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

此前，构建的具有活性的嵌合黄病毒含有 森林脑炎病毒结构蛋白CME或ME基因 其余的基因来自登革4型病毒(DEN4)。这个 Me嵌合体保留了其亲本TBEV对小鼠的神经毒力 它的M和E基因都是派生出来的，但它缺乏外周 TBEV的侵袭性。ME嵌合体发生突变 分析试图降低或消融神经毒力时表现出来的 病毒被直接接种到大脑中。三种截然不同的突变 与小鼠的显著减少独立相关 神经毒性。这些突变导致：(I)TBEV Prem裂解 正确处理M蛋白所需的位点；(Ii)TBEV E(包膜糖蛋白)糖基化位点；或(Iii)第一个DEN4 NS1 (非结构蛋白1)糖基化位点。这三个人中的每一个 弱化突变体在猿猴和蚊子中的生长都受到限制。 细胞。值得注意的是，非肠道接种这些疫苗会减弱 突变体诱导小鼠对致死性脑炎的完全抵抗力 通过随后对高度神经毒力的ME嵌合体的挑战。这些 观察结果提出了一种开发活体衰减TBEV的新策略 疫苗。 与高毒力的TBEV不同，发生的自然致病与拉加特有关 病毒(LGT)对小鼠的致病性明显较低。遗传病的分析 LGT可能会让我们确定 TBE病毒的神经侵袭性和神经毒力。沙门氏菌的RNA基因组 Lgt(tp21株)全长10940个核苷酸，含有一个开放读数 一个由3,414个氨基酸组成的多蛋白框架。5‘非编码区 是129个碱基，其中NTS.1-25和NTS。80-128是保守的 相关的TBEV或Powassan病毒(一种TBE)的相应区域 北美病毒)。LGT在3‘非编码区含有583个核苷酸 其中最后90个核苷酸在TbE复合体的病毒中保守。 研制一种安全有效的减毒活病毒是可能的。 构建表达LGT的DEN4-LGT嵌合病毒疫苗 抗原性。减弱与早先发现的突变类似的突变 将被引入嵌合病毒基因组和子代病毒 分析了免疫原性和毒力损失。

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.