of environmental impact from the use of live recombinant virus vaccines
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
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.
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.
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).
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.
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
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
Current opinion in Biotechnology, 2 (3), 1991, pp.
October 1991 September 1993
Institute for Animal Health
University of Liège (BE)
Central Veterinary Institute
Institute for Animal Health
University of Liverpool