[Show abstract][Hide abstract] ABSTRACT: The variation of highly pathogenic avian influenza H5N1 virus results in gradually increased virulence in poultry, and human cases continue to accumulate. The neuraminidase (NA) stalk region of influenza virus varies considerably and may associate with its virulence. The NA stalk region of all N1 subtype influenza A viruses can be divided into six different stalk-motifs, H5N1/2004-like (NA-wt), WSN-like, H5N1/97-like, PR/8-like, H7N1/99-like and H5N1/96-like. The NA-wt is a special NA stalk-motif which was first observed in H5N1 influenza virus in 2000, with a 20-amino acid deletion in the 49(th) to 68(th) positions of the stalk region. Here we show that there is a gradual increase of the special NA stalk-motif in H5N1 isolates from 2000 to 2007, and notably, the special stalk-motif is observed in all 173 H5N1 human isolates from 2004 to 2007. The recombinant H5N1 virus with the special stalk-motif possesses the highest virulence and pathogenicity in chicken and mice, while the recombinant viruses with the other stalk-motifs display attenuated phenotype. This indicates that the special stalk-motif has contributed to the high virulence and pathogenicity of H5N1 isolates since 2000. The gradually increasing emergence of the special NA stalk-motif in H5N1 isolates, especially in human isolates, deserves attention by all.
[Show abstract][Hide abstract] ABSTRACT: The swine influenza virus (SIV) strain A/Swine/TianJin/01/2004(H1N1) (A/S/TJ/04) was rescued successfully by an eight-plasmid rescue system. The cDNAs of SIV 8 gene segments were synthesized by RT-PCR and cloned into the RNA polymerase I/II bidirection expression vector PHW2000 independently, resulting in 8 recombinant plasmids. The 8 recombinant plasmids were cotransfected into COS-1 cell, 30 h later TPCK-trypsin was added to 0.5 microg/mL. The COS-1 cell and supernatant were harvested 48 h after cotransfection and were inoculated into the allantoic cavity of 9-day-old specific-pathogen free (SPF) chicken eggs. The allantoic fluid of dead eggs was harvested and passaged 3 generations in SPF chicken eggs to get infective virus. The successful rescue of A/S/TJ/04 SIV was identified by hemagglutination assay, hemagglutination inhibition assay, sequence analysis and electron microscope observation. The successful rescue of SIV built a platform for the research of the relationship between genome structure and function of SIV, the mechanisms of trans-species transmission of influenza virus and for the generation of new SIV as vaccine.
No preview · Article · Jun 2008 · Sheng wu gong cheng xue bao = Chinese journal of biotechnology
[Show abstract][Hide abstract] ABSTRACT: RNA interference (RNAi) is a powerful tool to silence gene expression. Small interfering RNA (siRNA)-induced RNA degradation has been recently used as an antivirus agent to inhibit specific virus replication. Here, we showed that several siRNAs specific for conserved regions of influenza virus matrix (M2) and nucleocapsid protein (NP) genes could effectively inhibit expression of the corresponding viral protein. We also evaluated the antiviral potential of these siRNAs targeting M2 and NP of H5N1 avian influenza virus (AIV), which are essential to viral replication. We investigated the inhibitory effect of M2-specific siRNAs and NP-specific siRNAs on influenza A virus (H5N1, H1N1 and H9N2) replication in Madin-Darby canine kidney (MDCK) cells and BALB/c mice. The results showed that treatment with these siRNAs could specifically inhibit influenza A virus replication in MDCK cells (0.51-1.63 TCID(50) reduction in virus titers), and delivery of pS-M48 and pS-NP1383 significantly reduced lung virus titers in the infected mice (16-50-fold reduction in lung virus titers) and partially protected the mice from lethal influenza virus challenge (a survival rate of 4/8 for H1N1 virus-infected mice and 2/8 for H5N1 virus infected mice). Moreover, the treatment of pS-M48 and pS-NP1383 could suppress replication of different subtypes of influenza A viruses, including a H5N1 highly pathogenic avian isolate strain. The results provided a basis for further development of siRNA for prophylaxis and therapy of influenza virus infection in humans and animals.
Full-text · Article · Dec 2007 · Antiviral Research