Population-level retrospective study of neurologically expressed disorders in ruminants before the onset of bovine spongiform encephalopathy (BSE) in Belgium, a BSE risk III country.

A retrospective epidemiological study (n = 7,875) of neurologically expressed disorders (NED) in ruminants before the onset of the bovine spongiform encephalopathy epidemic (years studied, 1980 to 1997) was carried out in Belgium. The archives of all veterinary laboratories and rabies and transmissible spongiform encephalopathy (TSE) epidemiosurveillance networks were consulted. For all species, a significantly higher number of NED with virological causes (rabies) was reported south of the Sambre-Meuse Valley. During the period 1992 to 1997, for which the data were complete, (i) the predicted annual incidence of NED varied significantly as a function of species and area (higher numbers in areas where rabies was present) but was always above 100 cases per million, and (ii) the mean incidence of suspected TSE cases and, among them, those investigated by histopathological examination varied significantly as a function of species and area. The positive predictive value of a presumptive clinical diagnosis of NED ranged from 0.13 (game) to 0.63 (sheep). Knowledge of the positive predictive value permits the definition of a reference point before certain actions (e.g., awareness and training campaigns) are undertaken. It also shows the usefulness of a systematic necropsy or complementary laboratory tests to establish an etiological diagnosis. TSE analysis of a small, targeted historical sampling (n = 48) permitted the confirmation of one case and uncovered another case of scrapie. The results of the present study help to develop and maintain the quality of the worldwide clinical epidemiological networks for TSE, especially in countries that in the past imported live animals, animal products, and feedstuffs from countries with TSE cases.

Comparison of visual microscopic and computer-automated fluorescence detection of rabies virus neutralizing antibodies.

The rapid fluorescent focus inhibition test (RFFIT) and the fluorescent antibody virus neutralization test (FAVNT) are both diagnostic tests for determining levels of rabies neutralizing antibodies. An automated method for determining fluorescence has been implemented to reduce the work time required for fluorescent visual microscopic observations. The automated method offers several advantages over conventional visual observation, such as the ability to rapidly test many samples. The antibody titers obtained with automated techniques were similar to those obtained with both the RFFIT (n = 165, r = 0.93, P < 0.001) and the FAVNT (n = 52, r = 0.99, P < 0.001).

Rabies prophylaxis.

In humans, rabies still is a fatally evolving encephalomyelitis caused by a Rhabdovirus of the genus Lyssavirus. In general, the disease is contracted through a contact with an infected mammal. Taxonomically, different rabies and closely related rabies-like viruses can be distinguished. New molecular identification techniques can be utilized as epidemiological tools to study the geographic distribution and presence in different reservoirs of the viruses. Antigenic diversity and new insights in the mechanisms of the immune response can have serious implication in vaccine strategies. Virus detection for diagnostic and epidemiological purposes can be done by immunofluorescency, by inoculating murine neuroblastoma cells and by using molecular techniques. Rabies is a zoonosis with a worldwide distribution. In Belgium, the epizootic is present in the Southern part of the country. Fox vaccination campaigns contributed significantly to the eradication of the virus from its natural reservoir. The importance of the prophylactic and therapeutic use of the vaccine, the control of wildlife animal reservoir and stringent public health measures to combat rabies is discussed. Due to stringent control measures, no endogenous case of human rabies have been reported since 1922 in Belgium.

Elimination of fox rabies from Belgium using a recombinant vaccinia-rabies vaccine: an update.

To improve both safety and stability of the vaccines used in the field to vaccinate foxes against rabies by the oral route, a recombinant vaccinia virus, expressing the glycoprotein of rabies virus (VVTGgRAB) has been developed. VVTGgRAB innocuity was verified in target species and in domestic animals as well as in numerous wild animal species that could compete with the red fox in consuming vaccine baits in Europe. Oral immunization of foxes, by distributing VVTGgRAB vaccine-baits, was undertaken in the whole of the infected area of Belgium (10,000 km2). Five campaigns of fox vaccination covering the whole infected area were carried out from the autumn of 1989 until 1991. Each time, 150,000 vaccine-baits were dispersed by air at a mean density of 15 per km2. These campaigns induced a drastic decrease in the incidence of rabies and the elimination of the disease from 80% of the initial infected area. Regarding the geographical evolution of rabies in Belgium and in adjacent regions in neighbouring countries, new spatial strategies for bait dispersal were planned for 1992, 1993 and 1994: successive restricted campaigns were carried out along political borders only. These campaigns induced a new decrease of incidence; no rabid foxes could be detected in 1993 in spite of an improved epidemiological surveillance. In 1994, rabies was confirmed again in 13 foxes collected in a region situated close to the French border. These cases demonstrate the persistence of a focus of rabies on the border and justify further restricted campaigns of vaccination.

Towards rabies elimination in Belgium by fox vaccination using a vaccinia-rabies glycoprotein recombinant virus.

Oral immunization of foxes by distributing vaccine baits was experimentally assessed and subsequently employed in the whole of the infected area of Belgium (10,000 km2). A vaccinia-rabies glycoprotein recombinant virus (VR-G) was used as vaccine because of its efficacy, safety and heat stability. Five 'full' campaigns of fox vaccination, carried out from 1989 until 1991, induced a drastic decrease in the incidence of rabies. The disease has disappeared from the major part of the initial infected area. In 1992 and 1993, three 'defence' campaigns, carried out along international borders, completely eliminated rabies virus infection from the fox population in 1993.

Large-scale eradication of rabies using recombinant vaccinia-rabies vaccine.

Rabies infection of domestic and wild animals is a serious problem throughout the world. The major disease vector in Europe is the red fox (Vulpes vulpes) and rabies control has focused on vaccinating and/or culling foxes. Culling has not been effective, and the distribution of five vaccine baits is the only appropriate method for the vaccination of wild foxes. Although some European countries have conducted field vaccination campaigns using attenuated rabies virus strains, their use has not been extensively approved because they retain pathogenicity for rodents and can revert to virulence. These strains cannot be used in North America because they are pathogenic for the striped skunk (Mephitis mephitis) and are ineffective in the raccoon (Procyon lotor). We have constructed a recombinant vaccinia virus, VVTGgRAB, expressing the surface glycoprotein (G) of rabies virus (ERA strain). The recombinant was a highly effective vaccine in experimental animals, in captive foxes and in raccoons. We report here the results of a large-scale campaign of fox vaccination in a 2,200 km2 region of southern Belgium, an area in which rabies is prevalent. After distribution, 81% of foxes inspected were positive for tetracycline, a biomarker included in the vaccine bait and, other than one rabid fox detected close to the periphery of the treated area, no case of rabies, either in foxes or in domestic livestock, has been reported in the area.

Use of recombinant vaccinia-rabies glycoprotein virus for oral vaccination of wildlife against rabies: innocuity to several non-target bait consuming species.

The pathogenicity of a vaccinia recombinant virus expressing the rabies glycoprotein (VVTGgRAB) was tested in several wild animal species which could compete with the natural rabies host, the red fox (Vulpes vulpes) in consuming vaccine baits in Europe. The following species were included in this study: wild boar (Sus scrofa), Eurasian badger (Meles meles), wood mouse (Apodemus sylvaticus), yellow-necked mouse (Apodemus flavicollis), bank vole (Clethrionomys glareolus), common vole (Microtus arvalis), field vole (Microtus agrestis), water vole (Arvicola terrestris), common buzzard (Buteo buteo), kestrel (Falco tinnunculus), carrion crow (Corvus corone), magpie (Pica pica) and jay (Garrulus glandarius). During the observation period, the 107 animals given the VVTGgRAB vaccine orally did not show any clinical signs. Daily monitoring for 28 days and postmortem examination did not result in the detection of pox lesions in the oral mucosa or the skin in mammals or the unfeathered portions of birds. VVTGgRAB seems to multiply in the mammalian species tested, since rabies seroconversion was observed in all of them. Birds failed to develop demonstrable rabies virus-neutralizing antibody. A seroconversion against vaccinia virus was observed in two of four wild boars. Serological results obtained in badgers and wild boars also demonstrates the absence of direct or indirect horizontal transmission of the recombinant virus. The potential of the recombinant virus for the immunization of badgers against rabies also was investigated. Only 50% of the badgers orally administered with 1 x 10(8.3) TCID50 of this vaccine were protected against rabies.

[Rabies, a public health problem]. / La rage, un problème de santé publique.

In West-Europe, rabies is mostly reported in foxes, and in less extend in different domestic animals including dogs, cats, cattle, sheep, goats and horses. The contamination to humans occurs through contact with rabid animals by bite, scratch and even contact with the wound. This paper reports upon the current situation of rabies disease in Belgium and the prophylactic means routinely applied and those actually under experiment. In 1988, 514 animals were found positive, among which 366 foxes (71%), 95 cattle (18%), 46 sheep (9%), 14 cats (3%) and 15 dogs (3%). Due to this epizootic situation, 1391 human vaccinations (845 curative- and 546 preventive vaccinations) were given. Recently, few cases have been encountered at the north side of the Sambre-Meuse. This streamline has hitherto been considered to offer an adequate natural barrier against the rabies disease spreading. Further discussed is the prophylactic oral vaccination in foxes using two vaccines types: the live attenuated rabies vaccine and the rabies virus glycoprotein gene- vaccinia virus recombinant vaccine.

A field trial in Belgium to control fox rabies by oral immunisation.

Campaigns of fox vaccination against rabies were carried out in Belgium in September 1986 and June and September 1987. The SAD B19 attenuated strain of rabies virus was inserted into baits which were distributed over an area of 2100 km2 at a density of 11 baits/km2. As recommended by the World Health Organisation, the efficacy and the innocuity of the method were controlled in the field and in the laboratory. Samples of blood and brain and jaw were taken from foxes which were shot or found dead in the vaccination area, for the diagnosis of rabies, the titration of antirabies antibody and the detection of tetracycline marker. In rabid animals, the virus strain was characterised by immunofluorescence using monoclonal antibodies. In September 1987, the uptake of the baits had reached 72 per cent by 14 days after distribution. Several wild species competed with foxes in taking the baits. After the last campaign, tetracycline was found in 65 per cent of the healthy foxes collected and rabies virus neutralising antibodies were detected in 77 per cent of them. In 1987, the incidence of rabies decreased markedly in the vaccination area compared with the untreated areas. No vaccine virus was isolated either from rabid animals or from 228 small mammals trapped in the vaccination area.

First field trial of fox vaccination against rabies using a vaccinia-rabies recombinant virus.

A field trial of fox vaccination against rabies using a vaccinia-rabies recombinant virus was carried out in Belgium on October 24, 1987. Each vaccine capsule contained a suspension of 10(8) TCID50 of the recombinant virus and was introduced into a chicken head. Each chicken head contained 150 mg of tetracycline as a marker of uptake. Two hundred and fifty heads were distributed in an area of 6 km2 situated within a military zone. The bait uptake was monitored for 15 days after the distribution. Sixty-three per cent of the chicken heads were taken by wild animals within that period. The trial was controlled according to the rules defined by the World Health Organisation.

Use of recombinant vaccinia-rabies virus for oral vaccination of fox cubs (Vulpes vulpes, L) against rabies.

Thirteen fox cubs were orally administered 10(7.2) plaque-forming units of live vaccinia-rabies glycoprotein recombinant virus. On Day 28 post-vaccination, all but 1 cub had produced rabies virus antibodies. Twelve animals were intramuscularly inoculated with 10(3.2) mouse intracerebral LD50 of rabies virus suspension on Days 33 (5 foxes), 180 (4 foxes) or 360 (3 foxes) after vaccination. Eleven of them resisted rabies challenge. Unvaccinated foxes, either put in contact with 1 vaccinated animal or used as controls, died after challenge applied on Day 33. The absence of horizontal transmission of this vaccine strain and its innocuity to cubs were also demonstrated.

[Experience with antirabies vaccination of foxes using the oral route coordinated among several European countries and perspectives on the use of recombinant vaccinia-rabies virus]. / Expérience de vaccination antirabique du renard par voie orale coordonnée entre plusieurs pays européens et perspectives d'utilisation du virus recombinant vaccine-rage.

Campaigns of fox vaccination against rabies were carried out in Belgium, grand-duchy of Luxembourg and France in September 1986, June and September 1987. The SAD B19 attenuated strain of rabies virus, contained in baits (Tübingen baits) was used as vaccine. Baits were distributed at a range density of 11 to 15 baits per km2. First results are very encouraging. A recombinant vaccinia virus harbouring the rabies virus glycoprotein gene has been developed. This recombinant virus can be given to the fox by the oral route and protects it against rabies virus challenge; it is also innocuous for the fox and other non-target European species. A first trial of fox vaccination against rabies using this recombinant vaccinia-rabies virus has been carried out in Belgium, on a military domain, in October 1987.

Immunization of young foxes against rabies: interaction between vaccination and natural infection.

A preliminary experiment of vaccination against rabies with a parenterally administered killed vaccine was performed in Belgium with young foxes (Vulpes vulpes L.) captured in an enzootic area (Province de Luxembourg). Out of 20 young foxes, 12 developed spontaneous rabies. Vaccination induced a rapid and important seroconversion in most of the non rabid animals but failed to stop the evolution of the disease in young foxes incubating rabies. On the contrary, vaccination seemed to accelerate the evolution.

Vaccination of young foxes (Vulpes vulpes, L.) against rabies: trials with inactivated vaccine administered by oral and parenteral routes.

Foxes aged between two and three months were vaccinated with an inactivated rabies vaccine. Three immunization procedures were attempted: a first group of animals was parenterally injected while two other groups daily ingested during ten days either an uncoated vaccine (commercially available liquid form: Rabisin) or a coated vaccine (gastro-resistant tablets) associated with a potential enteropathogenic virus (bovine rotavirus). The latter viral association was used in order to see if seroconversion can occur in these conditions. Rabies antibody titres and the length of survival postchallenge were recorded in each treated fox. Seroconversion was observed from approximately day 20 post-vaccination in two groups of cubs: satisfactory antibody titres were found in parenterally vaccinated foxes while lower titres characterized most of the cubs which ingested the uncoated liquid vaccine. Gastro-resistant tablets containing the viral association did not induce seroconversion. It was shown that bovine rotavirus did not multiply in the gut mucosa of young foxes, on the contrary to young dogs. Protection against experimental rabies was partially obtained in parenterally vaccinated foxes.