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Hand-foot-and-mouth disease (HFMD) is the most common enteroviral infection in South-East Asia. When evaluating the role of enterovirus 71 (EVA71) as an etiological agent of infectious disease in South Vietnam, we revealed a high proportion of EVA71 among identified species A enteroviruses found in 3542 samples from HFMD cases; 125 samples from cases of enteroviral meningitis; and 130 samples from acute flaccid paralysis (AFP) cases. These represent 50%, 54.8%, and 51.5%, respectively. According to molecular analysis, 90% of EVA71 were attributed to genotype C4 and 10% were attributed to genotype B5. The predominance of EVA71 circulation among the population proves the need to strengthen surveillance (with monitoring of enterovirus circulation for facilitation of HFMD outbreak prediction) and to increase the effectiveness of preventative measures by the implementation of vaccination against EVA71-associated infections. A phase III trial of a Taiwanese vaccine (EV71vac) in Taiwan and South Vietnam showed its safety, tolerability, and efficacy in children aged 2-71 months. This B4 genotype-based vaccine, which features cross-protection against B5 and C4 genotypes, and other existing EV71 vaccines can serve as a good approach to solving the HFMD problem, which is so important for Vietnam.

Aim: Compare frequency of isolation of polioviruses in children living in closed-type facilities (orphanages) before and after the change in poliomyelitis vaccination scheme., Materials and Methods: Feces samples of 207 children from 5 orphanages during immunization with oral poliomyelitis vaccine (OPV) and of 259 children from 4 orphanages during vaccination with inactivated poliomyelitis vaccine (IPV) were studied. Isolation and identification of polioviruses was carried out according to WHO recommendations., Results: In orphanages, where children were immunized with the oral vaccines, 21 polioviruses were isolated. In orphanages, where only inactivated vaccine was used, 10 polioviruses were isolated, the presence of polioviruses in these facilities is associated with their introduction from the outside. The percentage of poliovirus detection in children immunized with OPV was shown to be 16.9 ± 3.4% and was significantly higher than in children vaccinated with IPV (6.1 ± 1.9%). Polioviruses isolated from children immunized with OPV belonged to serotypes 1, 2 and 3 in 19.0, 14.3 and 66.7% of cases, respectively. Polioviruses detected in children immunized with IPV belonged to serotypes 1, 2 and 3 in 30, 40 and 30% of cases, respectively. All the isolated polioviruses turned out to be Sabin vaccine strains., Conclusion: Implementation of strict prophylaxis measures in orphanages is necessary in order to prevent the possibility of introduction, transmission and circulation of polioviruses. Improvement of control in children from closed-type facilities will ensure maintenance of Russian Federation status as the country free of poliomyelitis.

Aim: Study features of epidemic process and etiology of oral cavity and limb enterovirus exanthema group diseases in a number of territories of Northwestern Russia., Materials and Methods: Isolation and identification of non-poliomyelitis enteroviruses from material of patients was carried out according to WHO recommendations. Phenotyping and phylogenetic analysis of enteroviruses was carried out., Results: In 3 territories of Northwestern Russia oral cavity and limb enterovirus group diseases were registered. Children aged less than 14 years, predominately aged less than 3 years, were shown to be involved in the epidemic process. Coxsackie A16 enteroviruses from 27 samples of patients were isolated in cell cultures and identified by using specific sera. Coxsackie A16 enteroviruses from 16 samples were identified by using partial sequencing of VP1 genome area. Phylogenetic analysis has shown that the identified Coxsackie A16 viruses distributed among 2 phylogenetic groups., Conclusion: Coxsackie A16 enteroviruses that had never been detected in the region previously were established to be the etiologic factor of oral cavity and limb enterovirus exanthema group disease in the 3 territories of Northwestern Russia. The data obtained give evidence on the necessity of epidemiologic and virological control for enterovirus infection with the aim of obtaining novel information on the circulation of non-poliomyelitis enteroviruses in the population and the establishment of development patterns for epidemic process of this infection.

Aim: Genotyping and phylogenetic analysis of parvovirus B19 isolates isolated on the territories of Northwestern Federal District (NWFD) of Russia., Materials and Methods: 61 blood sera and 30 oropharyngeal lavages obtained from patients with maculopapular rash from various territories of NWFD were studied for the presence of parvovirus B 19 DNA (PVB 19). DNA isolation and amplification was carried out by standard techniques. DNA segment including fragment of non-structural gene NS1 and region of structural gene VPI (NS1-VPlu, 994 nucleotides) was sequenced, original sequences ofoligonucleotide primers were selected for this purpose. Phylogenetic analysis was carried out online on the website http://www.phylogeny.fr. Data for tree construction was obtained from GenBank., Results: PVB 19 DNAwas detected in 45% ofsamples. PVB19 genome segment was sequenced in 8 samples. All the PVB 19 isolates belong to a single cluster of 1A genotype. Isolate 57.12 from Komi Republic is similar to ISR-G strain isolated from Israel., Conclusion: Phylogenetic analysis showed a high degree of genetic similarity between PVB 19 isolates circulating on the territories of NWFD, their membership in the most widespread genotype in the world. Local and import cases ofparvovirus infection (PVI) were identified. The authors make a conclusion on the necessity to include PVI into the system of rubella and measles control.

Aim: Analysis of results of virological study of material from children of migrants and evaluation of intensity of immunity against polioviruses in these children., Materials and Methods: 1668 feces samples from patients with acute flaccid paralysis and contact individuals and 479 feces samples from healthy children from families of migrants, as well as 1012 blood sera of children aged 3 - 4 and 14 - 15 years living in the same territory of Russia, and 169 blood sera of children of migrants were studied. Polioviruses and non-polio enteroviruses were isolated by standard procedures recommended by WHO in 3 cell cultures - RD, L20B and Hep-2. Virus identification was carried out by microneutralization test with rabbit antisera against poliomyelitisvirus, RIVM (Bilthoven, Netherlands). For intra-type differentiation EIA and PCRwere used. Antibody titers were determined in microneutralization reaction with reference poliovirus vaccines strains in Hep-2 cell culture., Results: The frequency of detection of polioviruses in children of migrants was significantly higher than in patients with acute flaccid paralysis. In a larger percent of cases children of migrants did not have protective antibody titers against polioviruses of all the 3 serotypes., Conclusion: Migrants as a significant source of poliovirus detection may be an indicator group for detection of signs of unfavorable epidemic situation. Based on the results of epidemiologic surveillance of migrants the fact of import of wild poliovirus into North-West of Russia with the absence of poliomyelitis was proven, which confirms an important role of this form of monitoring in the system of poliomyelitis control.

Aim: Analyze the state of population immunity to polioviruses in children population of 14 administrative territories of the Russian Federation., Materials and Methods: Antibody titers against reference vaccine poliovirus serotypes 1, 2 and 3 were determined by using neutralization reaction in cell culture Hep-2., Results: Several years after the termination of conduction of National immunizations days the percent of seronegative children was shown to increase, especially to poliovirus serotype 3, in most of the territories. Possible reasons for fluctuation of geometric mean antibody titers against polioviruses in children in various territories as well as in various years in the same territory were established., Conclusion: Quality vaccine prophylaxis and control of population immunity state among children population play an important role in the maintenance of Russian Federation status as a country is free from poliomyelitis.

Aim: Control of poliovirus circulation by study of material from patients with acute flaccid paralysis and contact individuals, from children of risk groups; molecular characteristics of isolated polioviruses; monitoring of circulation of polioviruses and nonpoliomyelitis enteroviruses in population and the environment., Materials and Methods: Isolation and study of polioviruses and nonpoliomyelitis enteroviruses from various sources was performed in accordance with WHO recommendations., Results: Prolonged persistence and circulation of vaccine related strains of polioviruses in children is demonstrated. Enterovirus serotypes that circulate in the population and the environment more frequently are determined. CONCLUSION. Long term control of poliomyelitis and acute flaccid paralysis in combination with additional control variants in children from risk groups and objects of the environment allowed to obtain valuable data on poliovirus and nonpoliomyelitis enteroviruses circulation for the Program of eradication of poliomyelitis.

The rate of case investigation for measles-like illness (MLI) is an important indicator for the quality of measles surveillance in countries targeting measles elimination. However, a benchmark rate is still being discussed. We assessed different rates of investigation in 11 territories of the Russian Federation with low reported measles incidence during the previous 4-7 years. Each territory maintained their existing surveillance activities and also undertook additional surveillance activities for MLI over a 3-year period. The annual routine rate of investigation varied from 0·06 to 1·8/100,000 population; the overall rate of investigation, including enhanced surveillance, varied from 1·4 to 7·2/100,000. Forty-nine (30·8%) of 159 measles cases detected were identified through enhanced surveillance. Based on the results of this study, the Russian Federation concluded that a rate of routine investigation of 2/100,000 provided the best balance between available resources and sensitivity for detection of measles cases.

Examination of 281 healthy vaccinees from 6 children's homes could isolate 37 vaccine polioviruses. The detection rate for polioviruses in the children's homes, in which there were children born to HIV-positive mothers, was significantly higher than that in other children's homes. These children's homes showed the longest poliovirus excretion after vaccination. The RFLP analysis showed that 40% of the strains being examined were intertypic recombinant strains. The sequence analysis demonstrated that about 30% of the strains under study had a small number of nucleotide substitutions in the VP3-VP1 genomic region. The study revealed a correlation between the dynamics of poliovirus excretion after oral vaccination and the kinetics of formation of recombinant poliovirus strains.

Since the announcement of the WHO program for the global eradication of poliomyelitis and the establishment of epidemiological and virological surveillance, the emergence and circulation of pathogenic vaccine-derived polioviruses (VDPV) presenting >1% nucleotide divergence from the sequence of the original vaccine strain have been demonstrated in certain regions. We developed and used a multiple restriction fragment length polymorphism (RFLP) method to investigate the frequency of these VDPV in a population with a high level of oral poliovirus vaccine coverage in northwestern Russia. Modified RFLP profiles were found to be strongly correlated with the presence of mutations and recombination events in vaccine strains. We found that a substantial proportion of vaccine strains had high percentages of nucleotide substitutions (>0.5%), including a type 3 VDPV with 1.4% nucleotide divergence. These findings indicate that VDPV or pre-VDPV strains are not rare in certain populations with high levels of vaccine coverage. The multiple RFLP method appears to be a simple and rapid tool for monitoring such strains, which could jeopardize the benefits of the eradication program.

Specific antibody immune response to vaccination with commercial inactivated trivalent vaccine A(H1N1) + A(H3N2) + B (IgA, IgG, IgG, subclasses G1, G3, G4, and accumulation of antiCD8) was studied in subjects aged 20-95 years. The initial immune status before vaccination is significant for a positive immune response to the vaccine. Subjects responding to immunization by an increment in specific IgG had a much lower prevaccination level of these antibodies than subjects without these Ig conversion. Antibody immune response to vaccination depended on patient's age. All vaccinees aged 20-25 years developed an increment in IgG to at least one of influenza antigens used. Specific postvaccinal immune response to inactivated influenza vaccine included accumulation of G1, G3, and A antibodies, but not G4 or E antibodies. This latter fact suggests the absence of sensitizing effect of vaccination. In elderly subjects an increment in G1, G3, and A antibodies may not involve an increase in the total level of IgG. In part of elderly subjects secretion of specific antibodies was observed in the presence of increased concentration of antilymphocytic antibodies (antiCD8), indicating a possibility of autoimmune reactions in subjects of this age after injection of inactivated influenza vaccine.

The present investigation indicate that homologous polyclonal anti-idiotypic antibodies induce both humoral and cell-mediated immune response. Experiments on the exhaustion of immune sera with vaccine have revealed that anti-idiotypic antibodies induce not only specific antibodies to influenza virus antigens, but also antibodies to other epitopes of the globulin molecule of the anti-idiotype. Anti-idiotypic antibodies, when reintroduced into animals, induce the production of anti-influenza antibodies of the anamnestic type, but do not induce the formation of antihemagglutinins. The injection of influenza vaccine to animals, previously immunized with anti-idiotypic antibodies, induces the production of antihemagglutinins; an increase in the level of immune complexes and antibodies to anti-idiotype, i.e. anti-idiotypic antibodies, induces the development of immunological memory with respect to influenza virus antigens, including antihemagglutinins.

Antiviral properties were found with a preparation obtained from mussel meat by acid hydrolysis. In the experiments in vitro this mussel hydrolysate reduced the infectious activity of influenza viruses in neutralization tests by 3-6 log. Applications of the preparation to mice infected with a toxigenic influenza virus strain protected from death 66-84% of the animals, the survival rate in controls being from 9 to 16%. The protective effect of mussel hydrolysate was observed both after intranasal and oral administrations. The optimal protective effect was obtained when the preparation was used 5-24 hours before inoculation.

The injection of anti-influenza antibodies into rabbits induces a specific immune response, including the production of anti-idiotypic antibodies and then the production of immune complexes and anti-influenza antibodies. The injection of antibodies induces the development of immunological memory; as a result, the animals are primed to respond to the injection of influenza vaccine.

A safe, moderately reactogenic and immunologically effective scheme of complex (combined) immunization against meningitis A, diphtheria, typhoid fever, viral hepatitis A and influenza has been developed as the result of experimental and clinico-immunological studies. Depending on the epidemiological situation, the newly developed scheme can be used in two variants. According to the first variant of this scheme, the following preparations are injected subcutaneously into three different sites: a mixture of group A meningococcal vaccine and diphtheria toxoid, typhoid vaccine and influenza vaccine. The second variant of the scheme differs from its first variant in using intramuscular injection of normal human immunoglobulin instead of injection of influenza vaccine. Moreover, in practical realization these variants may be altered by excluding vaccines, unnecessary under present conditions. The newly developed scheme of vaccinal prophylaxis is recommended for practical use.

The injection of inactivated and live influenza virus into rabbits induces the formation of anti-idiotypic antibodies, appearing after anti-influenza hemagglutinins, in the blood. The presence of immune complexes antibody--anti-idiotypic antibody in the blood of the animals has been established. The booster immunization of the animals with influenza virus antigens produces a rise in the levels of both idiotypic and anti-idiotypic antibodies. The injection of autologous anti-idiotypic globulin into the primed animals ensures the induction of idiotypic and anti-idiotypic revaccinal reactions.

A comparative study of the antigenic characteristics of influenza A (H1N1) and A (H3N2) virus strains isolated during the same epidemic season, 1989-1990, in St. Petersburg and CzSFR was carried out by routine serological methods and by cross-challenge of immunized mice. Antigenic differences were found only among serosubtype A (H1N1) strains isolated in different areas. The study was carried out in connection with potential occurrence of influenza epidemic outbreaks from local sources.

The side-effect of nonvirion admixtures in inactivated influenza vaccines has been experimentally studied. The minimal sensitizing dose of allantoic admixtures other than ovalbumin has been experimentally determined in vivo. This dose has proved to be 4 times higher than the sensitizing dose of ovalbumin. The presence of sharply defined correlations between the concentration of the admixtures contained in inactivated influenza vaccines, the titers of antibodies to these admixtures and the severity of anaphylactic reactions has been established in guinea pigs after their multiple immunization.

Admixtures of free antigens have been shown to play the main role in the anaphylactogenic danger of vaccines. The immunogenic and anaphylactogenic action of such antigenic admixtures in corpuscular influenza vaccine can be observed after the immunization of animals in 2 or 3 injections. Host antigens incorporated into viral particles induce no anaphylactic reaction in guinea pigs after their immunization in 3 injections.

The immunogenicity of the Czechoslovak-made commercial vaccine Adinvira as well as that of the chromatographically concentrated vaccine Leningrad of Soviet origin was evaluated in an immunization project. The both vaccines were well tolerated after administration, the Soviet-made Leningrad brand featuring greater efficiency in eliciting an antibody response against influenza virus haemagglutinin.

Experiments in laboratory animals demonstrated safety, areactogenicity, and low allergenic potential of inactivated influenza chromatographic vaccine (IICV) administered by aerosol method. Aerosol immunization of white rats resulted in the development of the same level of secretory antibody as after intraperitoneal inoculation of IICV but the immunizing aspirational dose was 10-20 times as low as parenteral. Marked protection of mice was achieved after triple aerosol immunization of immunologically experienced animals.

Rooster monospecific sera to influenza virus hemagglutinin free from antibodies to the host (chick embryo) admixture components were prepared and satisfactorily used in radial immunodiffusion test for control of the levels of hemagglutinin in influenza vaccines. The indirect hemagglutination test with erythrocytic antibody diagnosticum has been proposed for the detection of low ovalbumin concentrations. The sensitivity of this method is less than 0.003 micrograms/ml ovalbumin which significantly surpasses other currently accepted methods for control of this component in inactivated influenza vaccines. Immunochemical methods established the presence in influenza vaccines of other "nonovalbumin" multicomponent admixtures. Their sensitizing activity was characterized and found to be 4-5 times weaker than that of ovalbumin.

Reactogenic and immunogenic properties of killed influenza vaccine prepared of various strains of influenza virus by the method of sorption chromatography on sliciate sorbents were studied on small groups of volunteers; there proved to be a moderate reactogenic and a marked immunogenic activity of the experimental batches of the preparation. The dose, the concentration and the method of administration of the vaccine to obtain the immune response were determined. The indices of humoral and local immunity following a single vaccination were studied. On the basis of parenteral immunization of donors with killed influenza vaccine there were obtained experimental samples of the antinfluenza gamma-globulin with an increased specific activity.

The safety, reactogenic properties and antigenic potency of inactivated whole-virion influenza vaccines produced in the USSR were studied on 1,117 schoolchildren in limited coded clinico-immunological observations. Inactivated chromatographic influenza vaccine obtained from strain A/Texas/1/77 (H3N2) and a variant of this vaccine, developed specially for children and obtained from strains A/Texas/1/77 and B/Leningrad/76, were used for immunization. Both preparations were introduced intradermally in a single injection in a volume of 0.1-0.2 ml by means of bi-3 jet injector. The content of hemagglutinin in this volume varied from 3.0 to 8.0 micrograms. Clinico-laboratory investigations demonstrated the safety of mono- and bivalent inactivated whole-virion influenza vaccine administered intradermally in a single injection to children of school age. The vaccines showed low reactogenicity and high antigenic potency in children aged 7-10 and 11-14 years, and the optimal doses of the preparations were selected for children of different age groups. The distinct prophylactic effectiveness of inactivated chemical chromatographic influenza vaccines in children aged 11-14 years was revealed 11 months after immunization, the index of immunological effectiveness being 1.7.

Corpuscular influenza vaccines have been found to contain chick embryo antigens differing from ovalbumin in their antigenic, immunogenic, and allergenic properties. The study has revealed that host antigens are present in vaccines in the form of free contaminants and as substances incorporated into viruses. The vaccine prepared by the method of sorption-elution on red blood cells with subsequent gel chromatography is characterized by the highest antigenic purity.

Two immunoaffinity chromatographic methods for the purification of corpuscular influenza vaccine from the admixture of chick embryo components have been examined. The isolation of the virus on immobilized antiviral antibodies has proved to be unsuitable for preparative purposes. The method for the purification of the vaccine from ovalbumin with the use of immobilized anti-ovalbumin antibodies has proved to be highly effective. When introduced into guinea pigs in 3 injections, the vaccine purified by immunosorption has been found to produce no anaphylactic reactions.