Vol 87, No 1 (2010)

Цель. Изучение микрофлоры нижних дыха- тельных путей разных возрастных групп детей, больных муковисцидозом, в динамике для вы- явления ее изменчивости и определения воз- можных источников инфекционных осложне- ний. Материалы и методы. Проанализирована 141 история болезни детей, больных муковис- цидозом, разных возрастных гру пп, прож и- вающих в ра з личны х рег иона х Российской Федерации. В динамике обследованы 84 ре- бен к а, бол ьн ы х м у ковисц и дозом, ж и т елей Москвы и Московской области, проходивших лечение амбулаторно и в стационаре. Для иден- тификации и характеристики микроорганиз- мов использованы микробиологические, мо- лек ул я рно-г енет и ческ ие и стат ист и ческ ие методы исследования. Результаты. Установлено, что хроническая синегнойная, стафилококко- вая или смешанная инфекция начинает диа- гностироваться у 25% детей у же в возрасте 1 — 4 лет, а к 18 годам выявляется у 80% больных м у ковисци дозом. В 2/3 сл у чаев вы явл яется ассоциация микроорганизмов, причем у го- спитализированных больных эти ассоциации представлены в 60% случаев 3 — 5 микроорга- низмами. В составе ассоциаций кроме основ- ны х возбудителей Pseudomonas aer uginosa и Staphylococcus aureus часто встречаются пред- ставители грамотрицательных неферментиру- ющих микроорганизмов — Burkholderia cepacia, Stenotrophomonas maltoph i l ia, Aci netobacter baumanii, что, вероятно, обусловлено тропиз- мом этих видов микроорганизмов к легочной ткани. Заключение. Для больных муковисцидо- зом характерным проявлением является хро- ническая смешанная инфекция. Расшифровка возможных механизмов инфицирования лег- ких больных муковисцидозом позволит раз- работать систему профилактики инфекцион- ных легочных осложнений у таких больных.

Aim. Genotyping of Hib strains isolated in regions of Russia as well as characterization of genetic relations of typed strains with strains isolated in other areas. Materials and methods. Genetic characterization of 31 strains of Hib isolated in Russian regions during 2005 - 2008 was performed by multilocus sequence typing. Results. Studied strains belonged to 11 variants of sequence types, 6 of which were described in previous studies, whereas other 5 were isolated for the first time during this study. The most common isolated strains were ST-92 (13 strains or 42%) and ST-6 (6 strains or 19%). Typed strains were distributed to two clonal complexes. Clonal complex «A1/A2» («ST-6») incorporates all typed strains except ST-93 strain belonging to clonal complex «B1b» («ST-93»). The majority of studied strains (19 or 61%) had difference from «central» sequence type of clonal complex «A1/ A2» («ST-6») on not more than one allele. Conclusion. Clonal structure of isolated strains is analogous to the one observed in Moscow and foreign strains.

Aim. To assess antibody levels against diphtheria, pertussis, and tetanus in pregnant women. Materials and methods. One hundred and two virtually healthy pregnant women aged 18-35 years were studied. Antibodies to diphtheria and tetanus were measured in passive hemagglutination reaction with diphtheria and tetanus diagnostic kits. Antibodies to Bordetella pertussis antigens were determined in hemagglutination assay (HA) and in enzyme immunoassay (EIA). Results. Protective titers of anti-diphtheria and anti-tetanus antibodies were detected in 91.2% and 94.1% of participants respectively, whereas high titers - in 24.5% and 27.4% respectively. Low levels of IgG to B.pertussis antigens measured by EIA were observed in 70.6% of participants whereas moderate and high levels - in 22.5% and 6.9% respectively. Conditionally protective levels of anti-pertussis antibodies measured by HA were detected in 10.8% of participants. Conclusion. Obtained results demonstrate high level of protection of pregnant women against diphtheria and tetanus and low level of anti-pertussis immunity.

Aim. To study features of specific immunity in patients with diphtheria using data from clinic as well as from animal experiments. Materials and methods. Serum samples of 80 patients hospitalized to Infec- tious Diseases Clinical Hospital No. 2 in Moscow and treated with anti-diphtheria serum (manufactured by «Immunogen» Concern, Stavropol) were studied. Localized diphtheria of the oropharynx was diagnosed in 29 patients, diffused diphtheria — in 8, subtoxic — in 3, grade 1 toxic — in 19, grade 2 toxic — in 12, grade 3 toxic — in 9. Experimental part of the study was performed on outbred rabbits weighted 3 — 3,5 kg. Level of antitoxin in serum was measured in reac- tion of passive hemagglutination using commercial antigenic erythrocyte diagnostic kit (manufactured by Mechnikov Research Institute of Vaccines and Sera, Moscow). Results. Intermittent administration of toxin to control rabbits which lack background im- munity did not lead to changes in their immune status and after administration of anti-diphtheria antitoxin kinetics of its level in serum was analogous to that observed after administration of antitoxin to intact animals. Highest level of antitoxin (1.0 — 2.0 IU/ml) was observed 1 — 3 days after its administration, and to day 13 — 15 antitoxin was not detected in serum samples. Diphtheria antitoxin in concentration from 0.03 to 40.0 IU/ml was detected in serum samples in 59 of 80 (74%) studied patients. Only in 21 patients (26%) the antitoxin was not detected. Conclusion. Presence of antitoxin in serum argue for active im- mune response to infection and activation of immune memory mechansisms, which allow to predict less severe course of the disease. Absence of antitoxin in serum of patient admitted to hospital points that infectious process is developing on the background of no immunity that predicts the probable development of severe diphtheria.

Aim. To determine level of Toll-like receptors (TLRs) expression in spleen and lymphatic nodes of mice after immunization by mucosal routes. Materials and methods. Mice were immunized with polycomponent vaccine Immunovac ® either by mucosal or subcutaneous route. Expression of TLRs in spleen, respiratory tract-associated lymphatic nodes as well as in small intestine was measured in immunized mice by flow cytomentry method. Results. After immunization of mice by subcutaneous, intranasal and oral routes level of TLRs expression was different. Significant expression of TLR9 and absence of TLR2 expression was noted after non-parenteral methods of immunization. After oral immunization expression of TLRs was identified in gut- and respiratory tract-associated lymphoid tissue as well as in spleen; after intranasal immunization – in respiratory tract-associated lymphoid tissue, and after subcutaneous immunization — in spleen and respiratory tract-associated lymphoid tissue. Conclusion. After oral immunization expression of TLRs was identified in all studied organs, including spleen. Involvement of spleen to this process allows to assume establishment of not only local but also systemic immunity.

Aim. To demonstrate the possibility of development of test based on phosphorescent analysis (PHOSPHAN) for simultaneous identification and differentiation of specific IgG to tick-borne encephalitis (TBE) and West Nile fever (WNF) viruses. Materials and methods. Twenty six serum samples from patients with TBE, twenty five from WNF, and sixty six from clinically healthy donors were used for the study. Immunologic analysis was performed in plate wells with active microzones «printed» on the wells’ bottom and corresponding the complex of virus-specific antigens with immobilized monoclonal antibodies; internal control of specificity was included in each well. Species specificity of antibodies was determined on the basis of not less than 2-fold elevation of value of positivity coefficient (P/N) of sample studied with homologous antigen compared to heterologous one. Results. PHOSPHAN provides simultaneous detec- tion of IgG in human serum to two related flaviviruses: TBE and WNF viruses. Usage of P/N criterion assessed with homologous and heterologous antigen allowed correct determination of species specificity of antibodies in 90% of serum samples from patients with TBE and WNF. Conclusion. PHOSPHAN allows to detect and differentiate IgG to TBE and WNF viruses during testing of one serum sample in one plate well without decrease of sensitivity compared to enzyme immunoassay with separated testing of samples on two viruses. This provides savings of biomaterial, which is an advantage compared to enzyme immunoassay.

Aim. To study the effect of growth medium content on antagonistic characteristics of «Pashkov» strain of B.pumilus . Materials and methods. The following strains were used in the study: B.subtitlis 3H, B.subtitlis 534, B.subtitlis 1719, B.pumilus «Pashkov». Test strains from collection of Tarasevich State Institute for Standardization and Control of Medical Biological Preparations were used for determination of antagonistic activity. Growing of B.pumilus cultures on medium (No. 9, No. 5, VK-2, Spizeisen, and L-broth) was performed by periodic cultivation during 72 h in conditions of continuous aeration at 37°C. Inhibiting effect of obtained samples was assessed on growth kinetics of test strains using microbiological analyzer with plan-table photometer «BIOSCREEN/ iEMS — reader MF» (LabMetod, Finland). Results. Expressed antagonistic characteristics were revealed during cultivation on media No. 9 and No. 5 with maximal expression on day 3. Addition of lactose to medium No. 9 resulted in increase of bacteriostatic effect (BSE) against both individual strains and all test strains — up to 16% in average. Selection of optimal concentrations of different components allowed to develop a new growth medium which provides maximal antagonistic activity in studied strain of B.pumilus . Conclusion. Effect of growth medium content on antagonistic characteristics against pathogenic and opportunistic bacteria was confirmed on the example of cultivation of spore-forming strain «Pashkov» of B.pumilus . Optimization of carbohydrate, salt and other components of growth medium allowed to increase up to 100% the BSE of «Pashkov» strain of B.pumilus against both individual and all test strains. This study opens perspectives for generation of com- pounds with directional probiotic effect.

Aim. Efficacy of different treatment regimens with equine diphtheria antitoxin (EDA) was assessed on clinical samples as well as in experiments on animals. Materials and methods. Protective properties and serum concentration kinetics of heterologous antibod- ies was studied on 12 rabbits and 51 guinea pigs after intramuscular injection of different doses of EDA, in serum samples from 26 patients, which received one intramuscular injection of EDA in various doses as well as in serum samples from 10 patients with diphtheria of different severity, which were treated with EDA in total course dose 100.000 - 1.500.000 IU. Antitoxin concentration in serum sample was measured with passive hemagglutination assay as well as Jensen toxin neutralization test on rabbits. Results. Experiments on laboratory animals received EDA in dose 150 IU/kg showed high protective effect. For example, rabbits with antitoxin level 1.0 - 1.25 IU/ ml in serum 24 hours after injection of EDA were 50 - 250 times resistant to dermonecrotic effect of diphtheria toxin compared with rabbits not received EDA. Guinea pig with antitoxin level 0.5 - 2.0 IU/ ml in serum 24 - 48 hours after injection of EDA in dose 150 IU/kg were all protected against 35 - 50 LD 50 of diphtheria toxin. After termination of EDA injection there was sharp decrease of antitoxin level and it was not detected in serum 7 days after. Increase of antitoxin level in serum of animals was not adequate to quantity of injected EDA. Study of serum samples from 26 patients received one intramuscular injec- tion of different doses of EDA showed that doses of antitoxin from 20.000 to 30.000 IU resulted in its presence in serum in concentration 0.5 - 3.0 IU/ml whereas injection of 50.000 IU or 70.000 - 100.000 IU resulted in serum concentrations 1.25 - 10.0 IU/ ml and 2.5 - 20.0 IU/ml respectively. Conclusion. Relatively low doses of EDA provided relatively high level of protection against diphtheria toxin that should be taken into account during treatment of diphtheria patients.

Modern version of I.Mechnikov’s hypothesis on association of somatic diseases with infectious agents is presented. List of bacteria and viruses associated with various types of cardiomyopathies, atherosclerosis, gastritis, gastric and duodenal ulcerative disease, type 1 diabetes mellitus. Literature data showing that influenza vaccination reduces number of fatal myocardial infarctions and strokes during winter seasons as well as number of hospitalizations due to exacerbations of chronic cardiovascular and cerebrov- ascular diseases are summarized. Data on probability of coincidence of influenza vaccination and sudden death in elderly persons are reviewed.

Data on development and study of adjuvants for influenza vaccines against avian and seasonal influenza are presented. Requirements to perspective adjuvants, their main groups (mineral salts of aluminium, squalene derivatives, polyglycosides, and immunomodulators) as well as experimental adjuvants are reviewed. Analysis of results of several clinical trials of adjuvanted vaccines against avian influenza is presented. On the basis of published data analysis conclusion about good perspective of influenza vaccines with new adjuvants was made.