[Rubella virus genetic determinant of attenuation].

Vaccination is the most effective and available way to prevent Rubella. Presently, 9 vaccine strains were registered. Nevertheless, the molecular mechanisms of the attenuation were poorly elucidated for the rubella virus. However, the study of these mechanisms identifying genotypic and phenotypic markers of attenuation, which together with sequence analysis could be used for the genetic stability control of vaccine strains, is still of current interest. Common trends of genetic changes in the process of adaptation to cold were found due to comparison of nucleic acid and amino acid sequences of the Russian strain C-77 with corresponding positions of the known rubella virus strains and its wild type progenitors, if available.

[Studying of molecular mechanisms of rubella virus attenuation evidence from Russian strain C-77].

Live attenuated rubella vaccine is used for vaccination. Temperature-sensitive (ts) phenotype was proved for almost all rubella vaccine strains, and the acquisition of the ts phenotype during cold adaptation was strongly correlated with the attenuation of the wild-type viruses. Nevertheless, the molecular mechanisms of the attenuation have been insufficiently understood for rubella virus. Study ofthese mechanisms, identifying genotypic markers of attenuation, which together with the sequence analyses could be used for genetic stability control of vaccine strains, is still of current interest. In this work, we determined nearly complete genome sequences of attenuated (ca) and the wildtype progenitor (wt) of the rubella virus strain C-77 isolated in Russia. Possible genetic determinants of attenuation were detected. Thus, 13 nucleotide differences leading to 6 amino acid substitutions were found. Four amino acid substitutions were found to be almost unique. Special consideration should be given to Tyr1042Cys substitution in the protease domain of C-77 strain, because it most probably plays the crucial role in acquisition of ts-phenotype.

[Monoclonal antibodies to rubella virus glycoprotein E1].

AIM: To obtain monoclonal antibodies (MCAs) to glycoprotein E1 of rubella virus, to assess their immunochemical characteristics and ability to use fluorescent MCA for rapid identification of rubella virus. MATERIALS AND METHODS: Rubella virus strain C-74 (Moscow), vaccine strains "Orlov" (Saint-Petersburg), Wistar RA 27/3 (USA) as well as strain Judith (Germany) were used. Viral antigens were obtained using diploid cells L-68 and cell lines VNK-21-F and Vero E6. MCAs were produced by conventional method and their isotype was determined: Immunoblotting, immunoenzume assay (IEA), hemagglutination inhibition assay (HIA) and immunofluorescence assay (IFA) were performed. RESULTS: Five monoclonal antibodies--Kh-252.1, Kh-347.2, Kh-183.3, Kh-214.4, Kh-187.5--to antigens of rubella virus strain C-74 were obtained. Isotypes of these antibodies were determined and their reactivity with native and denaturated antigens of other strains ("Orlov", Wistar RA 27/3, Judith) was characterized. IEA showed that all MCAs interacted with rubella virus glycoprotein E1 at high titers ranging from 1/1600 to 1/200,000. Immunoblotting demonstrated that 4 MCAs (Kh-252.1, Kh-347.2, Kh-183.3, Kh-214.4) had aforementioned feature. MCAs inhibited hemagglutinating activity of Judith strain in titer from 1/16 to 1/1024 in HIA. FITC conjugate of MCA Kh-347.2 (most sensitive variant) allowed to detect rubella virus in infected Vero E6 cells after 24 hours since infection, whereas FITC conjugates of 3 MCAs (Kh-183.3, Kh-214.4, Kh-187.5)--after 72 hours since infection. CONCLUSION: Use of FITC conjugates of MCAs is a perspective tool for identification of rubella virus glycoprotein E1 in infected cell cultures and nasopharyngeal swabs.

[Avidity of immunoglobulin G to rubella virus in post-vaccination and post-infection immunity].

Comparative evaluation of avidity of IgG to rubella virus in vaccinated persons, in patients with rubella or other exanthematous illness, and in healthy persons revealed similar patterns in post-vaccination and post-infection immunity. Virus specific low avidity IgG (index of avidity < or =30%) were detected in patients with rubella during 7 weeks after symptoms appeared as well as in vaccinated persons which were tested 6 weeks after vaccination. Low avidity antibodies in sera were detected in 96% of patients with rubella and in 75% of vaccinated persons which were not immune before immunization. Live attenuated vaccines Ervevax, Priorix, and MMR-II had similar ability to induce low avidity IgG to rubella virus. Increase of low avidity antibodies concentration was noted after immunization of children with low levels of antibodies before vaccination. After immunization of persons with high avidity antibodies in serum, index of avidity remained above threshold. Anamnestic high avidity IgG (index of avidity 51-100%) were detected in majority of immune healthy persons (96.4%) as well as in patients with exanthematous illnesses not related to rubella infection (93.6%). ELISA test-systems for detection of low avidity IgG to rubella virus allow to obtain reliable information about seroconversion rate and characteristics of immune response in vaccines. Detection of low avidity IgG in serum obtained 5-6 weeks after immunization points to primary immune response, whereas identification of high avidity antibodies reveals already immune persons.

[The significance of a mixed congenital viral infection in human antenatal and perinatal pathology]. / Znachenie smeshannoi vrozhdennoi virusnoi infektsii v antenatal'noi i perinatal'noi patologii cheloveka.

Examinations of 202 newborn babies for a representative group of viral infections by detection of viral antigens in cells of urine sediment and in the autopsy materials by indirect immunofluorescence permitted diagnosis of a congenital viral infection in 92% of patients with intrauterine and perinatal pathology; in 72.5% it was a mixed infection. In the patients the virus-virus associations were, as a rule, represented by enteroviruses of Coxsackie group and/or influenza A, B, and C viruses. Most frequently (83.3-100%) mixed virus infection was detected in newborn babies with the severest pathology (meningoencephalitis, encephalitis, sepsis, intrauterine pneumonia), as well as in fatal cases.

[Assessment by immunoenzyme analysis of the immune status of donors with reference to the viruses of rubella, measles and herpes simplex]. / Otsenka v immunofermentnom analize immunnogo statusa donorov k virusam krasnukhi, kori i gerpesa prostogo.

The results of testing the blood sera obtained from donors at a blood transfusion center in Moscow for the presence of antibodies to rubella, measles and herpes simplex viruses, carried out by means of the enzyme immunoassay with the use of the corresponding test systems, are presented. Antibodies to rubella, measles and herpes simplex viruses have been detected, respectively, in 81.5, 96.7 and 100% of blood sera. The proportion of sera with low, medium and high antibody titers has proved to be virtually the same with respect to antibodies to rubella and herpes simplex viruses, the sera with medium antibody titers constituting 59%. At the same time tests for measles antibodies have shown the prevalence of sera with low titers (49.2%) with the highest percentage of seronegative donors (18.5%, as compared with 3.3% in rubella and the absence of negative sera in herpes simplex).

[Difference in the capacity to repair DNA damage in human cells chronically infected with the rubella virus]. / Razlichie v sposobnosti k reparatsii povrezhdenii DNK kletok cheloveka, khronicheski infitsirovannykh virusom krasnukhi.

Centrifugation of cell lysates in alkaline sucrose gradients and chromatography on hydroxyapatite columns were used to demonstrate inhibition of reparation of mitomycin C-induced DNA damages at the stage of reunification of single-strand breaks of DNA in human HEp-2 cell cultures chronically infected with rubella virus. At the same time, reparation of single-strand breaks of DNA caused by bleomycin occurs with similar intensity both in chronically infected and noninfected HEp-2 cultures. The experimental results suggest that the chronic course of infection in human cells leads to disorders in reparative synthesis of cellular DNA and/or is due to disconnected effect of reparation enzymes in this system.

[Antibodies to viruses in children with diabetes mellitus]. / Antitela k nekotorym virusam u detei v dinamike sakharnogo diabeta.

Serological studies were carried out in the time course of insulin-dependent diabetes mellitus in 419 children, among whom paired sera from 66 were studied in the very beginning of diabetes mellitus. By seroconversion in 83% of the children early in the disease, different, frequently mixed virus infections were diagnosed: Coxsackie B2, B3, B4 (46%), rubella (41%), influenza A, B, C (38%), parainfluenza types 1-3 (35%), mumps (23%), adenovirus infection (18%), HB virus infection (4.5%). Acute respiratory diseases preceded or coincided with the onset of diabetes in half of the children with diagnosed acute respiratory and enterovirus infections. No clinical signs of rubella, mumps, or hepatitis immediately before the onset of diabetes or early in the disease were found. A possible role of virus infection diagnosed early in diabetes, in the etiology of chronic insulitis, in provocation of its exacerbation, and manifestation of diabetes mellitus is discussed.

[Properties of rubella virus persisting in cultures of human cells]. / Svoistva virusa krasnukhi, persistiruiushchego v kul'ture kletok cheloveka.

Comparative studies showed rubella virus persisting in chronically infected HEp-2-BK cell culture not to differ significantly from the original strain in the cytopathic effect in RK-13 and BHK-21/13S cells, the interfering capacity with heterologous viruses, and hemagglutinating properties, but to form smaller plaques. The persisting variant and the original virus had low sensitivity to heating at 56 degree C for 10 min. Reproduction of both viruses in RK-13 and BHK-21/32S cell cultures was slightly lower at 40 degree and 36 degree C. At the same time, virus replication in chronically infected cells was not temperature-dependent. The virus variants were shown to be completely antigenically identical by cross-neutralization tests, plaque production, and hemagglutination-inhibition with strain-specific sera.

[Chronic rubella virus-induced infection of human continuous cells]. / Khronicheskaia infektsiia perevivaemykh kletok cheloveka, indutsirovannaia virusom krasnukhi.

A stable chronically rubella virus-infected culture of HEp-2-BK cells existing for over 30 months has been obtained as a result of a single inoculation and further passages. This system is characterized by the lack of cell destruction and permanent production of infectious virus in titres of 2.5-6.3 lg PFU/ml. Synthesis of RNAs of the same classes as in the acute infection was demonstrated in the chronically infected cell culture (CICC) but virion RNA production was less marked. Only a portion of the cells in the population was found to carry the infectious virus. Virus-free cell clones were as susceptible to rubella virus as the control culture. The process of persistence proved to be resistant to virus-specific antibody as well as to the effect of higher (40 degree C) or lower (34 degree C) temperatures. Actinomycin D, 5-bromodeoxyuridine, and 5-iodo-2-deoxyuridine exerted no significant effect on virus reproduction in CICC which, combined with negative results of the experiments on transfection of the susceptible cultures with DNA preparations recovered from CICC, evidenced against the role of integration mechanisms in the establishment and maintenance of the HEp-2-BK system. A clear-cut interference with a heterologous virus was demonstrated in HEp-2-BK culture. The formation of the chronic form of infection in HEp-2-BK culture appears to be due to weak cytocidal properties of rubella virus and formation of endogenous interferon.

[Inhibition of the process of 4-nitroquinoline-1-oxide-induced DNA break repair in chronically virus-infected human cells]. / Ingibirovanie protsessa vossoedineniia razryvov DNK, indutsirovannykh 4-nitrokhinolin-1-oksidom v kletkakh cheloveka, khronicheski indutsirovannykh virusami.

Inhibition of DNA resynthesis after treatment of cell culture with 4-nitroquinoline-1-oxide was found to be due not to the induction of virus mutants repressing this system but to the selection in the cell population of cells predominantly with partially or completely defective system of reparation, or to the development of cellular reparative system because of the presence of viruses in the cell. In chronic infection of HEp-2 cells with tick-borne encephalitis, rubella, and rabies viruses the same phenomenon was observed, namely, inhibition of different stages of the reparation process, i.e. the mechanism of reparation is universal.

[Antibodies to rubella virus in the blood sera and in the synovial fluid of children with rheumatoid arthritis]. / Antitela k virusu krasnukhi v syvorotkakh krovi i v sinovial'noi zhidkosti u detei, bol'nykh revmatoidnym artritom.

Sera and synovial fluids from children with rheumatoid arthritis (RA) were examined for antibody to some viral and mycoplasmal antigens. Blood sera from the patients were found to have a selective excess of antibody to rubella virus as compared with the age norm. Antibody of this specificity was found in synovial fluids in titers significantly higher than those in the blood. Titers of other antibodies in the synovial fluids were, as a rule, slightly lower than in sera. In the time course of intercurrent respiratory diseases, despite the lack of rubella introduction into the wards, children with RA showed variations in the levels of antibody to rubella virus much more frequently than to other infectious agents. The foregoing data are discussed from the point of view of the etiological association of juvenile RA with rubella virus persistence. The antigen of the latter was found in snyovial membrane cells of 4 children by means of immunofluorescence procedure.

[Comparative study of the persistence of 3 RNA-containing viruses in a continuous cell culture of human origin]. / Sravnitel'noe izuchenie ersistentsii trekh RNK-soderzhashchikh virusov v perevivaemoi kul'ture kletok chelovecheskogo proiskhozhdeniia.

A comparative study of chronic infecton of HEp-2 cells with tick-borne encephalitis (TBE), rabies (RV), and rubella (RuV) viruses was carried out. Throughout the entire period of chronic infection (CI) no signs of specific cell destruction by these viruses were observed. The infectious virus was regularly demonstrated in the culture fluid and chronically infected cells. The antigenic properties of the persisting viruses did not differ from those of the original strains. The persisting TBE and rabies viruses replicated in the susceptible cells at a higher temperature and formed plaques of a smaller size than the original virus. The number of chronically infected cells producing infectious virus was always less than the number of cells containing the virus-specific antigen. In all three types of chronic infection the cells supported virus persistence at 40 degrees C. In TBE and RuV chronically infected cells interference with heterologous viruses was marked while in HEp-2-RV homologous interference caused by formation of defective interfering particles was observed. Treatment of the cells with BUDR resulted in activation of the infection only in the HEp-2-TBE system. Experiments on transfection of the sensitive cells by using DNA from HEp-2-RV and HEp-2-RuV gave negative results. The importance of various factors in the mechanism of virus persistence in the chronically infected cells under study is discussed.

[Seroepidemiologic study of immunity to rubella among the population of Tadzhikistan]. / Seroépidemiologicheskoe izuchenie immuniteta k krasnukhe u naseleniia Tadzhikistana.

The first serological survey of the human population of Tajikistan was carried out with the purpose of detection of antibody inhibiting hemagglutination of rubella virus. As a result, in over 2500 persons high intensity of formation of the herd immunity was established. A greater portion of the population (66%) acquired antibody in preschol age. The formation of the immune portion completed by 10--14 years of age. Among women of the child-bearing ages about 8--15% had no antibody to rubella virus. This value in combination with intensive spread of rubella among children indicates that in this republic the epidermiological situation with rubella appears to be unfavourable for adult women.