GAG-binding variants of tick-borne encephalitis virus.

Previously different authors described various flavivirus mutants with high affinity to cell glycosaminoglycans and low neuroinvasiveness in mice that were obtained consequently passages in cell cultures or in ticks. In present study the analysis of TBEV isolates has shown existence of GAG-binding variants in natural virus population. Affinity to GAG has been evaluated by sorption on heparin-Sepharose.

Microevolution of tick-borne encephalitis virus in course of host alternation.

Two tick-borne encephalitis (TBE) virus variants were studied: mouse brain-adapted strain EK-328 and its derivate adapted to Hyalomma marginatum ticks. The tick-adapted virus exhibited small-plaque phenotype and slower replication in PEK cells, higher yield in ticks, decreased neuroinvasiveness in mice, increased binding to heparin-sepharose. A total of 15 nucleotide substitutions distinguished genomes of these variants, six substitutions resulted in protein sequence alterations, and two were in 5'NTR.

[Change in the antigenic structure of surface glycoprotein E of tick-borne encephalitis virus during its adaptation to ticks and mammals].

Enzyme immunoassay and immunoprecipitation test of viral proteins, by applying a tick-borne encephalitis (TBE) viral protein E monoclonal antibody kit, have shown that TBE adaptation to ticks and mammals may cause a considerable change in the antigenic structure of surface glycoprotein E, by involving different antigenic domains, including a neutralizing epitope that seems to participate at the early stages of interaction of virions with the cell membrane.

[Residual neurovirulence of the chimera of Langat and Denge-4 flaviviruses in intracerebral infection of monkeys].

The residual neurovirulence of the previously obtained chimeric virus Tp21/DEN4, containing the RNA region encoding the pre M and E structural enzymes, strain Tp21, virus Langat, and the remaining part of the genome from the Denge 4 virus, was studied in experiments with monkeys Cercopithecus aethiops involving the intracerebral administration of the virus. The tick-borne encephalitis virus, strain Absettarov, was used as positive control. A comparative analysis of the experimental and published data showed the chimeric virus to be less virulent by its degree of morphological affection in the CNS zones, its spread into the CNS and by a percentage share of animals with viremia versus the Tp21 parent strain and Elantsev strain of the Langat virus.

Phylogenetic relationships of flaviviruses correlate with their epidemiology, disease association and biogeography.

Phylogenetic analysis of the Flavivirus genus, using either partial sequences of the non-structural 5 gene or the structural envelope gene, revealed an extensive series of clades defined by their epidemiology and disease associations. These phylogenies identified mosquito-borne, tick-borne and no-known-vector (NKV) virus clades, which could be further subdivided into clades defined by their principal vertebrate host. The mosquito-borne flaviviruses revealed two distinct epidemiological groups: (i) the neurotropic viruses, often associated with encephalitic disease in humans or livestock, correlated with the Culex species vector and bird reservoirs and (ii) the non-neurotropic viruses, associated with haemorrhagic disease in humans, correlated with the Aedes species vector and primate hosts.

[Vergina-like strains of tick-borne encephalitis in Russia].

The Greek strain Vergina representing an individual third serotype of tick-borne encephalitis (TBE) virus has been compared with 13 TBE strains isolated on the territory of Russia and Central Asia (in Kirghizia). A kit of deoxyoligonucleotide probes complementary to genome sites of Neudorfle strain of the TBE Central European subtype (protein C and prM genes) and of strain Sofyin of the Eastern subtype (protein E, C, M, prM, ns 1, ns 2b, ns 4b genes) was used in molecular hybridization of nucleic acids. Vergina strain was referred to the genetic variant VI prevalent in the western part of the East European plain, in Udmurtia, in the Altai mountains, and in West Siberia.

[Genomic changes in strains of the tick-borne encephalitis virus as a result of passages in mice].

Changes in genomes of TBE strains isolated from various sources at early stages of laboratory adaptation to white mice brain were demonstrated by molecular hybridization of nucleic acids with synthetic oligonucleotide probes, complementary RNA sites of reference TBE strains Sofyin and Neudorf. Of the 4 TBE strains passed 6 times through white mice brain in only 1 the level of RNA hybridization with 2 oligonucleotide probes was found changed, whereas in all the 3 tested strains the level of RNA hybridization with 1-2 oligonucleotide probes complementary to strain Sofyin was increased after 7-16 passages in the same system. The most noticeable changes in the genome were detected during readaptation to white mice of a TBE variant selected in passages on H.

[The electrophoretic mobility of the virus-specific proteins of tick-borne encephalitis virus strains isolated in different geographical regions of the CIS Nations].

Tick-borne encephalitis (TBE) virus strains isolated in different geographic areas were characterized by changes in electrophoretic mobility (EPM) of virus-specific proteins. Analysis of EPM of intracellular virus-specific protein E and other high molecular proteins of 42 TBE virus strains revealed similarity among the majority of them, with the exception of several strains including the Sophyin strain (Far-Eastern subtype). The most marked variability in the electrophoretic behavior of virus-specific proteins was observed with low molecular proteins prM, ns4b, ns2a.

[The phenomenon of antigenic defectiveness in naturally circulating strains of the tick-borne encephalitis virus and its possible connection to seronegative forms of the disease].

Ten strains of tick-borne encephalitis (TBE) virus isolated from single specimens of I. persulcatus ticks were studied. The strains were divided into antigenically complete (AC) and antigenically defective (AD), depending on the presence or absence of some virus antigens in concentrated virus preparations, characteristics in rocket immune electrophoresis (RIEP), rate and intensity of humoral immune response in monkeys and rabbits, and plaque size in SPEV cell culture.

[The properties of the particles formed in the reproduction of an acute in vitro infection with the tick-borne encephalitis virus adapted to H. plumbeum ticks].

The properties of virions produced in pig embryo kidney (PEK) cells inoculated with tick-borne encephalitis (TBE) virus strain EK-328 which had been passaged in white mice and its variant obtained by passages of TBE in H. plumbeum ticks (clone 718/574 H. pl.

[Changes in the host-dependent characteristics of the tick-borne encephalitis virus during its adaptation to ticks and its readaptation to white mice].

Features of multiplication in pig embryo kidney (PEK) cells of a variant of tick-borne encephalitis (TBE) virus previously selected by passages in H. plumbeum ticks and changes in the properties of the variant upon its repassages in white mouse brains were studied. The tick-adapted TBE variant differed from the original strain in the following characteristics: a lower yield of infectious virus and physical virus particles, altered time-course of infectious virus release from the cell, the lack of virions moving towards cathode in rocket immunoelectrophoresis (RIEP), reduced hemagglutinating activity of the virus, small plaque size in PEK culture, reduced level of virus replication in white mouse brain at a high cytopathic activity of the virus, and low synthesis of virus particles moving towards anode in RIEP.

[Electron microscopic study of a tick-borne encephalitis virus-infected cell culture exposed to tunicamycin].

Ultrastructural features of tick-borne encephalitis virus-infected pig embryo cell culture treated with different concentrations of tunicamycin at various intervals after infection were studied electron microscopically. The inhibition of glycosylation did not prevent virion formation in the infected cells. At the same time, treatment with tunicamycin led to marked accumulation of virus particles in cisterns and vacuoles of the Golgi complex and to a decrease in the number of virions released into the extracellular space.

[Action of tunicamycin on the infectivity and protein synthesis of the tick-borne encephalitis virus].

Tunicamycin added in concentrations of 0.5-1 microgram/ml into the maintenance medium with tick-borne encephalitis virus (TBE) propagated in PEK cell culture inhibited incorporation of labeled carbohydrates into glycoprotein V3 as well as into virus-specific proteins NV41/2, NV3, p22, p18, p13, and p12 and to a lower degree into proteins NV5 and NV4 irrespective of the time of its addition (immediately or 17 hours after virus adsorption on the cells). In the presence of tunicamycin, incorporation of radioactive amino acids into the intracellular virus-specific proteins was inhibited, and proteins with altered electrophoretic mobility appeared.

[Immunochemical characteristics of the antigens of a variant tick-borne encephalitis virus adapted to Hyalomma plumbeum ticks].

A comparative study of immunochemical properties of a preparation of the virion antigen of a variant of tick-borne encephalitis (TBE) virus adapted to H. plumbeum ticks (clone 718/574) and the parental TBE strain revealed the following features: (1) immunoelectrodiffusion test revealed no immunoprecipitation rockets directed towards cathode in the clone 718/574 preparation; anodic precipitates differed in shape from those of the parental strain; (2) diffuse precipitation in agar showed a reaction of incomplete identity between the virion antigen of clone 718/574 and the parental strain antigen, the clone 718/574 antigen forming only one precipitation band in contrast to two precipitation bands of the parental strain; (3) immune electron microscopy demonstrated a peculiar nature of the halo-overlay by immunoglobulins of clone 718/574 virions differing from that of the parental strain.

[Differences in the electrophoretic mobility of the low-molecular virus-specific proteins of viruses of the tick-borne encephalitis complex].

Analysis of low-molecular virus-specific proteins of all the known members of the tick-borne encephalitis (TBE) complex viruses: TBE (2 strains), Langat, louping-ill, Negishi, Kyasanur Forest disease, Omsk hemorrhagic fever (2 strains), and Powassan, was performed. The cells infected with the above viruses were found to contain low-molecular virus-specific NVX (with the exception of Powassan, Omsk hemorrhagic fever, and Kyasanur Forest disease viruses), NV21/2, NV2, NV11/2, and NV1 proteins. These proteins (except NV1) differ in the electrophoretic mobility in the viruses under study and in TBE strains No.

[Characteristics of a cell-culture infection with the tick-borne encephalitis virus with periodic replacement of the culture medium].

The following features were revealed in pig embryo kidney cell cultures infected with tick-borne encephalitis (TBE) virus in which the maintenance medium was changed periodically: more rapid accumulation of TBE virus-specific proteins, active proliferation of spherical vesicular structures and membrane elements of the endoplasmic reticulum, transformation of the latter into regularly arrayed complexes, a statistically significant increase in the yield of the infectious virus.

[Comparative study of the oligopeptide maps of virus-specific proteins of the viruses of the tick-borne encephalitis complex].

A comparative analysis of tryptic hydrolysates of virus-specific proteins NV5, V3 and NV3 of the tick-borne encephalitis complex viruses showed proteins NV5 to have considerable regions of similar amino acid sequences, V3 proteins to have significantly differing primary structures, and NV3 proteins to have different amino acid sequences.

[Comparative analysis of the electrophoretic mobility of the high-molecular virus-specific proteins of flaviviruses].

A comparative study of high molecular virus-specific proteins in cells infected with flaviviruses from 5 serological subgroups was carried out. Proteins NV5, NV4, and V3 were found to have similar electrophoretic mobility in viruses of the tick-borne encephalitis (TBE) complex with the exception of Powassan virus protein. Proteins NV5 and NV4 of mosquito-borne flaviviruses differ in electrophoretic mobility both from the corresponding proteins of the TBE complex viruses and from each other.

[Electron microscopic study of swine embryonic kidney cells infected with the tick-borne encephalitis virus].

Electron microscopic studies of morphological lesions in pig embryo kidney cells (PEK) infected with tick-borne encephalitis (TBE) virus as well as morphology and features of TBE morphogenesis after treatment with actinomycin D, cycloheximide and hypertonic NaC1 concentrations in the medium were carried out. Most marked morphological lesions were observed in the cells after combined effect of high NaC1 concentrations in the medium and inhibitors of protein synthesis. After all kinds of treatment, smooth-contour membrane structures were observed in TBE-infected cells.

[Characteristics of a low-molecular nonvirion ("soluble") antigen from the tick-borne encephalitis virus].

Pig embryo kidney and BHK-21 cells infected with tick-borne encephalitis virus synthesize a nonvirion antigen differing in its immunological properties from the virion antigens. The antigen has a high thermostability. According to the results of ultrafiltration and gel filtration, its molecular weight is approximately 70-100 kilodaltons.

[Heterogeneity of virus-specific flavivirus proteins].

The polyacrylamide gel analysis of large intracellular virus-specific proteins NV5, NV4, and the intracellular form of structural protein V3 established differences in the electrophoretic mobility of each of these proteins formed in cells infected with tick-borne encephalitis, Powassan, Langat, and West Nile viruses. It is assumed that these differences in the electrophoretic mobility of NV5, NV4 proteins, and the intracellular form of V3 protein reflect the differences in the primary structure of each of these proteins in the viruses examined.

[Differences in strains of tick-borne encephalitis virus with respect to the sensitivity of their hemagglutinins to the action of detergents].

Tick-borne encephalitis virus strains (TBE) differ in sensitivity of their hemagglutinating properties (HA) to the detergents: giamine 10-x triton X-100, bridge-58, sodium deoxycholate. These characteristics of different sensitivity to detergents were stable in virus passages in mice and apparently may be used as markers in comparative studies of different strains.

[Oligopeptide mapping of viruses of the tick-borns encephalitis complex].

Comparison of oligopeptide maps of several viruses from the tick-borne encephalitis complex revealed considerable similarity between them manifested in the presence of a common oligonucleotide framework. At the same time there are differences between viruses, each of them having its own oligopeptides lacking in other viruses. The viruses under comparison form groups: tick-borne encephalitis viruses proper, Langat and Powassan viruses, Negishi virus, louping-ill virus.