[The study of immune structure of Astrakhan region population to West Nile virus in 1998 and 1999]. / Izuchenie immunostruktury naseleniia Astrakhanskoi oblasti k virusu Zapadnogo Nila v 1998 i 1999 gg.

Immunostructure of the Astrakhan Region population to West Nile fever (WNF) was studied in the preepidemic period (1998) and after the outbreak (1999). Among the sera obtained in 1998, 63 (26.3%) were positive in neutralization reaction, 84 (27.1%) in enzyme immunoassay IgG and 20 (7.8%) in HAIR. IgM-antibodies were found in none of 142 samples. Overall number of donors having antibodies to WNF virus by three reactions reached 97 (31.6%). In the sera obtained in 1999, virus neutralizing antibodies were detected in 72(44.4%) cases, specific IgM antibodies detected by EIA_in 5(3.1%), IgG_in 44(27.1%), antihemagglutinins_in 11 (6.9%). The number of positive findings in 4 reactions in 1999 was 81(50.0%). The results of examination of the sera collected for two years (1998 and 1999) were the following: of 402 samples examined in NR positive were 135(33.6%), of 304 five (1.6%) were IgM positive, 128(27.1%) of 472 were IgG positive, and 31(7.4%) of 417 responded in HAIR. Overall index of humoral immunity for 2 years was 37.9% (in males and females 39.8 and 32.8%, respectively. In persons aged 20-29 years_36.9%, 50-59 years_42.9%. Thus, by 2-year results, antibodies to WNF virus occurred in 51.9% rural citizens and 35.0% city population.

[Characteristics of humoral immunity to rubella virus in residents of Moscow in 1998-1999]. / Pokazateli gumoral'nogo immuniteta k virusu krasnukhi u zhitelei Moskvy v 1998-1999gg.

A total of 400 blood samples were collected from residents of Moscow in 1998-1999, 369 from adults aged mainly 19-31 years and from children aged 5-12 years. The mean incidence of antirubella antibody was 76.5%; the value varied in different age groups. The highest percentage of antibody detection was observed in men aged 19-25 years (91.8%) and the lowest in pregnant women aged 18-38 years (66-67%). Estimation of antihemagglutinin titers in international Units showed that antibody titer 1:40 and higher protected from infection; such titers were detected in 73.6% examinees. In pregnant women the level of immunological defense was very low (only 56%), which necessitates urgent vaccination of adolescent girls and young women. The results of EIA were compatible with the results of indirect hemagglutination test (IHAT), but EIA was much more sensitive in cases with low titers of IHAT.

[Virus-like particles as a source for obtaining antigens for diagnosing rubella]. / Virusopodobnye chastitsy kak istochnikh polucheniia antigenov dlia diagnostiki krasnukhi.

Rubella diagnostic agents for hemagglutination inhibition (HI) and enzyme immunoassay (EIA) based on rubella virus-like particles (RVLP) have been developed. Noninfectious RVLPs containing three structural E1, E2, and C proteins were expressed in transfected CHO24S cell culture. HI titer in culture medium was 1:256. Tween-80 treatment and ether increased HI titer 4-6-fold and rendered the antigen higher stability. Immunogenic properties of RVLPs were similar to the native rubella virus in HI test with international reference human rubella serum and sera from convalescents after rubella. Serum of mice immunized with RVLP reacted similarly with RVLP antigens and native virus. Antigen for EIA from RVLP was prepared by concentrating RVLP from culture fluid and partial purification by ultracentrifugation. The results of human sera testing by HI and EIA with RVLP and native virus antigens coincided. RVLP are identical to antigenic structure of the virus, are stable and easily purified, and can therefore be used for commercial production of HI and EIA antigens.

[Genetic analysis of West Nile fever virus, isolated in the south of the Russian plain (Volgograd and Astrakhan regions) in 1999]. / Geneticheskii analiz virusov likhoradki Zapadnogo Nikia, vydelennykh na iuge Russkoi ravniny (Volgogradskaia i Astrakhanskaia oblasti) v 1999 g.

Two strains of West Nile virus, Vlg 27889 and Ast 986, were isolated from the brain of a dead man and from the blood of a patient, respectively, during an outbreak of serous meningitis and meningoencephalitis in July-September, 1999, in the Volgograd and Astrakhan regions. Analysis of parts of genome of the strains cloned from cell culture by reverse transcription and polymerase chain reaction demonstrated their identity and appurtenance to group I West Nile viruses.

[Isolation of the West Nile Fever virus from human patients during an epidemic outbreak in the Volgograd and Astrakhan regions]. / Vydelenie virusa likhoradki Zapadnogo Nila ot bol'nykh liudei v periodépidemicheskoi vspyshki v Volgogradskoi i Astrakhanskoi oblastiakh.

Two strains of West Nile virus LEIV 27889 Vig and Ast 986 were isolated from the brain of a dead subject and from the blood of a patient, respectively, during an outbreak of serous meningitis and meningoencephalitis in July-September, 1999, in the Volgograd region, Krasnodar territory, and Astrakhan region. These strains reacted with convalescent sera in hemagglutination inhibition test, which proves their etiological role in this outbreak.

[Epidemic outbreak of meningitis and meningoencephalitis, caused by West Nile virus, in the Krasnodar territory and Volgograd region (preliminary report)]. / Epidemicheskie vxpyshki meningita i meningoéntsefalita v Krasnodarskom krae i Volgogradskoi oblasti, vyzvannye virusom Zapadnogo Nila (Predvaritel'noe soobshchenie).

Sera from 102 inpatients from the Volgograd region (64) and Krasnodar region (38) were tested for antibodies to West Nile (WN) virus in hemagglutination inhibition (HI) test and for IgM and IgG antibodies in enzyme immunoassay (EIA). Diseases etiologically associated with WN virus were diagnosed in 81 patients: in 50 out of 64 in the Volgograd region and in 31 out of 38 in the Krasnodar region, which makes 79.4%. Specificity of antibodies to WN virus was confirmed in HI and EIA with WN antigens, related flaviviruses (Japanese encephalitis and yellow fever), and Sindbis alfavirus. A considerable number and the incidence of WN infection suggest that an epidemic caused by WN virus occurred in the Krasnodar and Volgograd regions in summer 1999.

[The effectiveness of lanthanide immunofluorescence and immunoenzyme analyses in differentiating viruses of the tick-borne encephalitis complex]. / Effektivnost' lantanidnogo immunofliuorestsentnogo i immunofermentnogo analizov pri differentsiatsii virusov kompleksa kleshchevogo éntsefalita.

Comparison of the efficacy of time-resolved fluoroimmunoassay (TR-FIA) and two variants of enzyme immunoassay: conventional (EIA-1) and one using biotin-streptavidin system (EIA-2), in differentiation of viruses of the tick-borne encephalitis complex showed that, their specificity being similar, the TR-FIA method was 2-4 times as sensitive as EIA-2 and 8-32 times as sensitive as EIA-1. When the reactivity of the antigens was assessed by titers, the differentiating capacity was found to be similar and to depend upon the specificity of "sandwich"-forming antibodies and the sensitivity of the immunoassay. When the reactivity of the antigens was assessed by relative reactivity in 4 test systems of different compositions, the differentiating capacity of TR-FIA was higher than that of EIA-1 and EIA-2 which allowed additional differentiation of 2 groups of viruses on the basis of qualitative characteristics of reactivity.

[A monoclonal antibody study of the receptor area of the Venezuelan encephalitis virus]. / Izuchenie retseptornoi oblasti virusa venesuél'skogo éntsefalita loshadei pri pomoshchi monoklonal'nykh antitel.

The receptor region for virus-cell interaction in Venezuelan equine encephalomyelitis (VEE) and Eastern equine encephalomyelitis (EEE) viruses was studied using a panel of 17 monoclonal antibodies (MCA). They were able to block agglutination of goose erythrocytes. The dominant role of glycoprotein E2 in the formation of viral receptor for EEE and VEE viruses was demonstrated. Competitive radioimmunoassay identified three antigenic sites in this region. These sites were also responsible for virus neutralization. MCAs to these sites protected outbred mice against lethal infection. The presence of a highly conservative region in VEE (site E2-3) and EEE (site E2a) which produced cross-reacting antibodies blocking hemagglutination of Western equine encephalomyelitis, Semliki Forest, Sindbis, Getah, Aura, Chikungunya, and Pixuna viruses was established. A hypothesis is suggested concerning the existence of similar regions for the entire alphavirus genus, and the role of this region in virus-cell interaction.

[The demonstration of the Venezuelan equine encephalomyelitis virus by the method of indirect lanthanide immunofluorescent analysis]. / Indikatsiia virusa venesuél'skogo éntsefalomielita loshadei metodom nepriamogo lantanidnogo immunofliuorestsentnogo analiza.

A test system of indirect time-resolved fluoroimmunoassay (TR-FIA) was developed and tested on an alpha-virus, Venezuelan equine encephalomyelitis virus. The indirect TR-FIA test system was shown to be highly effective in the detection of antigens of this virus. Not differing in specificity from the direct TR-FIA, the newly developed test system was 4 times as sensitive.

[The immunity generated in white rats vaccinated with strains of the Venezuelan equine encephalomyelitis virus]. / Immunitet, sozdavaemyi u belykh krys, vaktsinirovannykh shtammami virusov venesuél'skogo éntsefalomielita loshadei.

The existence of virulence gradient in the main members of Venezuelan equine encephalomyelitis virus complex (VEE) was established by subcutaneous inoculation of immunologically competent outbred rats weighing 50-70 g. The virulence of the strains may be judged by a parameter such as the weight of the animal in 5 or 10 days after inoculation. The highest degree of protection was observed in the animals immunized with strain 15, the least in those immunized with strain 230. The above results indicate that adult white rats may be used as a model for the evaluation of the efficacy of protective preparations against viruses of the VEE complex.

[The differentiation of viruses in the Venezuelan equine encephalomyelitis complex by using monoclonal antibodies and lanthanide immunofluorescence analysis]. / Differentsiatsiia virusov kompleksa venesuél'skogo éntsefalomielita loshadei s primeneniem monoklonal'nykh antitel i lantanidnogo immunofliuorestsentnogo analiza.

Potentialities of differentiation between Venezuelan equine encephalomyelitis (VEE) complex viruses by time-resolved fluoroimmunoassay and enzyme immunoassay were studied. For this, 4 test systems were used based on different combinations of native and labeled polyclonal antibodies to VEE virus, strain Trinidad, and monoclonal (MCA) antibody MAK 14-7 to protein EL of this virus. The maximal sensitivity and specificity was achieved in the test system formed from native MCA MAK 14-7 for sensitization of the solid phase and labeled polyclonal immunoglobulins for demonstration of the test results. This combination of antibodies allowed to differentiate the epidemic variant of VEF/Trinidad (IA) from epizootic variants of Mucambo (III), Pixuna (IV) and attenuated strain No. 230.

[The design and trial of conjugates for performing lanthanide immunofluorescence analysis]. / Konstruirovanie i aprobatsiia kon''iugatov pri postanovke lantanidnogo immunofliuorestsentnogo analiza.

The possibility of using a number of complexons for labeling of antibodies to Venezuelan equine encephalomyelitis virus and to adenovirus with europium ions was studied. The resultant conjugates, irrespective of the type of complexon, were shown to retain their immunochemical activity and could be used for lanthanide immunofluorescence analysis of virus-specific antigens.

[The use of sectional polystyrene plates in the set-up of lanthanide immunofluorescence analysis]. / Ispol'zovanie razbornykh polistirolovykh planshetov pri postanovke lantanidnogo immunofliuorstsentnogo analiza.

The authors have examined the possibility of using sectional polystyrene plates, made in this country, in time-resolved fluoroimmunoassay (tr-FIA) with Venezuelan equine encephalomyelitis and tick-borne encephalitis arboviruses, and with influenza A virus. The plates presensitized with specific antibodies were found fit for the detection of the antigens of the above viruses. These plates are not recommended for the detection of influenza A virus-specific proteins adsorbed directly onto the microplate surface.

[Monoclonal antibodies cross-reacting with the tick-borne encephalitis virus and with the Venezuelan equine encephalomyelitis virus]. / Monoklonal'nye antitela, perekrestno reagiruiushchie s virusom kleshchevogo éntsefalita i virusom venesuél'skogo éntsfalomielita loshadei.

Employment of radioimmunoassay led to the demonstration of serological crossing between tick-borne encephalitis (TBE) virus and Venezuelan equine encephalomyelitis (VEE) virus. Using hybridoma technology, three hybridomas were produced secreting monoclonal antibodies (MAb) cross-reacting with these two viruses. With MAb, the epitope of binding of these antibodies was shown to be located on protein E of TBE virus and protein E1 of VEE virus. Despite the low percentage (14%) of homology of amino acid sequences of these proteins, 12 areas with homology from 24% to 63% were demonstrated. Considering conservative replacements, homology of these areas was 53%-75%. The assumed existence of some of these areas in alpha-helical conformation may explain the observed immunological crossing.

[Virological study of cases of sandfly fever in Afghanistan]. / Virusologicheskoe izuchenie sluchaev moskitnykh likhoradok v Afganistane.

Three strains (Af-1008, Af-1038, and Af-130) of Neapolitan sandfly fever virus (NSFF) and 2 strains (Af-1028 and Af-83) of Sicilian sandfly fever virus (SSFF) were isolated from febrile patients among soldiers of the limited military contingent of Soviet troops in Afghanistan in May-August, 1986-1987. The viruses were isolated in neonate mice and identified by indirect immunofluorescence, complement-fixation tests and plaque reduction neutralization test in Vero E6 and SPEV cells. The new NSFF strains in serological tests differed slightly from the original Sabin strain and completely differed from Toskana virus. In the examinations of 104 paired sera from convalescents, 87 subjects showed a rise in antibody titres either to NSFF or to SSFF virus, and 16 patients had antibodies to both viruses but without any rises in titre. Among 60 subjects with a presumable diagnosis of sandfly fever 12 persons had an antibody No information on isolation of sandfly fever viruses in Afghanistan has been available hitherto.

[Antigenic analysis of tick-borne encephalitis virus group using monoclonal antibodies and immunofluorescence]. / Antigennyi analiz virusov gruppy kleshchevogo éntsefalita s ispol'zovaniem monoklonal'nykh antitel i immunofliuorestsentsii.

The study included 18 monoclonal antibodies (MAb) to E- or NS1-antigens tested by immunofluorescence with tick-borne encephalitis (TBE) complex viruses. MAb were induced to 3 strains of TBE virus: the pathogenic 4072 strain isolated from a patient; the Skalica strain of low pathogenicity; and the Neidorf strain isolated from ticks. According to their reactivity to complex viruses, MAb comprised 3 groups: monospecific for TBE virus (T6, T15) which detected tick-borne encephalitis virus alone; widely cross-reactive with 4-6 viruses of the complex (NEK, KEN, T7, T9); and partially complex-reactive (T11, T12, T13, T33/3) and bound to 2-3 viruses of the complex. T13 and T33/3 MAb reacted with the Omsk hemorrhagic fever virus to the same degree or stronger than with TBE virus. The cross-reactivity was more marked in anti-E-than in anti-NS1 MAb. The similarity of the Langat viruses and the Skalica strain was confirmed. Using anti-NS1 MAb in tests with non-fixed cells, the release of NS1-antigen was found to begin at hour 18 (time of observation). The results of the study may be useful for improvement of laboratory diagnosis of TBE and evaluation of the capacity of a vaccine to induce cross immunity to viruses of the TBE complex.

[Hybridomas producing monoclonal antibodies to Crimean hemorrhagic fever virus]. / Gibridomy, produtsiruiushchie monoklonal'nye antitela k virusu Krymskoi gemorrhagicheskoi likhoradki.

A series of hybridomas producing monoclonal antibodies to Crimean hemorrhagic fever virus was obtained and their cultural properties were characterized. HEMA-12 and HEMA-24 secreted monoclonal IgG2b antibodies, HEMA-101 secreted monoclonal IgG1 antibodies, HEMA-31, HEMA-9 and HEMA-11 secreted monoclonal IgG2 antibodies. According to the results of the indirect immunofluorescence test, the titer of specific immunoglobulins in the culture fluid was 1:16-1:32, but sometimes reached 1:64-1:128. The titer of antibodies in ascitic fluid amounted to dilutions of 10(4)-10(5). Hybridomas were cloned by the method of ultimate dilutions. The injection of 5-15 million HEMA cells into the abdominal cavity of BALB/c mice induced the formation of ascitic tumors in the animals within 7-11 days and the accumulation of ascitic fluid in a volume of 1-4 ml. Hybridomas, found to be capable of passage from mouse to mouse, underwent 5-16 passages during the term of observation.