Deletions in the putative cell receptor-binding domain of Sindbis virus strain MRE16 E2 glycoprotein reduce midgut infectivity in Aedes aegypti
Deletions in the putative cell receptor-binding domain of Sindbis virus strain MRE16 E2 glycoprotein reduce midgut infectivity in Aedes aegypti
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DOI：
10.1128/jvi.77.16.8872-8881.2003
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发表时间：
2003-08-01
影响因子：
5.4
通讯作者：
Olson, KE
中科院分区：
文献类型：
--
作者：
Myles, KM;Pierro, DJ;Olson, KE
辛德毕斯病毒（甲病毒属; Togaviridde）菌株MRE 16有效感染埃及伊蚊，埃及伊蚊摄取含有8至9 log（10）PFU病毒/ml的血粉。然而，这种病毒的小噬斑变体MRE 16 sp在以相同滴度经口感染后对蚊子的感染很差。为了确定MRE 16和MRE 16 sp病毒之间的遗传差异，我们对MRE 16 sp结构基因进行了测序，发现E2糖蛋白中跨越推定细胞受体结合结构域（CRBD）编码区3'端的90个核苷酸缺失。我们通过构建感染性克隆pMRE 16 icDeltaE 200-Y229和pMRE 16 ic，分别代表缺失和不缺失的MRE 16病毒基因组，研究了这种缺失在蚊子口腔感染中的作用。还构建了第三个感染性克隆pMRE 16 icDeltaE 200-C220，其含有仅延伸至CRBD编码区3 ′末端的较小缺失。pMRE 16 ic衍生的病毒在脊椎动物（BHK-21）和蚊子（C6/36）细胞中以与亲本病毒相同的效率复制，并经口感染A.埃及人。来源于pMRE 16 icDeltaE 200-Y229和pMRE 16 icDeltaE 200-C220的病毒在C6/36和BHK-21细胞中的复制效率比MRE 16 ic病毒低10至100倍。每个缺失突变体都不能很好地感染A.埃及伊蚊，并显着降低中肠感染和传播。然而，通过胸内接种感染后4天，所有病毒在蚊子体内产生几乎相同的滴度（类似于6.0 log（10）PFU/ml）。这些结果表明，E2 CRBD的缺失部分代表了A.埃及人。
The Sindbis virus (Alphavirus; Togaviridde) strain MRE16 efficiently infects Aedes aegypti mosquitoes that ingest a blood meal containing 8 to 9 log(10) PFU of virus/mI. However, a small-plaque variant of this virus, MRE16sp, poorly infects mosquitoes after oral infection with an equivalent titer. To determine the genetic differences between MRE16 and MRE16sp viruses, we have sequenced the MRE16sp structural genes and found a 90-nucleotide deletion in the E2 glycoprotein that spans the 3' end of the coding region for the putative cell-receptor binding domain (CRBD). We examined the role of this deletion in oral infection of mosquitoes by constructing infectious clones pMRE16icDeltaE200-Y229 and pMRE16ic, representing MRE16 virus genomes with and without the deletion, respectively. A third infectious clone, pMRE16icDeltaE200-C220, was also constructed that contained a smaller deletion extending only to the 3' terminus of the CRBD coding region. Virus derived from pMRE16ic replicated with the same efficiency as parental virus in vertebrate (BHK-21) and mosquito (C6/36) cells and orally infected A. aegypti. Viruses derived from pMRE16icDeltaE200-Y229 and pMRE16icDeltaE200-C220 replicated 10- to 100-fold less efficiently in C6/36 and BHK-21 cells than did MRE16ic virus. Each deletion mutant poorly infected A. aegypti and dramatically reduced midgut infectivity and dissemination. However, all viruses generated nearly equal titers (similar to6.0 log(10) PFU/ml) in mosquitoes 4 days after infection by intrathoracic inoculation. These results suggest that the deleted portion of the E2 CRBD represents an important determinant of MRE16 virus midgut infectivity in A. aegypti.