摘要
目的研究乙型脑炎病毒(JaGAr-01株和Nakayama株)持续感染变异株性状与E区基因序列的关系。方法将两种乙脑病毒野生株分别感染人肝癌KN73细胞,建立乙型脑炎病毒持续感染系。采用PFU法进行病毒滴度测定;为探讨持续感染病毒的增殖性,将两种病毒野生株及其持续感染株分别感染KN73细胞;利用E区特异引物以RT-PCR法得到两种病毒E区基因片段,应用基因测序反应进行序列分析,并对两种病毒株E区序列进行比较。结果在KN73细胞中两种持续感染株的增殖性比野生株明显低下。E区基因测序显示与JaGAr-01野生株比较,其持续感染变异株有4个氨基酸发生置换(第61位酪氨酸→天门冬氨酸,第219位组氨酸→酪氨酸,第384位缬氨酸→谷氨酸,第418位脯氨酸→丙氨酸);持续感染Nakayama株与其野生株相比有11个氨基酸发生置换(第51位精氨酸→丝氨酸,第61位酪氨酸→天门冬氨酸,第83位赖氨酸→谷氨酸,第123位丝氨酸→精氨酸,第209位精氨酸→赖氨酸,第227位脯氨酸→丝氨酸,第276位天门冬氨酸→丝氨酸,第290位精氨酸→赖氨酸,第387位赖氨酸→精氨酸,第418位亮氨酸→脯氨酸,第454位精氨酸→甘氨酸)。对比还显示基因变异后的持续感染JaGAr-01株和持续感染Nakayama株氨基酸排列的相同性达99.2%。结论乙脑病毒的持续感染变异株增殖性低于野生株;乙脑病毒变异株E区存在基因变异。
Objective To investigate the relationship of E sequence analysis of two different mutant strains of Japanese encephalitis virus and their characters during persistent infection. Methods Two wild strains of Japanese encephalitis viruses infected human hepatoma cell line respectively, the persistent infection model was established. Viral titers were examined by plaque methods using BHK cells. To explore the multiplication of the persistently-infected viruses, KN73 were infected by them. Nucleotides of the E coding region of four Japanese encephalitis strains, two wild and two mutant viruses, from infected cells were amplified by reverse transcription- polymerase chain reaction（RT-PCR） using specific primers. PCR products were sequenced by ABI-PRSM^TM 310 sequencing system and then E sequence of four strains were compared. Results The multiplication of two persistently-infected strains was obviously lower than the relevant wild strains in KN73 cells. Comparing to JaGAr-01 wild strains, four amino acids were replaced（E61Tyr→Asp, E219His→Tyr, E384Val→Glu, F418Pro→Ala） in the E sequence in JaGAr-01 persistent infection mutant strains. Eleven amino acid replacement（E51Arg→Ser, E61Tyr →Asp, E83Lys→Glu, E123Ser→ Arg, E209Arg→ Lys, E227Pro→ Sex, E276Asp→ Sex, E290Arg→ Lys, E3S7Lys→Arg, E418Leu→Pro, E454Arg→Gly） was also noted when comparing the E sequence between persistently infected Nakayama and its wild strains. Homology of amino acid sequence between mutant strains JaGAr-01 and Nakayama was 99.2% as well. Conclusion The multiplication of the persistently-infected mutant strains was lower than the wild strains. Genovariation existed in E region of persistently infected mutant Japanese encephalitis virus.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2006年第5期389-393,共5页
Chinese Journal of Microbiology and Immunology
