[Use of plant-origin components in roller cultivation of vaccine reassortant influenza virus strain H5N2].

AIM: To study the optimal conditions for roller cultivation of cold-adapted reassortant vaccine strain of influenza virus A/17/Duck/ Potsdam/86/92 (H5N2) in MDCK and Vero cell cultures grown on nutrient medium based on soy and rice flour hydrolysates obtained using trypsin and bromeline. MATERIALS AND METHODS: Vaccine strain was cultivated on MDCK and Vero cells in rollers in the presence of plant proteases. Obtained culture samples of vaccine strains were lyophilized and their infectivity was assessed. RESULTS: Cultivation of vaccine strain on MDCK and Vero cells grown in experimental media containing reduced quantity (2 and 3% respectively) of fetal calf serum ("Gibco", USA) resulted in high titers of the virus in the presence of plant proteases (4 mcg/ml of papain and 20 mcg/ml bromeline). CONCLUSION: Use of plant enzymes and nutrient media based on enzymic plant hydrolysates, including those obtained with bromeline, for cultivation of vaccine strain on MDCK and Vero cell cultures in rollers could make the manufacturing process of live influenza vaccines safer and more cost effective.

[Immunogenicity of influenza virus H5N2 vaccine strain samples produced by roller cultivation in media with plant derived components].

AIM: Study in CBA line mice of immunogenicity of cold adapted reassortant influenza virus H5N2 vaccine strain samples produced in rollers in MDCK and Vero cell cultures by using plant derived components. MATERIALS AND METHODS: Antibody levels in blood sera and nasal swabs, lungs and small intestine of experimental vaccine strain sample immunized mice were evaluated by using HI reaction in accordance with WHO recommendations. RESULT: Reassortant vaccine strain A/17/duck/Potsdam/86/92 (H5N2) produced in MDCK and Vero cells by using plant derived components (rice and soy flour hydrolyzate and plant protease based nutrient medium) after intranasal immunization of mice induced local and humoral antibodies, and the latter not only against homologous virus, but also against highly pathogenic avian influenza virus strains A/ Chicken/Suzdalka/Nov-11/2005 and A/Chicken/Kurgan/05/2005. CONCLUSION: Immunogenicity studies of reassortant influenza virus A/17/duck/Potsdam/86/92 (H5N2) vaccine strain samples cultivated in MDCK and Vero cells by using media with plant derived components in mice show high levels of humoral and secretory immunity.

[Development of new nutrient medium for MDCK and Vero cells based on soy hydrolysate obtained using bromeline and assessment of growth characteristics of influenza virus vaccine strains cultivated on them].

AIM: To develop nutrient medium for MDCK and Vero cells based on soy hydrolysate obtained using bromeline and to assess of growth characteristics of influenza virus vaccine strains cultivated on them. MATERIALS AND METHODS: Physico-chemical characteristics of hydrolysate were assessed according to FS 42-3874-99. Growth characteristics of nutrient medium based on soy hydrolysate and vaccine strains of influenza virus A/Solomon Islands/03/06 (H1N1), A/Wisconsin/67/2005 (H3N2) and B/Malaysia/2506/2004 were studied on MDCK and Vero cells. RESULTS: MDCK and Vero cells grew well on medium based on soy hydrolysate obtained using bromeline with decreased (to 2% and 3% respectively) content of fetal calf serum and allowed effective production of vaccine strains of influenza virus. CONCLUSION: Technology for producing of nutrient medium based on hydrolysate of soy flour obtained using bromeline was developed. This medium could successively used for cultivation of continued cell cultures MDCK and Vero used as substrate for tissue culture-based vaccines against influenza.

[Study of efficacy of anaferon pediatric in mice infected by pandemic influenza virus A(H1N1/09)v].

AIM: To study efficacy of anaferon pediatric in mice infected by pandemic influenza virus A(H1N1/09)v. MATERIALS AND METHODS: Influenza virus strain A/California/07/2009 (H1N1)v was used. Three groups of BALB/c mice intranasally inoculated with influenza virus were studied. First group received solution of Anaferon pediatric during 5 days before and 8 days after inoculation, 2nd group received Tamiflu during 5 days after inoculation. Distilled water was administered orally to mice from control group. RESULTS: It was shown that Anaferon pediatric used as preventive and treatment agent in mice intranasally inoculated with 100% infectious dose of influenza virus strain A/ California/07/2009 (H1N1)v had antiviral effect, which expressed in 10-fold decreased reproduction of influenza virus in lungs of infected mice compared to control group measured 4, 6, and 8 days after inoculation. CONCLUSION: Use of anaferon pediatric before and after inoculation with influenza virus A(H1N1/09)v was not less effective than use of Tamiflu after inoculation.

[In vitro and in vivo efficacy of Ingavirin against strains of pandemic influenza virus A(H1N1/09)v].

AIM: To study efficacy of Ingavirin in vitro and in vivo against strains of pandemic influenza virus A(H1N1/09)v and influenza virus A(H5N1) and A(H3N2). MATERIALS AND METHODS: Changes in hemagglutinating and cytopathic activity of influenza virus strains A(H1N1/09)v, A(H5N1) and A(H3N2) during their incubation in the presence of Ingavirin or Remantadin on MDCK cell culture were studied. In mice infected by influenza strains A(H1N1/09)v and A(H3N2) and orally treated with Ingavirin, Tamiflu or Remantadin virus titers in lungs were measured. RESULTS: There was decrease in hemagglutinating and cytopathic activity of influenza virus strains after incubation with Ingavirin in vitro. Ingavirin effectively inhibited reproduction of influenza virus strains A(H1N1/09)v and A(H3N2) in lungs of infected mice. Titers of these strains in lung homogenates decreased when Ingavirin was orally administered to infected mice. CONCLUSION: Strains of influenza virus A(H1N1/09)v were susceptible to Ingavirin and Tamiflu but resistant to Remantadin. Reference strains of A(H5N1) and A(H3N2) were susceptible to Ingavirin, Tamiflu and Remantadin.

[Genetical features of influenza virus A (H1N1) strain that caused the 2009 pandemic].

Genetical features of the A(H1N1) influenza virus strain that caused the 2009 pandemic are analyzed in the review. Mutations typical for this strain, unique and similar to influenza viruses of swine, avian and seasonal types, and phenotypic (pathologic) features associated with them, that are experimentally confirmed, are described. A possibility of reassortation of avian and swine influenza viruses and possible epidemiologic consequences are discussed.

[Study of antiviral activity of extracts obtained from basidial fungi against influenza viruses of different subtypes in experiments in vitro and in vivo].

AIM: To study antiviral activity of extracts obtained from basidial fungi against influenza viruses of different subtypes. MATERIALS AND METHODS: Antiviral activity of extracts obtained from basidial fungi against influenza virus A/chicken/Kurgan/05/2005 (H5N1) was determined in in vitro experiments. Changes in infectiousness of pandemic influenza virus A/Moscow/226/2009 (HIN1)v caused by extracts of basidial fungi was studied in experiments in vitro and in vivo. RESULTS: Seventy water extracts of basidial fungi were studied, of which 10 were able to inhibit infectiousness of influenza virus strain A/ chicken/Kurgan/05/2005 (H5N1) in MDCK cell culture. Also, several studied extracts decreased infectiousness of pandemic influenza virus strain A/ Moscow/226/2009 (H1N1)v in MDCK cells and inhibit its reproduction in lungs of infected mice. CONCLUSION: High antiviral activity of extracts obtained from basidial fungi against influenza viruses opens perspectives for development of drugs with preventive and treatment effects.

[Properties of CRF02_AG HIV-1 isolates circulating in Novosibirsk region].

AIM: Study of circulating 02_AG recombinant form HIV-1 isolates that have been rapidly spreading in Novosibirsk region during 3 recent years. MATERIALS AND METHODS: WHO protocol for primary HIV isolation was used, automatic sequencer was used for genetic characterization of isolates. Virus specific RNA were isolated and env HIV-1 region DNA fragments were processed. Phylogenetic analysis was also performed. RESULTS: CRF_02AG HIV-1 isolated from peripheral blood of HIV-1 positive patients belonged to CCR5 tropic viruses and had various reproduction characteristics. Most of the HIV isolated were rapidly replicating virus variants characterized by an ability to accumulate high levels of virus protein p24 in cultural fluid. Infectivity and reproductive properties of HIV isolates were confirmed in experimental infection by using clarified cultural liquid of mononuclear cells from healthy donors. Phylogenetic analysis of CRF02_AG HIV-1 variants isolated in Novosibirsk region in 2007 - 2010 showed the formation of a separate outbreak in the area caused by emergence of CRF02_AG HIV-1 in human population. CONCLUSION: A collection of genetically and biologically characterized CRF02_AG HIV-1 isolates that has not been spreading previously in Russia.

[Evaluation of immunogenicity and safety of 2 immunizations with allantoic intranasal live influenza vaccine Ultragrivac].

AIM: Evaluate reactogenicity, safety and immunogenicity in phase 2 clinical trials of 2 immunization schedules with Ultragrivac--an allantoic intranasal life influenza vaccine based on A/17/ duck/Potsdam/86/92 [17/H5] reassortant strain. MATERIALS AND METHODS: 4 groups of volunteers participated in the study: group 1--40 individuals were vaccinated twice with a 10 day interval; group 2--40 individuals were vaccinated twice with a 21 day interval; group 3 (control)--10 individuals received placebo twice with a 10 day interval; group 4 (control)--10 individuals received placebo twice with a 21 day interval. Local (secretory IgA), cellular and humoral immune response were evaluated. Humoral immunity was evaluated by the intensity of increase of geometric mean antibody titers against 2 influenza virus strains A/17/duck/Potsdam/86/92 [17/H5] and A/chicken/Suzdalka/Nov-1 1/2005 (H5N1), and by the level of significant (4 times or more) antibody seroconversions after the vaccination. RESULTS: After the use of Ultragrivac the level of secretory IgA in the nasal cavity of vaccinated volunteers in the groups with revaccination intervals of 10 and 21 days increased significantly. The second immunization with 10 or 21 day intervals significantly increased postvaccinal humoral immune response. Humoral immune response induction after 2 vaccinations with 10 day interval was no less effective than with 21 day interval. CONCLUSION: Ultragrivac allantoic intranasal live influenza vaccine is areactogenic, harmless for vaccinated individuals, safe for those around, and has immunogenic properties against not only homologous virus A(H5N2), but also against influenza strain A(H5N1).

[Studies of properties of pandemic influenza virus A/H1N1 circulated in Russian Federation in 2009 - 2010].

AIM: Studies of cultural, virologic, antigenic properties of 89 samples of pandemic influenza A(H1N1) 2009 virus isolated in Russian Federation from May 2009 to March 2010. MATERIALS AND METHODS: Properties of isolated samples were compared with those of the reference strain A/ California/04/2009 (H1N1). RESULTS: Studies of biological properties and analysis of genome nucleotide sequences of the isolated samples showed that those strains are closely related to the reference strain. CONCLUSION: Monitoring of genetic, virologic and antigenic properties of pandemic influenza A(H1N1) 2009 virus isolates carried out from May 2009 to March 2010 did not reveal significant changes in the abovementioned properties of the virus or emergence of mutations that can lead to such changes.

[Pandemic influenza virus A (H1N1) in Amur region in autumn 2009].

AIM: Isolation and study of molecular genetic characteristics of pandemic influenza virus A (H1N1) circulated in Amur region in autumn 2009 as well as testing of serum samples taken from citizens of this region during November- December 2009 in order to measure levels of antibodies to socially significant serotypes of influenza A virus. MATERIALS AND METHODS: Strain of pandemic influenza virus A/Blagoveschensk/01/2009 (H1N1) was isolated on MDCK cell culture and nucleotide sequences of all eight segments of viral genome were determined. Five hundred seventy-six serum samples taken in Amur region in autumn 2009 were tested by hemagglutination inhibition assay. RESULTS: Nucleotide sequence of A/Blagovechensk/01/2009 (H1N1) strain was 99.7% identical to reference influenza virus strain A/California/04/2009. Diagnostically significant titers of antibodies to pandemic influenza virus were observed in 46.3% of persons younger 30 years old and in 20.1% older persons. Antibodies to seasonal influenza virus H1N1 and H3N2 were detected in 39.5 and 29.8% of persons respectively. CONCLUSION: Final seroepidemiological picture of distribution of pandemic virus in Amur region matches with the one for seasonal influenza virus A (H1N1): > 60% of seropositive persons were registered in age group < 18 years old, and this proportion increases with increasing age.

[In vitro efficacy of Ingavirin against the pandemic influenza virus A(H1N1/09)v].

Ingavirin was shown to be efficient in inhibition of the influenza virus strains A/California/04/2009 (H1N1)v, A/California/07/2009 (H1N1)v, A/Moscow/225/2009 (H1N1)v and A/Moscow/226/2009 (H1N1)v, as well as the strains A/Chicken/Kurgan/05/2005 (H5N1) and A/Aichi/2/68 (H3N2) in the MDCK cell culture. The hemagglutinin and cytopathic activity of the influenza virus strains decreased at entering Ingavirin in vitro.

[In vivo efficacy of Ingavirin against pandemic A (H1N1/09)v influenza virus].

Ingavirin was shown to be efficient in inhibition of the pandemic influenza virus strains A/California/04/2009 (H1N1)v, A/California/07/2009 (H1N1)v, A/Moscow/225/2009 (H1N1)v and A/Moscow/226/2009 (H1N1)v. as well as the influenza virus strain A/Aichi/2/68 (H3N2) in the lungs of the infected mice. After oral administration of Ingavirin the titers of the influenza virus strains in the lung homogenates lowered.

[Experimental evaluation of the biological risks of introduction of the genetically modified microorganism (GMM) B. subtilis VKPM B-7092 into the environment].

Experimental evaluation of the biological risks of introducing the genetically modified microorganism (GMM) B. subtilis VKPM B-7092, an active ingredient of the probiotic VETOM 1.1, into an open system was performed. The following features of the GMM were studied: the survival rate of the GMM in bovine gastroenteric tract; its influence on the microbiocenosis; the species composition of microflora of the gastroenteric tract of the animal species; the possibility of transfer of the DNA fragment cloned in the B. subtilis bacterium and containing the gene of human leukocyte alpha2 interferon to the representatives of intestinal microflora of animals fed on the probiotic VETOM 1.1, as well as the GMM transfer to other microorganism species spread in the areas of potential getting of the GMM into the environment (soil). The study revealed no negative effects of the GMM on the animal organism and the environment, including remote aftereffects.

[Experimental evaluation of the protective properties of some nonwoven materials for respiratory protection means and prospects of their use].

The protective properties of nonwoven materials (Spandbond, SMS) used to manufacture 3-5-layer medical masks, by using model physical and bacterial test aerosols, were experimentally assessed. It was shown that the more layers of the materials, the less permeable they became for test aerosols. Three-five-layer masks made from SMS at a density of 42 g/m2 were found to have higher protective properties for oil mist and fine aerosol than those made from Spandbond at a density of 25 g/m2. Five-layer SMS materials at a density of 42 g/m2 have the highest values of bacterial aerosol retention.

[Evaluation of the protective properties of nonwoven materials for throw-away medical overalls].

The permeability of varying density felts (Spandbond, SMS, Tyvek, Sontara) for throw-away medical overalls was experimentally assessed under statistic and dynamic conditions, by using model test aerosols, including bacterial one. Their permeability was shown to decrease with the higher density of the materials under study. Laminated Span-bond contributes to an abrupt reduce in the bacterial penetration of this kind of tissues and enhances their protective properties. The felts CMC, Tyvek, Sontara, and laminated Spanbond were found to have the highest protective properties. In terms of their barrier and protective properties, the felts surpass the tissues used to make re-usable protective medical overalls.

[Experimental evaluation of possible transmission of human alpha-2 interferon artificial gene to the other microorganism species]. / Eksperimental'naia otsenka vozmozhnosti peredachi iskusstvennogo gena leikotsitarnogo interferona-al'fa2 cheloveka v drugie vidy mikroorganizmov.

Described within the case study are experimental evaluation data on the remote results of introduction of gene-engineered microorganisms (GEM) of the B-7092 B. subtilis strain into the environment with the follow-up ranging from 1 to 7 years. No bacteria of the above GEM were detected (among selected 19 bacterial cultures) in soil samples from an agricultural farm, where the VETOM 1.1 probiotic was used for the treatment and prevention of cattle; primary and precise identification (two-round PCR with specific primers) was applied. The introduction of the B-7092 B. subtilis strain into the environment did not result in its unlimited growth and proliferation in soil. No transfer of plasmid genes (genes of leukocytic human alpha-2 interferon) from the B-7092 B. subtilis strain GEM to microorganism species available in the environment (soil) was registered. No recombinant plasmid DNA from lysed GEM cells was detected in soil samples.

[Experimental study of therapeutical properties and safety of king crab collagenase-containing ointment]. / Eksperimental'noe izuchenie lechebnykh svoistv mazi, soderzhashchei kollagenazu kamchatskogo kraba.

The effects of ointment containing king crab (Paralithodes camtschatica) collagenase on intact skin, thermal, and pyonecrotic wounds were studied in rats by using hematological, biochemical, immunological, and morphological methods. The ointment for the skin and viscera was shown to be safe. It is highly effective in debriding the infected wounds. Different concentrations of collagenase were tested. The concentration of collagenase was recommended to be 0.2 mg/g ointment for use.