Laboratory Lead — Mikhail P. Grudinin, PhD
Phone: +7 (812) 234–42–51
E-mail: grudinin@influenza.spb.ru
The Laboratory of Molecular Virology and Genetic Engineering, formed in 1987, is dedicated to the research of genetic variety, molecular biologic properties and evolution of influenza A, B and other viruses.
n the course of 2006–2009 epidemic seasons, more than 200 influenza A and B strains have been analyzed. 110 A/H1N1 strains were studied for Oseltamivir resistance. It was established that during 2008–2009 epidemic season in Russia, influenza A H1N1(clade 2B and 2C), pandemic A/H1N1v, H3N2 and influenza B viruses were in circulation. Among A/H1N1 strains, clade 2B prevailed. A/H3N2 strains were of D genotype, showed resistance to Rimantadine and sensitivity to Oseltamivir. No Perth/16-like strains, recommended for vaccine composition, were isolated among Russian A/H3N2 population. 2009 saw the return of Vic87-like influenza B strains (III–II), which replaced Yam88-like strains that were dominant in 2008. All studied pandemic á/H1N1v strains were resistant to Rimantadine (S31N mutation) and sensitive to Oseltamivir.
It was proven that certain influenza proteins (HA, NA and M) are capable of causing pathologic abnormalities similar to that of ILI infections both in vitro and in vivo. HA, NA and M proteins are also capable of modulating fibrinolytic and anticoagulant activity of blood plasma, changing activity of erythrocytes and thrombocytes , amplifying contraction frequency of lymphatic vessels, blocking T-Cells’ CD-4 receptor(HA by 20% and HM by 70%), causing proliferation of peripheral blood leucocytes and CD-25 receptor expression. This can be explained by the presence in viral protein structure of amino acid sequences that can mimicry the hosts’ protein and peptide amino acid sequences. A new study concerning the damaging effect of influenza virus on the vessel endothelium has started. The data acquired through this study shows that influenza A viruses (H5N1, H3N2, H1N1) can fully reproduce in endothelial cell culture and inflict damage, causing apoptosis.
The Laboratory developed test samples of oligonucleotid chip for detecting influenza subtype A/H5N1 in clinical conditions. With the help of reverse genetics, 6 vaccine candidates were acquired, which are 5:3 and 6:2 reassortants of high-yield PR/8/34 virus, influenza A/Kurgan/5/05 (H5N1) and A/Texas/04/09 (H1N1) viruses.
Determination of primary nucleotide sequences of full-size genomes of H5N1 isolates from Kurgan city and Crimea region, collected during avian influenza outbreak in 2005, was conducted. Phylogenetic analysis of primary nucleotide sequences showed the homology of northwest Siberian strain hemagglutinin with strains isolated the same year in Qinghai province, China. These isolates cluster with strains of so called «Qinghai Group» and belong to A/H5N1 clade 2 subclade 2 (WHO) and are close to WHO-recommended An A/Bar headed goose/Quinghai/1A/2005-like virus candidate, which was acquired through reverse genetics.
Novel techniques for regulating self-assembly of polypeptides were introduced.
Influenza vectors expressing Mycobacterium tuberculosis ESAT-6 protein were created. Prophylactic and therapeutic efficacy of recombinant influenza vectors on experimental tuberculosis infection was shown.
A pioneering biological system for expressing recombinant delta antigen gene was developed. The system simultaneously synthesizes small and large HDV antigens by eukaryotic expression type, based on E. coli XL1-Blue strain, carrying Sup E mutation, ÒUC-D and pGEX-D plasmids.
Lab scientists amplified, sequenced and cloned genes, which encode structural part of 14.3.3 proteins and prions, and a full-size HDV antigen gene (serum of infected human).
Genotype II HDV, previously isolated only in Japan and Taiwan, was isolated in Yakutia. Phylogenetic analysis of the isolate with the help of ClustalW, PHILIP and PAUP showed that the Russian isolate forms its own branch between Japanese and Taiwanese isolates. Phylogenetic analysis of full-size HBV genome sequence, acquired from GenBank/EMBL, showed high recombinant frequency in HDV evolutional history. For 9 mosaic genomes probable recombination sites were mapped. 6 mosaic genomes isolated in southeast Asia formed as a result of B and C genotype recombination. 3 mosaic genomes are A/D recombinants. They were isolated in Italy, where A and D genotypes constitute 90% of HBV population. In Saint-Petersburg, Russia, an A/D recombinant genome was isolated. 3 specimen from southeast Asia were identified as C recombinants. Studies showed that due to human migration HBV genotype spectrum, found in Saint-Petersburg and Western Europe, is expanding.
