NEW APPROACHES TO VACCINES AGAINST THE TICK-BORNE ENCEPHALITIS VIRUS COMPLEX

以前，构建了活的嵌合黄病毒，其含有 蜱传脑炎病毒（TBEV）结构蛋白CME或ME基因 其余的基因来源于登革4型病毒（DEN 4）。 的 ME嵌合体保留了其TBEV亲本小鼠的神经毒力， 它的M和E基因是来自，但它缺乏外围的， TBEV的侵袭性。 ME嵌合体进行突变， 分析，试图减少或消除神经毒力表现， 病毒被直接注入大脑。 三种不同的突变 独立地与小鼠 神经毒性 这些突变消除了：（i）TBEV PreM切割， 位点，其是M蛋白的适当加工所需的;（ii）TBEV E（包膜糖蛋白）糖基化位点;或（iii）第一DEN 4 NS 1 （非结构蛋白1）糖基化位点。 每个3 减毒突变体在猴和蚊子中的生长都受到限制 细胞 值得注意的是，用这些减毒的 突变体诱导小鼠对致死性脑炎的完全抗性， 通过随后用高神经毒性ME嵌合体攻击。 这些 观察结果表明，一种新的策略，发展减毒活TBEV 疫苗 与高毒性TBEV不同，天然存在的相关Langat 病毒（LGT）对小鼠的致病性明显较低。 的遗传分析 LGT可以帮助我们确定 TBE病毒的神经侵袭性和神经毒力。 的RNA基因组 LGT（菌株TP 21）的长度为10940 nt，包含开放的阅读码 由3，414个氨基酸组成的多聚蛋白的框架。 5'非编码区 长度为129 nt，其中nt。1-25和NTS。80-128是保守的， 相关TBEV或Powassan病毒（TBE）的相应区域 北美病毒）。 LGT在3'端非编码区含有583个核苷酸 其中最后90个核苷酸在TBE复合体的病毒中是保守的。 有可能研制出安全有效的TBEV减毒活疫苗 通过构建表达LGT的DEN 4-LGT嵌合病毒的疫苗 抗原性 与先前发现的突变相似的减弱突变 将被引入嵌合病毒基因组和子代病毒中 分析免疫原性和毒力丧失。

Previously, viable chimeric flaviviruses were constructed that contained tick-borne encephalitis virus (TBEV) structural protein CME or ME genes with the remaining genes derived from dengue type 4 virus (DEN4). The ME chimera retained the neurovirulence for mice of its TBEV parent from which its M and E genes were derived, but it lacked the peripheral invasiveness of TBEV. The ME chimera was subjected to mutational analysis in an attempt to reduce or ablate neurovirulence manifest when virus is inoculated directly into the brain. Three distinct mutations were independently associated with marked reduction of mouse neurovirulence. These mutations ablated: (i) the TBEV PreM cleavage site which is required for proper processing of M protein; (ii) the TBEV E (envelope glycoprotein) glycosylation site; or (iii) the first DEN4 NS1 (non-structural protein one) glycosylation site. Each of the 3 attenuated mutants was restricted in growth in both simian and mosquito cells. Significantly, parenteral inoculation with these attenuated mutants induced complete resistance in mice to fatal encephalitis caused by subsequent challenge with the highly neurovirulent ME chimera. These observations suggest a new strategy for developing a live attenuated TBEV vaccine. Unlike the highly virulent TBEV, the naturrally occurring related Langat virus (LGT) is markedly less pathogenic for mice. Genetic analysis of the LGT may allow us to identify the molecular basis for neuroinvasiveness and neurovirulence of TBE viruses. The RNA genome of LGT (strain TP21) is 10940 nt in length and contains an open reading frame for a polyprotein of 3,414 amino acids. The 5' noncoding region is 129 nt in length of which nts. 1-25 and nts. 80-128 are conserved in the corresponding regions of the related TBEV or Powassan virus (a TBE virus of North America). LGT contains 583 nt in the 3' noncoding region of which the last 90 nt are conserved among viruses of the TBE complex. It may be possible to develop a safe and effective live attenuated TBEV vaccine by constructing DEN4-LGT chimeric viruses that express LGT antigenicity. Attenuating mutations similar to those identified earlier will be introduced in the chimeric virus genome and progeny virus analyzed for immunogenicity and loss of virulence.