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目的分析2013年流感疫苗生产用甲型H3N2(NYMCX-223A)毒株主要抗原血凝素(hemagglutinin,HA)和神经氨酸酶(neuraminidase,NA)基因特征,并检测该疫苗株主种子批、工作种子批及疫苗病毒收获液的传代稳定性。方法对制备的H3N2疫苗株的主种子批、工作种子批及疫苗病毒收获液进行全面的生物学特性检测;采用RT-PCR法从主种子批、工作种子批及疫苗病毒收获液中扩增HA及NA基因片段,并进行全基因序列测定及分析;应用MEGA 5.05软件对7株不同年份的H3N2亚型疫苗株的HA氨基酸序列进行比对,绘制基因种系发生树。结果 H3N2疫苗株的主种子批、工作种子批抗原性与2013年度WHO推荐毒株相一致,其他生物学特性均符合《中国药典》三部(2010版)标准;主要抗原HA基因序列长度为1 701 bp,编码566个氨基酸;NA基因序列长度为1 410 bp,编码469个氨基酸,各代次流感病毒HA和NA基因核苷酸和氨基酸序列均一致,与GenBank公布的序列完全一致,同源性为100%。2013年与2012年H3N2疫苗株HA氨基酸序列相比,同源性为97.5%。2013年与2012年H3N2疫苗株具有较远的进化距离,与同源性结果相对应。结论 2013年甲型H3N2(NYMCX-223A)流感疫苗株主要抗原基因传代稳定,一般生物学特性符合《中国药典》三部(2010版)要求。2013年与2012年H3N2疫苗株序列差异较大。
Objective To analyze the characteristics of the major antigen hemagglutinin (HA) and neuraminidase (NA) genes of influenza A virus H3N2 (NYMCX-223A) in 2013 for influenza vaccine production. Seeding batch and vaccine virus harvest liquid passage stability. Methods The biological characteristics of the main seed batch, working seed batch and vaccine virus harvested liquid of the prepared H3N2 vaccine strain were tested. The main HA seedlings, working seed lots and vaccine virus harvested liquid were amplified by RT-PCR And NA gene fragment were amplified and sequenced. The MEGA 5.05 software was used to compare the HA amino acid sequences of seven H3N2 subtype vaccine strains of different years to draw the gene phylogenetic tree. Results The main seed batches and working seed batches of H3N2 vaccine strains were consistent with the WHO recommended strains in 2013 and all the other biological characteristics were in line with the standards of Chinese Pharmacopoeia (2010 edition). The length of the major antigen HA gene was 1 701 bp, encoding a protein of 566 amino acids. The length of NA gene was 1 410 bp, encoding 469 amino acids. The nucleotide and amino acid sequences of HA and NA genes of each sub-influenza virus were the same, which were identical with those published in GenBank. Sex is 100%. The homology of the HA amino acid sequence of H3N2 vaccine strains in 2013 and 2012 was 97.5%. The 2013 and 2012 H3N2 vaccine strains had far evolutionary distances, corresponding to homology results. Conclusion The main antigen genes of influenza A H3N2 (NYMCX-223A) vaccine strains were successfully passaged in 2013 and the general biological characteristics are in line with the requirements of the third edition of the Chinese Pharmacopoeia (2010 edition). The sequences of H3N2 vaccine strains in 2013 and 2012 are quite different.