Investigations of the antigenic structure and genome of influenza A (H1N1) viruses isolated in the Mongolian People's Republic in 1982, 1983, 1986 and 1987 from children with acute respiratory diseases using monoclonal antibodies and nucleotide sequencing revealed 4 strains identical to the prototype strain A/PR/8/34 (H1N1), variant M. Sinai, and 8 strains closely similar to the epidemic strain A/USSR/90/77 (H1N1) in the antigenic structure and A/Leningrad/54 (H1N1) in the primary structure of HA.

A new technique of DNA sequencing by hybridization with oligonucleotide matrix (SHOM) which could also be applied for DNA mapping and fingerprinting, mutant diagnostics, etc., has been tested in model experiments. A dot matrix was prepared which contained 9 overlapping octanucleotides (8-mers) complementary to a common 17-mer. Each of the 8-mers was immobilized as individual dot in thin layer of polyacrylamide gel fixed on a glass plate. The matrix was hybridized with the 32P-labeled 17-mer and three other 17-mers differing from the first one by a single base change. The hybridization enabled us to distinguish perfect duplexes from those containing mismatches in 32 out of 35 cases. These results are discussed with respect to the applicability of the approach for sequencing. It was shown that hybridization of DNA with an immobilized 8-mer in the presence of a labeled 5-mer led to the formation of a stable duplex with the 5-mer only if the 5- and the 8-mers were in continuous stacking making a perfect nicked duplex 13 (5+8) base pairs long. These experiments and computer simulations suggest that continuous stacking hybridization may increase the efficiency of sequencing so that random or natural coding DNA fragments about 1000 bases long could be sequenced in more than 97% of cases. Miniaturized matrices or sequencing chips were designed, where oligonucleotides were immobilized within 100 x 100 micron dots disposed at 100 micron intervals. Hybridization of fluorescently labeled DNA fragments with microchips may simplify sequencing and ensure sensitivity of at least 10 attomoles per dot. The perspectives and limitations of SHOM are discussed.

In an attempt to develop a reliable system for DNA sequence analysis with multiple hybridization probes, oligonucleotides down to 8 bases long were covalently immobilized in a thin layer of polyacrylamide gel fixed on a glass plate. It was shown possible to detect single base changes in DNA by hybridization of the immobilized oligonucleotides with radioactively and fluorescently labeled DNA fragments. Moreover, it was found that dissociation temperatures of differently GC-rich duplexes could be equalized by appropriate choice of immobilized oligonucleotides concentrations. A model accounting for this phenomenon is presented. In order to make the system more compact, a rectangular matrix of 200 mm dots of immobilized oligonucleotides ("hybridization chip") was designed which offered the sensitivity of 20 attomoles per dot for fluorescent DNA fragment. The applications and perspectives of the approach are discussed.

Investigations of the antigenic structure and genome of influenza A (H1N1) viruses isolated in the Mongolian People's Republic in 1982, 1983, 1986 and 1987 from children with acute respiratory diseases using monoclonal antibodies and nucleotide sequencing revealed 4 strains identical to the prototype strain A/PR/8/34 (H1N1), variant M. Sinai, and 8 strains closely similar to the epidemic strain A/USSR/90/77 (H1N1) in the antigenic structure and A/Leningrad/54 (H1N1) in the primary structure of HA.

A molecular analysis was made of genomes of influenza A (H1N1) virus strains, the causative agents of an epidemic in Leningrad, 1986. The primary structure of hemagglutinin gene of two of these strains, A/Leningrad/624/86 and A/Leningrad/621/86, was established, as well as partial primary structure of PB1 gene of certain current strains of the A (H1N1) subtype. A hypothesis of a "shift" of PB1 gene in 1950-1957 is suggested.

The hemagglutinin gene primary structure of influenza virus A/Riga/9977/86 (H3N2) belonging to the "Coen/84" antigenic subgroup was determined by primer sequencing. A comparative analysis confirmed that the reversions of amino acids in the late stages of the H3 influenza virus subtype antigenic drift became more frequent and the antigenic variants remained in epidemic circulation longer. The possible role of some mutations is discussed.

The authors own results on the variety of the genomic primary structures in human influenza A viruses participating in the epidemic process, including the atypical viruses. The comparative studies revealed new trends in the HA gene antigenic drift on the late stages and the PB1 gene shift. Modifications occurring in the primary structure of the influenza A viruses native genomes during laboratory treatment (adaptation to new hosts, vaccine preparation, egg passaging) have been analyzed. Sequencing of several types of "antigenic anachronisms" revealed the direct links between some of such viruses and the anthropogenic pollution of the biosphere by vaccine strains. Modifications in the HA genes of influenza A viruses during the persistent infection have also been studied.

Electrophoresis in polyacrylamide gel of the reference influenza A/Victoria/35/72 (H3N2) virus and its persisting variants (PV) showed that the PV isolated on the 158th day from the moment of persistence modelling (PV158) had mutation in the gene of hemagglutinin (HA). This mutation is manifested by incomplete HA synthesis at 40 degrees C and increase of mobility of the light HA subunit (HA2). Analysis of nucleotide sequence of the greater part of HA gene of PV158 virus revealed 5 nucleotide substitutions four of which were significant. Three substitutions were found in Hal: 219 (Ser----Phe), 220 (Arg----Gly), 226 (Leu----Gln) and one in HA2: 156 (Thr----Asn). The importance of these mutations in the determination of the PV phenotype is discussed.

A molecular analysis was made of genomes of influenza A (H1N1) virus strains, the causative agents of an epidemic in Leningrad, 1986. The primary structure of hemagglutinin gene of two of these strains, A/Leningrad/624/86 and A/Leningrad/621/86, was established, as well as partial primary structure of PB1 gene of certain current strains of the A (H1N1) subtype. A hypothesis of a "shift" of PB1 gene in 1950-1957 is suggested.

The nucleotide sequence of the cDNA for the viral RNA region coding for the main antigenic protein of the epidemic stomatitis virus of Asia 1 serotype has been identified. The amino acid sequences in the regions of VP1 protein antigenic determinants of the serotype Asia 1 virus and other serotypes viruses have been compared.

The primary structure of hemagglutinin of the members of three main antigenic groups of current influenza A (H3N2) viruses was determined. A phenomenon of "antigenic mimicry" was found to be due to the accumulation of reversions in hemagglutinin structure at late stages of antigenic drift.