OBJECTIVEIn order to investigate the relationship between selection pressure and the prevalence of antigenic clusters, we sequenced and analyzed the H3N2 influenza virus from China between 1992 and 2012.

METHODSThe H3N2 influenza virus (n = 1206) in China from 1992 to 2012 was analyzed, include global selection pressure and sites positive selection pressure analysis.

RESULTSConsidering all the H3N2 influenza viruses during these 21 years, a total of four amino acid sites subject to positive selection. The global selection pressure varies with the variation of different antigenic clusters and three years with peak bottom selection pressure were identified.

CONCLUSIONThe global selection pressure rise from the peak bottom, a new antigenic clusters will appear andprevalent in the population, indicating the best time to replace the vaccine strain.

Antigens, Viral ; immunology ; China ; Influenza A Virus, H3N2 Subtype ; genetics ; immunology ; Influenza Vaccines ; Selection, Genetic ; Time Factors

OBJECTIVETo evaluate the safety and immunological effect of domestic split influenza virus vaccine.

METHODSAll 606 subjects were divided into three groups by under 6, 16-60 and above 60 years old. Each age group was divided as study group (n = 213), control group 1 (n = 195) and control group 2 (n= 198) by Table of Random Number, one domestic vaccine and two imported vaccines were respectively inoculated in three group people. The differences of clinical side effect rate, antibody positive rate, protective rate and geometric mean titer (GMT) of these three vaccines were compared by using the statistical software with statistical significance of P < 0.05.

RESULTSThe side effect rate of study group, control group 1 and control group 2 was 3.76% (8/213), 4.10% (8/195), and 3.54% (7/198), respectively without statistical significance(chi2 = 0.87, P =0.93). The positive seroconversion rates of H1N1, H3N2 and B in these three groups were respectively 89.2% (190/213), 63.4% (135/213), 86.4% (184/213), 88.7% (173/195), 61.5% (120/195), 87.2% (170/195), 87.9% (174/198), 61.6% (122/198) and 84.8% (168/198). There were no statistical significance in the total positive seroconversion rate of each antibody type (chi2(H1N1) = 0.94, P(H1N1) = 0.63; chi2(H3N2) = 0.94, P(H3N2) = 0.63; chi2(B) = 0.75, P(B) = 0.69). The average growth multiple of H1N1, H3N2 and B in these three groups were 10.7, 7.3, 8.4, 10.5, 6.3, 8.3, 10.2, 7.1, 8.8 times. There were no statistical significances in the GMT growth multiple of each antibody type (F(H1N1) = 0.35, P(H1N1) = 0.70; F(H3N2) = 2.22, P(H3N2) = 0.11; F(B) = 1.51, P(B) = 0.35). The antibody protective rates of H1N1, H3N2 and B were 100% (213/213), 70.0% (149/213), 95.3% (203/213), 100% (195/195), 66.7% (130/195), 97.9% (191/195), 99.5% (197/198), 66.2% (131/198), 96.5% (191/198) respectively. There was no statistical difference among the three vaccines (chi2(H1N1) = 2.04, P(H1N1) = 0.36; chi2(H3N2) = 0.74, P(H3N2) = 0.69; chi2(B) = 0.42, P(B) = 0.82).

CONCLUSIONThe domestic influenza split vaccine might be suitable for colony vaccination for its having clinical safety and immunological effect.

Adolescent ; Adult ; Child ; Humans ; Influenza A Virus, H1N1 Subtype ; immunology ; Influenza A Virus, H3N2 Subtype ; immunology ; Influenza Vaccines ; adverse effects ; immunology ; Influenza, Human ; prevention & control ; Middle Aged ; Young Adult

Adult ; Antibodies, Viral ; blood ; immunology ; Cross Reactions ; Hemagglutinin Glycoproteins, Influenza Virus ; immunology ; Humans ; Influenza A Virus, H1N1 Subtype ; immunology ; Influenza A Virus, H3N2 Subtype ; immunology ; Influenza B virus ; immunology ; Influenza Vaccines ; immunology ; Orthomyxoviridae ; immunology ; Seasons ; Vaccination

OBJECTIVETo screen the influenza A (H3N2) mimotopes by using phage display library.

METHODSUsing influenza A (H3N2) monoclonal antibody as selective molecule, a 7 mer phage peptide library was biopanned and positive clones were selected by ELISA, competition assay and DNA sequencing.

RESULTS21 positive clones were chosen for DNA sequencing. From the experiment and sequencing comparison results, one epitope was comfirmed as mimotope of influenza A (H3N2).

CONCLUSIONInfluenza A (H3N2) mimotope was obtained by phage peptide library screening. The result provide a new approach for new Influenza virals vaccine development.

Epitope Mapping ; Humans ; Influenza A Virus, H3N2 Subtype ; chemistry ; genetics ; immunology ; Peptide Library

Abattoirs ; manpower ; Adult ; Antibodies, Viral ; blood ; China ; Humans ; Influenza A Virus, H1N1 Subtype ; immunology ; Influenza A Virus, H3N2 Subtype ; immunology ; Influenza A Virus, H3N8 Subtype ; immunology ; Influenza A Virus, H5N1 Subtype ; immunology ; Influenza A Virus, H9N2 Subtype ; immunology ; Influenza A virus ; classification ; immunology ; Influenza, Human ; epidemiology ; prevention & control ; virology ; Middle Aged ; Population Surveillance ; methods ; Seroepidemiologic Studies ; Young Adult

OBJECTIVETo analyse seasonal influenza epidemic situation in 2006, and to analyse the genetic and antigenic characteristics of viral hemagglutinin (HA) gene.

METHODSThe single-way hemagglutination inhibition (HI) tests were used to test the antigenic characteristics of these viruses from influenza surveillance network, and the HA1 genes were sequenced based on the antigenic test results according to different isolation times and sites.

RESULTSThe influenza virus types A and B co-circulated in 2006. influenza A H1N1 subtype and Victoria-like B influenza circulated preponderantly during this epidemic season. The HA1 gene sequence of H1N1 viruses showed that 192, 193, 196, 198 positions (located at antigenic site B) have an amino acid substitute, compared with the last circulating strain A/Hubeihongshan/53/2005(H1N1). Two amino acid changes at 142 and 144 positions compared with A/Yunnan/1145/2005 (H3N2). There was no change in influenza B viruses either Victoria-like B or Yamagata-like B virus, i.e . antigenic characteristics is analogous to B/shenzhen/155/2005 and B/tianjin/144/2005, respectively.

CONCLUSIONThe H1N1 and H3N2 influenza viruses had changing antigenic and genetic characteristics in 2006. Influenza virus types B did not change in 2006.

Amino Acids ; analysis ; China ; Hemagglutination Inhibition Tests ; Hemagglutinin Glycoproteins, Influenza Virus ; chemistry ; genetics ; immunology ; Influenza A Virus, H1N1 Subtype ; immunology ; isolation & purification ; Influenza A Virus, H3N2 Subtype ; immunology ; isolation & purification ; Influenza B virus ; immunology ; isolation & purification ; Time Factors

OBJECTIVETo elevate the immunological effect of subunit influenza vaccine in infants and aged people (over 60) using liposomal adjuvant in the context of its relatively low immunity and to investigate the relation between vaccine antigens and liposomal characteristics.

METHODSSeveral formulations of liposomal subunit influenza vaccine were prepared. Their relevant characteristics were investigated to optimize the preparation method. Antisera obtained from immunizinged mice were used to evaluate the antibody titers of various samples by HI and ELISA.

RESULTSLiposomal trivalent influenza vaccine prepared by film evaporation in combinedation with freeze-drying significantly increased its immunological effect in SPF Balb/c mice. Liposomal vaccine stimulated the antibody titer of H3N2, H1N1, and B much stronger than conventional influenza vaccine. As a result, liposomal vaccine (mean size: 4.5-5.5 microm, entrapment efficiency: 30%-40%) significantly increased the immunological effect of subunit influenza vaccine.

CONCLUSIONThe immune effect of liposomal vaccine depends on different antigens, and enhanced immunity is not positively correlated with the mean size of liposome or its entrapped efficiency.

Animals ; Influenza A Virus, H1N1 Subtype ; immunology ; Influenza A Virus, H3N2 Subtype ; immunology ; Influenza B virus ; immunology ; Influenza Vaccines ; administration & dosage ; immunology ; Liposomes ; Mice ; Mice, Inbred BALB C ; Orthomyxoviridae Infections ; prevention & control ; Specific Pathogen-Free Organisms ; Vaccines, Subunit ; administration & dosage ; immunology

Influenza A virus is an enveloped virus that belongs to the Orthomyxoviridae family. It has 8 negative RNA segments that encode 16 viral proteins. The viral polymerase consists of 3 proteins (PB 1, PB2 and PA) which plays an important role in the transcription and replication of the influenza A virus. Polymerase basic protein 1 (PB 1) is a critical member of viral polymerase complex. In order to further study the function of PB1, we need to prepare the PB1 antibody with good quality. Therefore, we amplified PB1 conserved region (nt1648-2265) by PCR and cloned it into pET-30a vector, and transformed into Escherichia coli BL2 1. The expression of His tagged PB 1 protein was induced by IPTG, and His-PB 1 proteins were purified by Ni-NTA resin. For preparation of PB 1 protein antiserum, rabbits were immunized with His-PB 1 fusion protein 3 times. Then the titer of PB 1 polyclonal antibody was measured by indirect ELISA. The antibody was purified by membrane affinity purification and subjected to immunoblotting analysis. Data showed that PB1 antibody can recognize PB 1 protein from WSN virus infected or pCMV FLAG-PB 1 transfected cells. Meanwhile, PB 1 antibody can also recognize specifically other subtype strains of influenza A virus such as H9N2 and H3N2. PB 1 polyclonal antibody we generated will be a useful tool to study the biological function of PB1.
Animals ; Antibodies, Viral ; biosynthesis ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; metabolism ; Genetic Vectors ; Influenza A Virus, H3N2 Subtype ; Influenza A Virus, H9N2 Subtype ; Plasmids ; Rabbits ; Viral Proteins ; immunology

To construct a recombinant vaccinia virus RVJ1175M2 expressing influenza A3 virus (H3N2) M2 gene, full length gene encoding influenza virus (H3N2) M2 protein was amplified with PCR and cloned into plasmid pJSC1175 which was used for homologous recombination with vaccinia virus Tiantan strain. Along with this, a recombinant vaccinia virus RVJ1175M2 containing the M2 gene was subsequently constructed. It was identified by PCR that the gene of M2 protein was inserted into the TK locus of vaccinia virus Tiantan strain correctly and M2 protein was expressed by recombinant vaccinia virus RVJ1175M2 effectively. Two electrophoretic bands of M2 protein expressed by the infected HeLa cells, one of 15kD and the other of 13kD in accordance with related documents, was deteced by Western-blot. M2 protein distributing on the surface of the infected cells was demonstrated by immunofluorescence and flow cytometry. The results suggested that recombinant vaccinia virus RVJ1175M2 could express M2 protein effectively, this laid a foundation for comparative research on the immune effect of universal vaccine of influenza virus with different kinds of vaccine expressing M2 protein.
HeLa Cells ; Humans ; Influenza A Virus, H3N2 Subtype ; genetics ; Influenza Vaccines ; immunology ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis ; Vaccines, Synthetic ; immunology ; Vaccinia virus ; genetics ; Viral Matrix Proteins ; genetics

The molecular characterization and phylogenetic analysis of hemagglutinin (HA) genes of human influenza H3N2 viruses in Guangdong, China from 2007 to 2010 were studied in this study. By space-time sampling of strains, the HA genes of H3N2 strains from Guangdong were sequenced and searched from Internet, and then the variation and evolution of HA genes were conducted by Lasergene 7.1 and Mega 5.05 and evolutionary rates were analyzed by epidemiological data. The phylogenetic tree was established by alignment of 17 Guangdong strains and 26 global reference strains. Ks rates and Ka rates of HA genes were 2.06 x 10(-3)-2.23 x 10(-3) Nt/Year and 1.05 x 10(-3)-1.21 x 10(3) Nt/Year during 2007-2010, while the velocity of HA1 evolution of Ka was 3. 13 times than that of HA2 evolution. Compared with HA of vaccine strain A/Perth/16/2009, the genetic homologies of Guangdong strains in 2009 reached to 98.8%-99.7% and of Guangdong strains in 2010 reached to 98.0%-98.4%. There were some amino acid substitutions in five epitope regions of HA1 during 2007-2010, especially in B region (N160K) and D region (K174R/N); the K189E/N/Q and T228A in RBS (receptor-binding site) occurred in 2010 as two glycoproteins sites substituted impacted on the HA1 antigenicity. The antigenicity of epidemic H3N2 strains in 2010 was to some degree different that of the vaccine strain A/ Perth/16/2009. According to that there were variations of B and D epitopes and two sites of RBS and two glycoprotein in Guangdong H3N2 HA1 genes, WHO/ CDC should recommend new representative strains during 2011-2012 influenza seasons if H3N2 HA genes further evolve in the near future.
Amino Acid Substitution ; China ; Disulfides ; chemistry ; Epitopes ; genetics ; Evolution, Molecular ; Hemagglutinin Glycoproteins, Influenza Virus ; chemistry ; genetics ; immunology ; Humans ; Influenza A Virus, H3N2 Subtype ; genetics ; Mutation ; Phylogeny