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目的对肾综合征出血热灭活疫苗毒株84Fli株的M和S基因进行核苷酸序列分析,了解该毒种的基因稳定性。方法根据汉坦病毒标准株设计特异的PCR引物,用RT-PCR技术分段扩增疫苗株84FLi株的M和S基因片段,PCR产物纯化后直接测序,并进行遗传进化分析。结果疫苗株84FLi株M基因片段的核苷酸序列与GenBank中收录的HTN型毒株的M基因核苷酸序列的同源性为83.5%～99.9%,而与SEO型毒株的同源性为70.2%～72.0%。其S基因与HTN型毒株的同源性为85.9%～99.6%,与SEO型毒株的同源性为69.4%～70.8%。疫苗株84FLi株的M和S基因的氨基酸序列与GenBank中收录的HTN型汉坦病毒84FLi株M和S基因的氨基酸序列的同源性分别为99.9%和99.5%。从遗传进化树上可以看出,疫苗株84FLi株与GenBank中收录的84FLi株位于一个独立的分支上,与其他HTN型病毒亚型分布于不同分支上,与其他HTN型病毒亚型亲缘关系较远。结论肾综合征出血热灭活疫苗毒株84FLi株M和S基因片段的核苷酸和氨基酸序列未发生较大变异,说明在传代过程中该疫苗株在基因水平上并未发生较大改变。
Objective To analyze the nucleotide and amino acid sequence of M and S gene of 84Fli strain of inactivated vaccine of hemorrhagic fever with renal syndrome and to understand the gene stability of this strain. Methods Specific PCR primers were designed according to Hantavirus standard strains. The M and S gene fragments of vaccine strain 84FLi were amplified by RT-PCR. The PCR products were directly purified after sequencing and analyzed by genetic evolution. Results The nucleotide sequence of M gene fragment of vaccine strain 84FLi was 83.5% -99.9% with the nucleotide sequence of M gene of HTN strain in GenBank, but the homology with the SEO strain 70.2% ~ 72.0%. The homology between S gene and HTN strain was 85.9% -99.6%, and the homology with SEO strain was 69.4% -70.8%. The amino acid sequences of the M and S genes of the vaccine strain 84FLi were 99.9% and 99.5% homologous to the amino acid sequences of the M and S genes of the HTFL-type Hantavirus 84FLi strain contained in GenBank, respectively. It can be seen from the phylogenetic tree that the vaccine strain 84FLi is located on an independent branch with the 84FLi strain contained in GenBank and is distributed on different branches with other HTN virus subtypes and has a higher genetic relationship with other HTN virus subtypes far. Conclusion The nucleotide and amino acid sequences of the M and S gene fragments of 84FLi strain isolated from the inactivated vaccine for hemorrhagic fever with renal syndrome do not change significantly, indicating that the vaccine strain did not change significantly at the gene level during passage.