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Objective:To compare the optimal conditions, virus yield, viral titer and cell metabolism between culturing influenza virus H1N1 vaccine strain in MDCK and MDCK-G1 cells.Methods:The optimal culture conditions were investigated using chessboard method. The hemagglutination titer, half of the tissue infection dose (TCID 50) and the metabolism of glucose and lactic acid were monitored and compared between the two cell lines. Results:After MDCK-G1 cells were inoculated with H1N1 at the multiplicity of infection (MOI) of 0.001 with the presence of 1 μg/ml of trypsin, the hemagglutination titer reached the peak of 1∶512 at 72 h and the viral titer was 10 7.4TCID 50/ml. In the MDCK cell line group, the hemagglutination titer reached the peak of 1∶256 at 72 h and the viral titer was 10 6.6TCID 50/ml when using H1N1 at MOI=0.0001 and 1 μg/ml of trypsin. Conclusions:MDCK-G1 cells were more suitable than MDCK cells for the proliferation of influenza virus. This study provided reference data for further research on cell-derived influenza vaccine.
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Objective@#To investigate the best amount of TPCK trypsin in Madin Darby canine kidney (MDCK) cell suspension for the culture of H7N9 avian influenza virus.@*Methods@#Different concentrations of TPCK trypsin were added during the periods of cell growth and virus production. Their effects on cell growth, viability, glucose and lactate metabolism, and hemagglutination titer were monitored every 12 h. Inter-batch differences were analyzed. The amount of trypsin added in the cell growth phase was 0, 1 μg/ml, 2 μg/ml, 4 μg/ml, 6 μg/ml, 8 μg/ml, 10 μg/ml and 15 μg/ml. The amount of trypsin added during the virus production period was 0, 0.5 μg/ml, 1 μg/ml, 1.5 μg/ml, 2 μg/ml and 2.5 μg/ml. When the hemagglutination titers were same, the adding amount was further optimized at different multiplicity of infection (MOI) of 0.001, 0.005, 0.025 and 0.05.@*Results@#No significant linear effects of TPCK trypsin concentration on cell number, viability, and glucose and lactate metabolism were observed. No toxicity to cell growth was observed when TPCK trypsin concentration reached 15 μg/ml. After the inoculation of H7N9 avian influenza virus, the hemagglutination titers in the 1 μg/ml, 1.5 μg/ml, 2 μg/ml and 2.5 μg/ml TPCK trypsin groups reached the peaks at 48 h, which were 1∶26.5. At 60 h, the hemagglutination titers of the latter two groups decreased faster than those of the former two groups. When the MOI was 0.005, the hemagglutination titer of the 1.5 μg/ml group at 48 h was 26.5 higher than 26 in the 1 μg/ml group under the same condition. There were differences between different batches of TPCK trypsin.@*Conclusions@#Adding 1 μg/ml and 1.5 μg/ml of trypsin could better promote the proliferation of H7N9 avian influenza virus, and 1.5 μg/ml of trypsin had a wider range of MOI applicability.
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Objective@#To reduce the residual proteins and DNA of host cells in the preparation of H5N1 influenza A virus.@*Methods@#Core 700 was firstly used to remove residual host cell proteins, and then Capto Q was used to remove host cell DNA. Several batches of H5N1 influenza A virus cultured in Madin-Darby canine kidney (MDCK) cells were purified using this method. The efficiency of purification was evaluated using many methods including quantitative real-time PCR, hemagglutination (HA) test and single radial immunodiffusion assay. Moreover, Benzonase nuclease was used for comparison.@*Results@#Without the use of Benzonase nuclease, the overall removal rates of host cell DNA and residual proteins were 99.62% and 98.1%, and the HA antigen recovery rate was 66.96%.@*Conclusions@#This study established a purification strategy with good effect for cell-based influenza vaccines. It can efficiently remove host cell DNA and proteins and achieve a high HA recovery rate. The purification result is no worse than that of adding Benzonase nuclease, suggesting the potential of its application in actual vaccine production.
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Objective To reduce the residual proteins and DNA of host cells in the preparation of H5N1 influenza A virus. Methods Core 700 was firstly used to remove residual host cell proteins, and then Capto Q was used to remove host cell DNA. Several batches of H5N1 influenza A virus cultured in Ma-din-Darby canine kidney ( MDCK) cells were purified using this method. The efficiency of purification was evaluated using many methods including quantitative real-time PCR, hemagglutination ( HA) test and single radial immunodiffusion assay. Moreover, Benzonase nuclease was used for comparison. Results Without the use of Benzonase nuclease, the overall removal rates of host cell DNA and residual proteins were 99. 62％ and 98. 1％, and the HA antigen recovery rate was 66. 96％. Conclusions This study established a purification strategy with good effect for cell-based influenza vaccines. It can efficiently remove host cell DNA and proteins and achieve a high HA recovery rate. The purification result is no worse than that of adding Benzonase nuclease, suggesting the potential of its application in actual vaccine production.
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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.
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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.
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To study the complete genomic sequence, genomic characteristics, and genetic variation of the bovine papillomavirus 2 genotype (BPV-2) Aks-01 strain at the molecular level, genotyping of this strain from the skin samples of cows in southern Xinjiang (China) was first detected by the polymerase chain reaction with FAP59/FAP64 primers. Based on the complete genome of the BPV-2 reference strain, specific primers and sequencing primers were designed, and the complete genome of the Aks-01 strain amplified and sequenced. Sequence analyses showed that genotyping of the Aks-01 strain belonged to BPV-2. The Aks-01 strain had the structural characteristics of BPV-2. The 7944-bp full-length genomic sequence of the Aks-01 strain was compiled using DNAStar™. The sequence of the Aks-01 strain had 98% similarity to the reference strain from GenBank. The Aks-01 strain was most closely related to BPV-1 and BPV-13. BPV-2, BPV-1 and BPV-13 were grouped within the genus Deltapapillomavirus. The Aks-01 strain is the first BPV-2 strain reported in southern Xinjiang.