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Kinetic haemagglutination inhibition technique as a means of detecting antigenic variations among strains of Nigerian flaviviruses.

Seven isolates of West Nile virus and an additional isolate tentatively identified at the outset as West Nile virus, were compared by Casals' modified haemagglutination inhibition technique. Two of the isolates were nearly identical; a second set of five isolates cross-reacted among themselves within a wide range and differed from the first two; the last isolate was significantly different from the others.

Antigens, Viral

Protease treatment and chemical crosslinking of a flavivirus: tick borne encephalitis virus.

Tick-borne encephalitis virus was treated with pronase or thermolysin. The resulting particles were banded in sucrose gradients and analyzed for polypeptide composition. Both enzymes caused a reduction in particle density from 1.19 to 1.15--1.16 g/cm3. No loss of viral lipid or nucleic acid could be observed. SDS-polyacrylamidegel electrophoresis showed that only the core protein V2 was unchanged whereas the envelope proteins V3 and V1 had disappeared from their original positions in the PAGE profile. Instead a new peptide(s) with molecular weight of 4000--6000 was found in which hydrophobic amino-acids were enriched. Crosslinking by dimethyl-3.3'-dithiobispropionimidate (DTBP) made the virus resistent to solubilization of the envelope proteins by TX-100. This could be interpreted by the formation of a dense envelope protein network around the nucleocapsid preventing its liberation by TX-100. Some data however indicate that direct crosslinking of at least one of the envelope proteins with the core cannot be excluded.

Cross-Linking Reagents

Studies of louping-ill virus (Flavivirus group) in wild red grouse (Lagopus lagopus scoticus).

Studies were made to find evidence of louping-ill virus infection in free-living red grouse and relate this to their breeding success. In areas where ticks were abundant 61 (84%) adult grouse had antibody to the virus compared with 1 (10%) in areas where ticks were relatively scarce. Of 162 chicks tested 25 were shown to be viraemic. Infected chicks were of significantly less weight than comparably aged uninfected birds and the probability that they died was much greater than that of uninfected birds. It is concluded that the relatively poor breeding success in areas of high tick numbers was principally due to infection with louping-ill virus. The susceptibility of the red grouse to infection is discussed.

Animal Population Groups

Immune response in rabbits to virion and nonvirion antigens of the Flavivirus kunjin.

The nature of the antibodies formed in rabbits in response to the following Kunjin virus antigens was examined: infectious suckling mouse brain (SMB), purified virion or rapidly sedimenting hemagglutinin (RHA), slowly sedimenting hemagglutinin (SHA), and envelope fragments prepared from RHA disrupted by 0.1 or 0.2% sodium deoxycholate (DOC). The hemagglutination-inhibiting (HI) and neutralizing antibody responses to SMB, RHA, and large envelope fragments (0.1% DOC) were remarkably uniform, antibodies appearing at the same time, attaining similar HI titers (lowest to envelope), and being of similar avidity early and late in the respone. The 19S (immunoglobulin M) antibodies to all antigens were always relatively type-specific, whereas the 7S (immunoglobulin G) antibodies were always broadly cross-reactive in HI tests. These results confirm that the envelope antigen is the principal antigen involved in the stimulation of protective neutralizing antibodies and contains both type- and group-specific antigenic determinants. The results also establish that there is no significant advantage in using purified RHA or SHA either for immunization or as hemagglutinin antigens in attempts to obtain greater specificity in the HI test. No differences were detected in the antibody responses to infective Kunjin virus, within the range 1,400 to 10(9) plaque-forming units (PFU). Below 1,400 PFU, there was no detectable response. Inactivated virus (10(6) PFU) also stimulated the normal antibody response. In contrast, small envelope fragments (derived with 0.2% DOC) and a detergent-solubilized extract of infected cells were unable to stimulate a detectable antibody response and the small envelope fragments may have induced low dose tolerance in one of two rabbits.

Animals

Unique peptide maps of the three largest proteins specified by the flavivirus Kunjin.

Tryptic digests of four polypeptides found in Kunjin virus-infected Vero cells, NV5, NV4, V3, and NV3, were compared by peptide mapping. The polypeptides to be analyzed were labeled with radioactive methionine and separated by electrophoresis through polyacrylamide gels containing sodium dodecyl sulfate. Because infection of Vero cells by Kunjin virus does not inhibit host cell protein synthesis, radioactively labeled viral polypeptides prepared from infected cells migrate coincidentally during sodium dodecyl sulfate-gel electrophoresis with some of the labeled host proteins. Thus, the genuine viral methionine-containing peptides in tryptic digests of viral proteins have been identified by co-analyzing polypeptides from [3H]methionine-labeled uninfected cells and [35S]methionine-labeled infected cells and determining the 35S/3H ratio in the peptides resolved in two dimensions on thin-layer chromatography plates. The peptide map of NV3 demonstrated that it is host coded, whereas NV5, NV4, and V3 have unique peptide maps and, therefore, account for approximately one-half of the coding potential of Kunjin virus RNA.

Arboviruses

Effects of tilorone hydrochloride on experimental flavivirus infections in mice.

The effect of tilorone hydrochloride on experimental infections of mice with West Nile, Langat and Dengue type 2 viruses were studied. The drug significantly reduced the mortality after intraperitoneal inoculation of West Nile virus, but failed to affect intracerebral infection with Langat or Dengue viruses. Tilorone partially reversed cyclophosphan-enhanced mortality after extraneural inoculation of West Nile and Langat viruses. The possible modes of action of tilorone, interferon-inducing ability and stimulation of humoral immunity, are discussed.

Animals

[2 aspects of immunity in experimental flavivirus infections].

Experimentally, two aspects protective and damaging, of the immune response were demonstrated in the course of development of infection in mice inoculated with tickborne encephalitis (TBE) virus, Langat, yellow fever, dengue type 2 or West Nile viruses. The experiments were carried out in animals in which the functions of T- and B-lymphocytes, were temporarily inhibited with cyclophosphane (CP). It was demonstrated that the protective or damaging role of the immune response depended on the method of inoculation of the animals, the virus properties, and characteristics of the mouse strain. The conditions optimal for the development of immunopathological reactions in one infection (TBE) were not identical for those in another infection even when caused by an antigenically related virus (LANGAT). In mice of the AKR strain the possibility of producing a therapeutic effect upon treatment with CP of the animals inoculated with TBE virus was demonstrated.

Animals

Cross reactions among flaviviruses in macrophage migration inhibition assay.

Adult Swiss albino mice were immunized intravenously with flaviviviruses. Cellular immune response was studied by macrophage migration inhibition assay. Migration of cells from sensitized mice was significantly inhibited in the presence of homologous virus or antigen. Migration inhibition was also significant with serologically related viruses, thus establishing cross reaction in cellular immune response.

Animals

Serologic examination of human population in South Moravia (Czechoslovakia) on the presence of antibodies to arboviruses of the Alfavirus, Flavivirus, Turlock groups and Bunyamwera supergroup.

A statistically representative group of human population from the South Moravian region (CSSR) was examined on the presence of antibodies against the following arboviruses: Lednice, Sindbis, tick-borne encephalitis, West Nile, Tahyna and Calovo. The ascertained absence of antibodies to the Lednice virus is an evidence that at present this virus is of no epidemiological importance in the region studied. No antibodies to Sindbis, West Nile and Calovo viruses were demonstrated. 17.8 to 42.0% of antibodies to Tahyna virus were detected in all age groups examined. The total infection rate was 26.0%. Antibodies to the tick-borne encephalitis virus occurred primarily in persons over 40 years. The total infection rate was 5.5%.

Adolescent

[Change in the nature of the persistence of alpha- and flaviviruses in the body of BALB/c strain mice with a mixed infection with leukovirus].

The features of pathogenesis of infection caused in adult Balb/c mice intraperitoneally infected with Sindbis virus, virulent or attenuated strains of West Nile (WN) virus, individually or in combination with Rauscher leukemia virus (RLV) were studied. The influence of the latter on the course of togavirus infections was characterized by 3 features: (a) different effects on the visceral and neural phases of the pathogenesis (increased period of viremia and virus reproduction in the viscera did not lead to stimulation of virus reproduction in the CNS); (b) changes in the time of togavirus persistence in the infectious form; (c) the dependence of the observed effect on the togavirus properties.

Animals