【摘要】： 戊型肝炎病毒(HEV)是戊型肝炎的病原體,隨著HEV流行病學(xué)研究的深入,發(fā)現(xiàn)HEV的4個(gè)主要基因型存在著明顯的易感宿主差異:基因1型(HEV-1)和基因2型(HEV-2)僅分離于人類,能導(dǎo)致大規(guī)模的戊肝爆發(fā)流行,實(shí)驗(yàn)動(dòng)物目前只成功感染非人靈長(zhǎng)類,而基因3型(HEV-3)和基因4型(HEV-4)人畜共患,僅見(jiàn)于小規(guī)模流行和臨床散發(fā)。我們將HEV分為兩類:包括HEV-1/2的H(Human)類和包括HEV-3/4的Z(Zoonosis)類。 首先我們通過(guò)Western blot、體外捕獲PCR、ELISA阻斷實(shí)驗(yàn)及合成的多肽庫(kù)對(duì)實(shí)驗(yàn)室制備的多株抗HEV單抗的識(shí)別表位進(jìn)行系統(tǒng)的研究,結(jié)果發(fā)現(xiàn)12株線性單抗識(shí)別的位都位于ORF2 aa408-458之間,17株構(gòu)象型單抗識(shí)別表位都定位于ORF2 aa459-606之間,其中15株的識(shí)別表位位于天然病毒表面。 現(xiàn)有研究結(jié)果表明HEV的主要中和表位區(qū)域集中于ORF2的aa459-606之間,并且也是主要介導(dǎo)HEV與嗜性細(xì)胞吸附的區(qū)域。本研究通過(guò)比較這兩類HEV ORF2 aa368-606區(qū)段,發(fā)現(xiàn)存在4個(gè)類保守的差異位點(diǎn),均位于HEV的主要中和表位區(qū)域,分別是Ser483Thr、Val492Met、Ser497Thr和Ala599Gly。以能形成類病毒顆粒的HEV 239(ORF2 aa368-606)為基礎(chǔ),對(duì)這四個(gè)位點(diǎn)進(jìn)行定點(diǎn)替換突變,并以這15株能夠捕獲HEV-1和/或HEV-4的單克隆抗體比較各種突變體的免疫反應(yīng)性,結(jié)果表明僅aa497的差異造成了這兩類HEV中和表位構(gòu)象的部分差異,提示aa497及其相關(guān)的病毒表面結(jié)構(gòu)差異可能在H類和Z類HEV宿主選擇中扮演重要角色。
文內(nèi)圖片：
圖片說(shuō)明：HEV基因組Fig1GenomicorganizationofHEV(A)ingenotype1-3and(B)ingenotype4
[Abstract]:Hepatitis E virus (HEV) is the pathogen of hepatitis E. with the deepening of HEV epidemiological research, it is found that there are obvious susceptible host differences in the four main genotypes of HEV: gene type 1 (HEV-1) and gene type 2 (HEV-2) are only isolated from human beings, which can lead to large-scale outbreak of hepatitis E. At present, experimental animals are only successfully infected with non-human primates. Gene 3 (HEV-3) and gene 4 (HEV-4) zoonosis are only found in small-scale epidemic and clinical dissemination. We divide HEV into two categories: the H (Human) class of HEV-1/2 and the Z (Zoonosis) class of HEV-3/4. Firstly, we systematically studied the recognition epitopes of several strains of anti-HEV monoclonal antibodies prepared in laboratory by Western blot, capture PCR,ELISA blocking experiment in vitro and the synthetic polypeptide library. The results showed that the recognition epitopes of 12 strains of linear monoclonal antibodies were located between ORF2 aa408-458 and 17 strains of constructed monoclonal antibodies were located between ORF2 aa459-606, and 15 of them were located on the surface of natural viruses. The existing results show that the main neutralizing epitope region of HEV is concentrated between the aa459-606 of ORF2, and it is also the region that mediates the adsorption of HEV with sex-like cells. In this study, by comparing the two types of HEV ORF2 aa368-606 regions, it was found that there were four conserved difference sites, all of which were located in the main neutralizing epitope regions of HEV, Ser483Thr,Val492Met,Ser497Thr and Ala599Gly., respectively. The site-directed substitution mutations of these four loci were carried out on the basis of HEV-239( ORF2 aa368-606), which can form viral particles, and the immunoreactivity of various mutants was compared with the 15 monoclonal antibodies that could capture HEV-1 and / or HEV-4. The results showed that only the difference of aa497 resulted in some differences in the conformational patterns of neutralizing epitopes between the two types of HEV. It is suggested that the surface structure difference of aa497 and its related viruses may play an important role in the host selection of class H and Z HEV.
【學(xué)位授予單位】：廈門大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2007
【分類號(hào)】：R392

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