Objective: To evaluate the immunogenicity of inactivated quadrivalent influenza vaccine (QIV) in adults aged 18-64 years, through a Meta-analysis. Methods: Literature was retrieved by searching the Medline, Cochrane Library, Science Direct in the past decade. All the studies were under random control trial (RCT) and including data related to immunogenicity which involving sero-protection rate (SPR) and sero-conversion rate (SCR) of the QIV, versus inactivated trivalent influenza vaccine (TIV) in the population aged 18 to 64. Revman 5.3 software was employed to manipulate the pooled date of the included literature. Result: A total of 8 studies for the SPR and SCR of the shared strains (two A lineage and one B lineage) were included. There appeared no significant differences in the response rates between the two vaccines. As for QIV versus TIV (B/Yamagata), the pooled RR of the SPR for B/Victoria was 1.28 (95%CI: 1.08-1.51, P<0.05), with the pooled RR of the SCR for B/Victoria as 1.94 (95%CI: 1.50-2.50, P<0.05). For QIV versus TIV (B/Victoria), the pooled RR of the SPR for B/Yamagata as 1.10 (95%CI: 1.02-1.18, P<0.05), and the pooled RR of SCR for B/Yamagata as 1.99 (95%CI: 1.34-2.97, P<0.05). Conclusion: In the population aged 18-64 years, inactivated QIV was equivalently immunogenic against the shared three strains included in the activated TIV while a superior immunogenic effect was noticed in the vaccine strain which did not include the inactivated QIV.
Adolescent ; Adult ; Antibodies, Viral/blood* ; Drug-Related Side Effects and Adverse Reactions ; Hemagglutination Inhibition Tests ; Humans ; Influenza A virus/immunology* ; Influenza B virus/immunology* ; Influenza Vaccines/immunology* ; Influenza, Human/prevention & control* ; Middle Aged ; Vaccines, Inactivated/immunology* ; Young Adult

Preparation of maternal strain A/PR/8/34 HA antiserum for influenza virus classical reassortment. A/PR/8/34 virus was digested by bromelain after inactivation and purification. 5%-20% sucrose continuous density gradient centrifugation method was used to purify HA protein. SIRD method was used to select the target protein. SDS-PAGE method was used to identified HA protein. High Immunogenic A/PR/8/34 HA protein was successfully prepared and HI titer reached 10240. High purity HA antiserum was identified by SIRD method. The key reagent in the classical reassortment of influenza virus was prepared, and the complete set of technical methods were explored, which laid the foundation for the independent research and development of seasonal influenza vaccine strains of China.
Animals ; Antibodies, Viral ; immunology ; Electrophoresis, Polyacrylamide Gel ; Female ; Hemagglutination Inhibition Tests ; Hemagglutinin Glycoproteins, Influenza Virus ; analysis ; immunology ; Humans ; Influenza A Virus, H1N1 Subtype ; genetics ; immunology ; Influenza, Human ; immunology ; virology ; Rabbits ; Reassortant Viruses ; genetics ; immunology

OBJECTIVETo investigate the preventive effects of Qiangzhi Decoction (, QZD) on influenza A pneumonia through inhibition of inflammatory cytokine storm in vivo and in vitro.

METHODSOne hundred ICR mice were randomly divided into the virus control, the Tamiflu control and the QZD high-, medium-, and low-dose groups. Mice were infected intranasally with influenza virus (H1N1) at 10 median lethal dose (LD50). QZD and Tamiflu were administered intragastrically twice daily from day 0 to day 7 after infection. The virus control group was treated with distilled water alone under the same condition. The number of surviving mice was recorded daily for 14 days after viral infection. The histological damage and viral replication and the expression of inflammatory cytokines were monitored. Additionally, the suppression capacity on the secretion of regulated on activation normal T cells expressed and secreted (RANTES) and tumor necrosis factor-α (TNF-α) in epithelial and macrophage cell-lines were evaluated.

RESULTSCompared with the virus control group, the survival rate of the QZD groups significantly improved in a dose-dependent manner (P<0.05), the viral titers in lung tissue was inhibited (P<0.05), and the production of inflammatory cytokines interferon-γ (IFN-γ), interleukin-6 (IL-6), TNF-α, and intercellular adhesion molecule-1 (ICAM-1) were suppressed (P<0.05). Meanwhile, the secretion of RANTETS and TNF-α by epithelial and macrophage cell-lines was inhibited with the treatment of QZD respectively in vitro (p<0.05) CONCLUSIONS: The preventive effects of QZD on influenza virus infection might be due to its unique cytokine inhibition mechanism. QZD may have significant therapeutic potential in combination with antiviral drugs.

Animals ; Cell Line ; Cell Survival ; drug effects ; Chemokine CCL5 ; metabolism ; Chemokines ; metabolism ; Cytokines ; metabolism ; Dogs ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Enzyme-Linked Immunosorbent Assay ; Hemagglutination, Viral ; drug effects ; Humans ; Inflammation ; pathology ; Influenza A Virus, H1N1 Subtype ; drug effects ; physiology ; Influenza A Virus, H1N2 Subtype ; drug effects ; Lung ; drug effects ; pathology ; Madin Darby Canine Kidney Cells ; Mice, Inbred ICR ; Orthomyxoviridae Infections ; complications ; pathology ; prevention & control ; Pneumonia ; complications ; pathology ; prevention & control ; Protective Agents ; pharmacology ; therapeutic use ; Survival Rate ; Tumor Necrosis Factor-alpha ; pharmacology

Hemagglutinin (HA) contains a head domain with a high degree of variability and a relatively conserved stem region. HA is the major viral antigen on the surface of the influenza virus. To define the biologic activities of chimeric HA bearing different head domains and stem regions or their potential use, a HA chimeric gene containing the head domain of the H7 subtype virus and stem region of the H3 subtype virus was modified and expressed using a baculovirus expression vector. Then, the secreted protein was purified and its biologic activities characterized. Approximately 1.4 mg/mL cH7/3 HA could be obtained, and its molecular weight was ≈ 70 kD. The trimer form of cH7/3 protein had hemagglutination activity and could be recognized by specific antibodies. The method described here can be used for further studies on the screening of HA stem-reactive antibodies or the development of vaccines with conserved epitopes.
Antibodies, Viral ; immunology ; Baculoviridae ; genetics ; metabolism ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Hemagglutination ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; immunology ; Humans ; Influenza Vaccines ; genetics ; immunology ; Influenza, Human ; prevention & control ; virology

In order to study the proliferation inhibition effect of H5N1 subtype avian influenza virus (AIV) with small interfere RNA (siRNA), a total of 4 siRNAs were designed in accordance with the NP and PA genes of H5N1 subtype AIV, the siRNAs were then transfected to chicken embryo fibroblast(CEF), CEF was infected with H5N1 subtype AIV after 6 hrs. Virus titer of cell supernatant was tested at 16-56hrs post infection, and pathological changes of the cells was observed; mRNA levels of NP, PA, HA and p13-actin gene were tested at 36hrs post infection. The results showed that these 4 siRNAs could inhibit the prolif-eration of H5N1 subtype AIV in CEF in varying degrees, and one siRNA targeting PA was best per-formed. The experimental results also showed that the inhibition effect was decreased with the time prolonged. This research provides a basis for further studying RNAi on AIV prevention and control.
Actins ; genetics ; Animals ; Chick Embryo ; DNA Primers ; genetics ; Fibroblasts ; virology ; Hemagglutination ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; Hemagglutinins ; genetics ; Humans ; Influenza A Virus, H5N1 Subtype ; genetics ; growth & development ; physiology ; RNA Interference ; RNA Replicase ; genetics ; RNA, Small Interfering ; chemical synthesis ; genetics ; RNA-Binding Proteins ; genetics ; Real-Time Polymerase Chain Reaction ; Specific Pathogen-Free Organisms ; Transfection ; Viral Core Proteins ; genetics ; Viral Proteins ; genetics ; Virus Replication

A recombinant hemagglutinin-neuraminidase (rHN) protein from Newcastle disease virus (NDV) with hemagglutination (HA) activity was expressed in Spodoptera frugiperda cells using a baculovirus expression system. The rHN protein extracted from infected cells was used as an antigen in a hemagglutination inhibition (HI) test for the detection and titration of NDV-specific antibodies present in chicken sera. The rHN antigen produced high HA titers of 2(13) per 25 microL, which were similar to those of the NDV antigen produced using chicken eggs, and it remained stable without significant loss of the HA activity for at least 12 weeks at 4degrees C. The rHN-based HI assay specifically detected NDV antibodies, but not the sera of other avian pathogens, with a specificity and sensitivity of 100% and 98.0%, respectively, in known positive and negative chicken sera (n = 430). Compared with an NDV-based HI assay, the rHN-based HI assay had a relative sensitivity and specificity of 96.1% and 95.5%, respectively, when applied to field chicken sera. The HI titers of the rHN-based HI assay were highly correlated with those in an NDV-based HI assay (r = 0.927). Overall, these results indicate that rHN protein provides a useful alternative to NDV antigen in HI assays.
Animals ; Antibodies, Viral/*blood ; Antigens, Viral/*diagnostic use/genetics/metabolism ; Baculoviridae/genetics ; Chickens ; HN Protein/*diagnostic use/genetics/metabolism ; Hemagglutination Inhibition Tests/*methods/veterinary ; Newcastle Disease/*diagnosis/immunology/virology ; Newcastle disease virus/genetics/*immunology/metabolism ; Poultry Diseases/*diagnosis/immunology/virology ; Recombinant Proteins/diagnostic use/genetics/metabolism ; Sf9 Cells ; Spodoptera

In chicken fibroblast cell (CEF) cultures with antiserum against Newcastle disease virus (NDV) strain TZ060107, the virus was passed serially for 50 passages in 3 independent lineages. HN and F genes were amplified and sequenced every 10 passages. The derived virus A1-50 with most mutations among 3 lineages was further passed for another 50 passages in CEF with or without antiserum against A1-50, each in 3 independent lineages. Sequence comparisons for HN and F genes of 60, 70, 80, 90 and 100 passages indicated that the ratio of nonsynonymous mutations (NS) vs synonymous mutations (S) for HN genes in the lineages passed with antiserum against A1-50 was 5.25, which was obviously higher than 2. 375 of NS/ S in the lineages without the antiserum. The stable NS mutations occurred in the first 50 passages with the antiserum against the original TZ060107 were still maintained and one more new stable NS mutation appeared. For the F gene, 3 new stable NS mutations occurred during the second 50 passages in lineages with antiserum against A1-50 when the original NS mutations obtained in the first 50 passages with antiserum against TZ060107 still existed. Cross hemagglutination inhibition (HI) between original virus and its derivative viruses indicated that the more continuous passages in cell culture with antiserum passed, the bigger difference of antigenicity between the virus and the original virus had.
Amino Acid Sequence ; Animals ; Antibodies, Viral ; immunology ; Base Sequence ; Chickens ; Evolution, Molecular ; HN Protein ; genetics ; immunology ; Hemagglutination Inhibition Tests ; Molecular Sequence Data ; Mutation ; Newcastle Disease ; immunology ; virology ; Newcastle disease virus ; genetics ; immunology ; Poultry Diseases ; Viral Fusion Proteins ; genetics ; immunology

A transient four-plasmid cotransfection system was used to construct avian influenza A (H5N1) pseudotyped viral particle (H5N1Pp) by incorporating hemagglutinin (HA) protein and neuraminidase (NA) protein from H5N1 avian influenza virus onto Murine leukemia virus pseudotyped viral particles, the transmission electron microscopy, infectivity titer assay, hemagglutination assay, neutralization assay of H5N1Pp were studied. We established a pseudotyped H5N1 viral particle at a high titer of 10(8) Pp/mL, the morphology,the hemagglutination activity and neutralization specificity of H5N1Pp is simililar to wild H5N1 virus. The research result sets a platform for studying this virus, including its receptors, the functional analysis of HA and NA, neutralizing antibodies and anti-H5N1 drug development.
Animals ; Birds ; Cricetinae ; Genetic Engineering ; HEK293 Cells ; Hemagglutination ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; Humans ; Influenza A Virus, H5N1 Subtype ; genetics ; physiology ; Influenza in Birds ; virology ; Neutralization Tests ; Transfection ; Viral Load ; genetics ; Virion ; genetics

BACKGROUND: Validation of hemagglutination inhibition (HI) assays is important for evaluating antibody responses to influenza virus, and selection of erythrocytes for use in these assays is important. This study aimed to determine the correlation between receptor binding specificity and effectiveness of the HI assay for detecting antibody response to pandemic influenza H1N1 (pH1N1) virus. METHODS: Hemagglutination (HA) tests were performed using erythrocytes from 6 species. Subsequently, 8 hemagglutinating units of pH1N1 from each species were titrated by real-time reverse transcription-PCR. To investigate the effect of erythrocyte binding preference on HI antibody titers, comparisons of HI with microneutralization (MN) assays were performed. RESULTS: Goose erythrocytes showed most specific binding with pH1N1, while HA titers using human erythrocytes were comparable to those using turkey erythrocytes. The erythrocyte binding efficiency was shown to have an impact on antibody detection. Comparing MN titers, HI titers using turkey erythrocytes yielded the most accurate results, while those using goose erythrocytes produced the highest geometric mean titer. Human blood group O erythrocytes lacking a specific antibody yielded results most comparable to those obtained using turkey erythrocytes. Further, pre-existing antibody to pH1N1 and different erythrocyte species can distort HI assay results. CONCLUSIONS: HI assay, using turkey and human erythrocytes, yielded the most comparable and applicable results for pH1N1 than those by MN assay, and using goose erythrocytes may lead to overestimated titers. Selection of appropriate erythrocyte species for HI assay allows construction of a more reliable database, which is essential for further investigations and control of virus epidemics.
Adult ; Animals ; Antibodies, Neutralizing/immunology ; Antibodies, Viral/*analysis/immunology ; Chickens ; Erythrocytes/*metabolism ; Female ; Geese ; *Hemagglutination Inhibition Tests ; Horses ; Humans ; Influenza A Virus, H1N1 Subtype/genetics/immunology/*metabolism ; Influenza, Human/epidemiology/immunology/virology ; Male ; Middle Aged ; Neutralization Tests ; Pandemics ; Swine ; Turkeys

In our previous study, a panel of 52 broadly cross-reactive H5-specific monoclonal antibodies (MAbs) were generated and characterized. The 13D4, one of these MAbs, has been demonstrated to protect mice against lethal challenge by 4 strains of H5N1 avian influenza virus representing the currently prevailing genetic populations, clades 1, 2.1, 2.2, and 2.3. Here, we further cloned the gene of the 13D4 MAb and constructed a single-chain variable fragment. Then, the 13D4 single-chain antibody (scFv) was expressed in secretory maner in Pichia pastoris. The supernatant of the culture was concentrated and subjected to ammonium sulfate precipitation. The purity of the 13D4 scFv was around 90% in SDS-PAGE following ion-exchange chromatography. We further investigated its binding property using hemagglutination inhibition (HI) test and blocking ELISA. The results indicated that the 13D4 scFv shared the same binding sites and comparable HI titer with the prototype murine 13D4 Mab. In conclusion, an anti-H5 single-chain wide-spectrum neutralizing antibody is prepared successfully in yeast system.
Antibodies, Viral ; genetics ; Hemagglutination Inhibition Tests ; Immunoglobulin Fragments ; genetics ; immunology ; Influenza A Virus, H5N1 Subtype ; immunology ; Pichia ; genetics ; Single-Chain Antibodies ; genetics ; immunology