phosphorylation sites and hydrophobic peaks despite the fact that the two viruses evolved on different continents.

Molecular cloning of the mRNA coding for the G protein of the viral haemorrhagic septicaemia (VHS) of salmonids.

Viral haemorrhagic septicaemia virus (VHSV), a rhabdovirus, is a major threat for continental European trout fish farming. The development of a recombinant subunit vaccine could solve that problem. The neutralizing epitopes are located on the glycoprotein or G protein, the surface antigen. The G protein has a molecular weight of 65 kDa, reduced to 55 kDa by deglycosylation. cDNA was synthetized from mRNA of VHS virus infected cells, and cloned in E. coli. The viral cDNA was recognized by positive hybridization with a labelled probe made from infected cell RNA, and negative hybridization with labelled cDNA made from cellular RNA. The Northern blot hybridization with different clones on VHS infected cell RNA revealed two VHS mRNA whose lengths, 2.0 and 1.5 kb, were compatible with the mRNA length for G and N proteins respectively. This mRNA must contain about 400 bp of untranslated sequence.