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EC-sponsored Research on Safety of Genetically Modified Organisms - A Review of Results
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image Assessment of environmental impact from the use of live recombinant virus vaccines

Background and objectives

Vaccinia virus, a member of the Orthopoxivirus genus has been successfully used as a live vaccine for the prevention and the eradication of one of the most serious human diseases ever known, smallpox. Technical advances have permitted the development of vaccinia virus as a cloning and expression vector; vaccinia recombinants bearing foreign antigen-coding sequences have been developed to immunise against several diseases such as rabies and AIDS. The glycoprotein of rabies virus is the only rabies antigen capable of inducing the production of rabies virus-neutralising antibodies and has been shown to be able to confer immunity against rabies. The rabies virus glycoprotein gene has been inserted into vaccinia virus to generate a selectable recombinant to be used as a vaccine. This recombinant was named VVTGgRAB.

Control of infections in wild animals is difficult to achieve, mainly because such animals are free ranging, making them difficult to handle by conventional veterinary means. Vaccination is the most appropriate way of controlling infections at a population level, but in these cases the only practical route of administration is the oral route. We used the vaccinia virus to test the environmental impact of vaccination of wildlife with genetically modified viruses.


Fox. Fox.



Approach and methodology

It is preferable that a vaccine virus used for oral vaccination of wildlife not be horizontally transmitted to unvaccinated animals. In order to test for horizontal transmission, unvaccinated control animals were held in close contact with vaccinated ones.

It is also of major importance to preclude risks such as the emergence of asymptomatic carriers of wild rabies virus. This situation could occur in the field by vaccination of naturally infected animals during the incubation period. Therefore, the influence of vaccination with VVTGgRAB on the onset of the disease and on the delay before death in foxes previously infected with wild rabies virus, was investigated.

When assessing a recombinant virus for use in vaccination, it is also of great importance to detect any variation in tissue specificity compared with the parental vector strain. Experiments have been designed to determine the multiplication site in foxes of the recombinant virus compared to the parental strain by virus isolation, titration and indirect immunofluorescence. The polymerase chain reaction was also used to detect specific virus DNA in several fox organs. Foxes were fed with 108 CCID50 of either VVTGgRAB or vaccinia virus, and were euthanasied 12, 24, 48 or 96h after inoculation by the oral route.
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Main findings and outcome

No transmission of VVTGgRAB was found to occur in adult or young foxes, with the exception of one adult fox bitten by a freshly inoculated animal. All foxes, vaccinated or not, died from rabies. Animals vaccinated early after challenge died after a shorter period of incubation than unvaccinated controls. On the other hand, animals vaccinated belatedly after challenge died after the controls. The results show that “early” and “late” death phenomena occur as a consequence of interactions between oral vaccination with VVTGgRAB and rabies infection, but preclude the risk of the emergence of asymptomatic carriers of wild-rabies virus after vaccination.

VVTGgRAB or vaccinia virus could be detected during the first 48h following vaccination by the oral route, but only in the tonsils, buccal mucosa and soft palate. As no virus could be detected in the salivary glands of foxes (parotid or maxillary), the risk of transmission from one animal to another through saliva can be assumed to be negligible. Furthermore, the fact that VVTGgRAB only multiplies in restricted sites minimises the potential risk of recombination with other wild orthopoxviruses. In these experiments, no difference was observed between the multiplication sites of VVTGgRAB and vaccinia virus, demonstrating that the recombination event did not modify the tropism of the virus. Additionally, virus was never detected in the brain.

Several non-target wild species have been chosen for testing in Europe because of their opportunistic feeding behaviour and their presence in the areas where the vaccine must be distributed. Clinical signs of rabies and/or pox-inflicted lesions were never observed in these vaccinated animals during the observation period (28 days minimum after vaccination).
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Conclusions

Before using the recombinant vaccinia-rabies vaccine in the field to vaccinate foxes against rabies and thereby to reduce the incidence of rabies, we performed many safety tests. The vaccine was safe for the target species, the fox, as well as for numerous non-target species. The virus did not spread horizontally. We also examined differences in tissue tropism compared to the parental vector virus and the interference of vaccination on the outcome of rabies. All the experiments performed supported the safety of this vaccine for use in oral vaccination of wildlife against rabies.

 

Major publications

Brochier B., Blancou J., Aubert M.F.A., Kieny M.P., Desmettre P., Pastoret P.P., "Interaction between rabies infection and oral administration of vaccinia-rabies recombinant virus to foxes (Vulpes vulpes)”.
J. Gen. Virol.,
70, 1989, pp. 1601-1604.

Brochier B., Blancou J., Thomas I., Languet B., Artois M., Kieny M.P., Lecocq J.P., Costy F., Desmettre Ph., Chappuis G., Pastoret P.P., “Use of recombinant vaccinia-rabies glycoprotein virus for oral vaccination of wildlife against rabies: innocuity to several non-target bait consuming species”.
J. Wildl. Dis.,
25, 1989, pp. 540-547.

Thomas I., Brochier B., Languet B., Blancou J., Péharpré D., Kieny M.-P., Desmettre P., Chappuis G., Pastoret P.-P., "Primary multiplication site of the vaccinia-rabies glycoprotein recombinant virus administered to foxes by the oral route".
J. Gen. Virol.,
71, 1990, pp. 37-42.

Pastoret P.P., Brochier B., Blancou J., Artois M., Aubert M., Kieny M.P., Lecocq J.P., Languet B., Chappuis G., Desmettre P., “Development and deliberate release of a vaccinia-rabies recombinant virus for the oral vaccination of foxes against rabies”, in Recombinant Poxviruses, Matthew M. Binns and Geoffrey L. Smith (eds.), CRC Press, Baton Roca, USA, 1992, pp. 163-206.

Pastoret P.P., Brochier B., “Biological control of wild animal infections”.
Current opinion in Biotechnology, 2 (3), 1991, pp. 465-469.
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imageResearch project
 

Contract number
BIOT-CT91-0289

Period
October 1991 – September 1993

Coordinator
D.N. Black
Institute for Animal Health
Woking (UK)

 
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Partners


P.P. Pastoret
University of Liège (BE)

A.L.J. Gielkens
Central Veterinary Institute
Lelystad (NL)

M.A. Skinner
Institute for Animal Health
Huntingdon (UK)

R.M. Gaskell
University of Liverpool
Neston (UK)

 
 
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