[Monoclonal antibodies to hemagglutinin of influenza A/H7N3 virus (Orthomyxoviridae: Alphainfluenzavirus: Influenza A virus)].

INTRODUCTION: Variants of influenza virus A/H7 have the same high pandemic potential as A/H5. However, the information about the antigenic structure of H7 hemagglutinin (НА) is considerably inferior in quantitative terms to similar data for H5 НА.The aims of the study were development and characterization of the monoclonal antibodies (MAbs) panel for HA subtype H7 of the influenza A virus. MATERIAL AND METHODS: Viruses were accumulated in 10-day-old chicken embryos. Purification and concentration of the virus, determination of protein concentration, preparation of MAbs and ascitic fluids, hemagglutination and hemagglutination inhibition (HI) tests, assessment of antibodies' activity in indirect enzyme-linked immunosorbent assay (ELISA), as well as determination of MAbs isotypes and neutralization reaction (NR) were carried out by standard methods. RESULTS: The obtained MAbs to А/mallard/Netherlands/12/2000 (H7N3) strain were studied in HI test with a set of strains of different years of isolation belonging to different evolutionary groups. MAbs had a reduced reactivity compared to the immunogen-virus for all the studied strains. Cross-interaction of MAbs 9E11 and 9G12 in HI test with influenza A/H15 virus has been observed. DISCUSSION: Influenza A agent with H7 HA variant could serve as a potential cause of a future pandemic. Development of the MAbs panel for subtype H7 HA is an urgent task for both veterinary medicine and public health. CONCLUSION: The obtained MAbs can be used not only for epitope mapping of the H7 HA molecule (currently insufficiently studied) and as reagents for diagnostic assays, but also for determining common («universal¼) epitopes in HA of different strains of this subtype.

[Change of phenotypic properties of escape mutants and readaptants of influenza virus A (H1N1)pdm09 under the influence of selected mutations in the molecule of hemagglutinin.]

INTRODUCTION: After the emergence and spread of pandemic H1N1 viruses in 2009, antigenic epitopes recognized by neutralizing antibodies against the hemagglutinin of influenza A/Moscow/01/09(H1N1)pdm09 viruses were studied. PURPOSE: The purpose of the study was to obtain readapted variants of the virus from a low-virulent escapemutant that has an increased affinity of the avian and the human types cellular receptors compared to the wild type and the comparative study of their antigenic and receptor specificity. MATERIAL AND METHODS: Viruses were accumulated in 10-day-old chicken embryos. The MAB panel against HA of influenza virus strain A/IIV-Moscow/01/09(H1N1)sw1 was used in the form of ascites fluids from mice. Immunization of mice, HI testing, elution of viruses from chicken erythrocytes, PCR and sequencing of readapted variants were performed by standard methods. RESULTS: The amino acid substitution A198E acquired in the process of readaptation leads to changes in the antigenic specificity. A correlation was found between a decrease in virulence of a low-virulent escape mutant associated with the substitution D190N in the hemagglutinin molecule and an increase in the hemagglutinating titer to inhibitors in normal mouse serum. Viruses with low affinity of cellular receptor analogs and carrying amino acid substitutions have an increased ability to elute from chicken erythrocytes. DISCUSSION: The results discuss the effect of mutations in the HA molecule of the influenza A(H1N1) pdm09 virus to the change in antigen specificity; virulence for mice, adsorption-elution at cellular receptors. CONCLUSION: A comparative study of the antigenic specificity and receptor-binding activity of the escape mutants was conducted for the hemagglutinin of the influenza virus A/Moscow/01/2009 (H1N1)swl, and the readapted variants obtained for one of the escape mutants with reduced virulence for mouse. Monitoring the pleiotropic effect of mutations in the hemagglutinin H1 molecule is necessary to predict variants of the virus with pandemic potential.

[Adaptation of hepatitis A virus to nonprimate cells]. / Adaptatsiia virusa gepatita A k neprimatnym kletkam.

Hepatitis A virus (HAV) was adapted to nonprimate BHK-21 cell line (Syrian hamster kidney). Enzyme immunoassay, immunoblotting, and slot hybridization demonstrated the capacity of HM175 culture strain to stable reproduction in this cell line. More than 50 passages of adapted HAV were carried out, which showed no changes in the basic cultural characteristics of the virus. The data permit a conclusion on the possibility of HAV reproduction in nonprimate cells.

[The reproduction of the hepatitis A virus in cell cultures]. / Reproduktsiia virusa gepatita A v kletochnykh kul'turakh.

A comparative analysis of reproduction of 9 strains of hepatitis A viruses (HSA-15, CF-979, MI, MBB 11/5, H-141, KMW-1, GBM, IH-26, WR-61) from different regions of the world in cell cultures (PLC/PRF/5, HEL-240, FRhK-4, MK) revealed the differences in the capacity of the viruses for reproduction in these cell lines. The factors influencing HAV reproduction in cell cultures such as temperature, medium, and sera were studied. In one-cycle infection, accumulation of vRNA and formation of virus particles were analysed.

[The adaptation of an isolate of the hepatitis A virus to reproduction in cell cultures]. / Adaptatsiia izoliata virusa gepatitia A k reproduktsii v kletochnykh kul'turakh.

A HAV strain derived from a patient in Moscow multiplied in a continuous cell line PLC/PRF/5 at 32 degrees C (variant MI-1) and at 37 degrees C (variant MI-1.1). These two variants of the strain MI retained the ability for reproduction both at 32 degrees C and 37 degrees C after passages in cell culture. The strain MI also replicates in MK cells. Negative results on HAV cultivation were obtained in HEF-240 and FRhK-4 cell cultures. The MI-1 and MI-1.1 variants are typical of hepatitis A viruses by their physicochemical properties, the results of ELISA, protein electrophoresis and dot hybridization tests.

[Cultivation of hepatitis A virus in various cell lines and isolation of DNA sequences]. / Kul'tivirovanie virusa gepatita A v razlichnykh kletochnykh liniiakh i poluchenie DNK-posledovatel'nostei.

Using the MBB/11/5 strain originally adapted to the PLC/PRF/5 cell line, reproduction of the virus in diploid cells of human embryo fibroblasts, continuous primate lines (RAMT, FRhK-4) and human urinary bladder tumor (T-24) lines was studied. We obtained HAV DNA sequences (about 99% from vRNA) for the MBB/11/5 strain which were used as a probe for demonstration of vRNA synthesis in the infected cells.