A previously unknown mechanism in viral pathogenesis leading to effective new vaccines and post-exposure immune treatments of viral infections.

Investigations with the rabies virus identified a previously unknown mechanism of viral pathogenesis. After ultracentrifugation of a suspension of rabid dog brain and rabies vaccine strains, the supernatant was found to contain active components, as evaluated in an in vitro plaque test. The unexpected detection of active components in non-sedimented material prompted further research and the finding that these components, and not the complete virus, were responsible for paralysis and death. Vaccination of cattle with existing rabies vaccines showed that even low titres of antibodies against these components provided protection after challenge. In a control group of non-vaccinated cows, cows that had low titres of these antibodies survived rabies challenge. These low titres could not be detected with the usual serum neutralization test in mice but only with a plaque reduction test, which is more sensitive. A hyperimmune serum raised in rabbits against active components isolated from the brain of a rabid dog was injected intracerebrally into mice that had been previously injected intramuscularly with a rabies virus. This delayed post-exposure treatment was still effective against advanced rabies (virus already in the brain, but not yet paralytic symptoms). This promising finding makes the development of a new inactivated rabies vaccine possible and opens the way for post-exposure treatment for humans, particularly in developing countries where rabies is still a major problem. The role of these active components in other viral diseases, such as human immunodeficiency virus, should be investigated.

The efficacy of a new single post-exposure treatment of rabies in mice without vaccination.

Local application of rabies immune serum and isoprinosine, an immunomodulator with antiviral activity was effective in mice infected with a sylvatic rabies virus. In this way, a single medical or veterinary treatment is only required, which is particularly important for developing but also for developed countries. The importance of using a post-exposure potency test to monitor rabies vaccines is emphasized. The same principle could be applied to other emerging viral infections of humans (for example, human immunodeficiency virus infection) and animals, for which no effective vaccines are available at this moment.

[Myxomatosis in the sub-antarctic islands of Kerguelen, without vectors, thirty years after its introduction]. / La myxomatose dans les îles subantarctiques de Kerguelen, en l'absence de vecteurs, trente années après son introduction.

Myxoma virus was introduced into the Kerguelen archipelago in 1955-1956. Thirty years after its introduction, the virus is present in most areas inhabited by rabbits. Rabbit fleas and mosquitoes are absent from this group of islands and the disease is transmitted by contact. The timing of the beginning of new myxomatosis outbreaks, the absence of real epizootics as well as the higher percentage of infected males over females are specific observations in favour of this mode of transmission. The majority of 34 isolates tested between 1984 and 1988 are of intermediate virulence (Grades IIIA-IIIB). In these conditions, the impact of myxomatosis virus on rabbit populations estimated on two sites is low. Myxomatosis therefore plays only a minor role in the regulation of rabbit populations.

Isolation and plaque purification of a syncytial forming component of myxoma virus.

Using the endpoint dilution and plaque purification technique performed on rabbit kidney cells, two stable syncytial clones of myxoma virus have been isolated. No eosinophilic intracytoplasmic inclusion bodies nor intranuclear granulations could be detected in cells infected with these clones. Traditional plaques with the central area of detached cells surrounded by round cells also could not be recognized. However, when enough virus was added, one large complete syncytium developed in 24 on the monolayer. Syncytial activity was associated with the entire particle. This was the first time that myxoma virus is reported to demonstrate such an intense syncytial activity.

[Isolation and in vitro and in vivo characteristics of a clone of a wild Aujeszky disease virus resistant to bromovinyldeoxyuridine]. / Isolement et caractères in vitro et in vivo d'un clone de virus sauvage de la maladie d'Aujeszky résistant à la bromovinyldésoxyuridine.

The antiviral activity of bromovinyldeoxyuridine has been studied on the wild type of pseudorabies virus. A heterogenicity of viral populations has been observed. A resistant clone presented special characteristics both in vitro (large syncytia, round cells) and in vivo (reduced pathogenicity and immunogenic power).

[The phenomenon of "early and stimulated death" in mice infected and then vaccinated against rabies]. / Le phénomène de "mort précoce et stimulée" chez la souris infectée puis vaccinée contre la rage.

Live (diluted) and inactivated rabies vaccines of low antigen content induce early and enhanced death in mice, inoculated before vaccination with a wild type of rabies virus. Such vaccines, which neither induce interferon nor protect, produce a low level of antibodies which appear later than with vaccines of higher antigenicity. It is recommended to examine rabies vaccines not only by the usual (pre-exposure) potency test-NIH test or modified NIH test (one vaccination), but also by a post-exposure potency test.

A plaque-purified rabies virus (strains V-319) derived from a vampire bat (Desmodus rotundus) in Mexico: vaccine potential.

A plaque-purified experimental rabies vaccine was developed from an isolate (strain V-319) made from a naturally infected vampire bat (Desmodus rotundus). Two different vaccines were prepared; one was live virus and the second was an inactivated rabies virus preparation. The live virus vaccine, as well as a betapropiolactone-inactivated vaccine, gave complete protection to challenge inoculation after 1 year. In contrast, greater than 80% of the non-vaccinated experimental control cattle died of rabies. The live virus vaccine could be given at doses as low as 10(5) PFU without loss of efficacy. It did not cause adverse reactions. More than 10,000 cattle have been vaccinated with the live virus vaccine under field conditions. No rabies deaths occurred in vaccinated cattle during a 2-year postvaccinal period. The serological responses of vaccinated cattle indicated protection that endured at least 1 year.

A rabies virus persistent infection in BHK21 cells.

A rabies virus persistent infection in BHK21 S13 cells was established and maintained in culture for more than 4 years. Initially, the cultures produced a large plaque virus similar to that produced by the original virus, but between the 10th and 20th passage, this was replaced by a small plaque variant. By the 200th passage, infectious virus could no longer be detected in the medium. After further cell passages (greater than or equal to 300) no infectious particles could be detected in the medium. At various passage levels, the persistently infected cells were labelled with [35S]methionine and the virus antigens immunoprecipitated and analysed by polyacrylamide gel electrophoresis. No changes in the virus polypeptides were observed in the establishment of the persistent state. However, after the 20th passage (predominance of small plaque variant) there was an increase in the size of the glycoprotein. This was followed (164th passage) by a change in the MI polypeptide which was subsequently further modified in the defective state (greater than or equal to 300 passages). Virus isolated from the 400th passage by treatment of the cells with DEAE-dextran, was also modified in the glycoprotein and M1 polypeptides and contained less L polypeptide than the original virus. This virus grew more slowly, to a lower titre and was no longer pathogenic in suckling mice.

A potency test which simulates natural exposure for measuring post-exposure activity of rabies vaccines. A proposal for preparing a relevant international reference preparation.

There is a basic difference between the mechanism of immunity at the pre- and post-exposure level. Due to the fact that vaccination precedes challenge in the available potency tests (Habel and NIH) they do not measure post-exposure activity of rabies vaccines. For these reasons a potency test in mice has been developed which measures post-exposure activity of rabies vaccines. In this test natural conditions have been simulated by using intramuscular inoculation of a street virus (fox salivary gland origin), killing approximately 50% of the control mice, whereas intraperitoneal inoculation with 0.5 ml of undiluted inactivated rabies vaccine within 24 hours after street virus infection, should protect all mice. A human rabies vaccine prepared in HDC and already licensed for post-exposure vaccination showed only some post-exposure activity in this newly developed potency test, whereas an experimental inactivated rabies vaccine protected all mice. The International rabies Reference Vaccine did not protect after exposure but, on the contrary, enhanced the rabies street virus infection (shorter incubation period and more mice died of rabies than in the controls); it is therefore unsuitable for measuring post-exposure potency. Authorities concerned with rabies vaccine control are invited to use this newly developed post-exposure potency test in their laboratories and if agreed upon its value, to prepare an adequate batch of inactivated tissue culture rabies vaccine which meets this test-requirement. Such a batch could be a candidate for the establishment of an international reference preparation measuring post-exposure potency of rabies tissue culture vaccines.

Comparative studies of atherosclerosis in swine.

A survey of spontaneously occurring fatty streaks and fibrous plaques, considered as atherosclerosis, in 1637 swine in different European countries and the USA, using a standardized procedure, was undertaken to determine whether significant differences exist in the occurrence and extent of the disease in various groups of animals. At the same time a preliminary study on the possible relation of any differences observed in atherosclerosis to certain environmental and constitutional factors was carried out with the ultimate goal of contributing to the understanding of analogous problems in man.STATISTICALLY SIGNIFICANT INCREASES OF FATTY STREAKS AND FIBROUS PLAQUES WERE NOTED IN RELATION TO: (a) increasing age, starting at 6 to 7 months, the earliest age period studied; (b) geographical locality; and (c) considerable as compared with moderate or slight physical activity at 1 year of age. Although not statistically significant, there was also a suggestive trend towards more atherosclerosis in pigs consuming soft water as compared with those consuming hard water.While these correlations may represent contributory factors to the increases of the changes noted in the abdominal aortas, it is not possible to pinpoint the importance of individual components because of the limited data and the large number of variables involved in this preliminary study. Studies in swine and other animals are being encouraged in which all variables but one are being kept constant to determine their possible role in the development of atherosclerotic lesions.