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研究狂犬病病毒CTNCEC25株的增殖能力、病毒的感染力及致病力等表型特征,为将该毒株应用于疫苗生产提供基础信息。将CTNCEC25株及其母株CTN-1株分别接种非洲绿猴肾细胞(Vero细胞)、原代鸡胚成纤维细胞(Primary chicken embryo cell,CEC)和小鼠神经瘤母细胞(Mouse neuroblastoma N2acells,N2a),观察其细胞病变特性及其复制规律;以脑内注射的方式接种动物,观察其对乳鼠、小鼠、豚鼠及家兔等动物的致病力;同时还将CTNCEC25株在CEC上连续传20代,在乳鼠脑内传3代,观察其增殖能力、病毒感染力及致病力的稳定性。研究结果表明,与母株CTN-1株相比,CTNCEC25株在Vero细胞和N2a细胞上的增殖并没有显著差异,但在CEC上,CTNCEC25株具有明显优势,其72h滴度可达10~(7.5~7.6) FFU/mL,而CTN-1株滴度只有10~(5.8) FFU/mL;此外CTNCEC25株在N2a细胞、Vero细胞及CEC上均可产生明显的细胞病变。脑内致病力实验结果表明,CTNCEC25株虽然对1~3日龄的乳鼠仍然具有较强的脑内致病性,但对成年小鼠的脑内致病力大大减弱,对豚鼠和家兔不致病。经细胞和乳鼠脑内连续传代,CTNCEC25株在CEC上增殖稳定,其减弱的毒力并未回升,无弱毒返祖现象。综上所述,本研究初步证实CTNCEC25株是一株高度减毒、不易返祖的弱毒株,其在CEC上可稳定、快速增殖,病毒滴度高(>10~(7.0) FFU/mL),是一株安全性较高的狂犬病疫苗候选毒株。
To study the phenotypic characteristics of rabies virus CTNCEC25 strain, its virulence and virulence and provide the basic information for its application in vaccine production. The CTNCEC25 strain and its parent CTN-1 strain were inoculated with Vero cells, primary chicken embryo cells (CEC) and mouse neuroblastoma N2 cells N2a) were used to observe the cytopathic features and their replication rules. The animals were inoculated intracerebrally to observe the pathogenicity of the CTNCEC25 strains on the CECs of mice, mice, guinea pigs and rabbits. 20 consecutive generations, in the third generation of neonatal rat brain, observe its proliferation, viral infectivity and pathogenicity stability. The results showed that there was no significant difference in the proliferation of CTNCEC25 strains between Vero cells and N2a cells compared with the parent strain CTN-1 strain, but CTNCEC25 strain had obvious advantages on CEC, 7.5 ~ 7.6) FFU / mL, while the titer of CTN-1 strain was only 10 ~ (5.8) FFU / mL. In addition, CTNCEC25 strain could produce obvious cytopathic effect on N2a cells, Vero cells and CECs. The results of in-vivo pathogenicity test showed that although the CTNCEC25 strain still has strong intracerebral pathogenicity in 1-3 days old suckling mice, its pathogenicity in the brain of adult mice was greatly weakened, Rabbit is not sick. After continuous passage in the brain of neonatal and neonatal rats, the CTNCEC25 strain proliferated steadily on CEC, its weakened virulence did not rise, and no attenuated anapogenetic phenomenon was found. In conclusion, CTNCEC25 strain is a highly attenuated and not easily revertant attenuated strain that can be stably and rapidly proliferated on CEC with a high virus titer (> 10-7.0 FFU / mL) , Is a highly safe rabies vaccine candidate strains.