輪狀病毒不同基因型間交叉反應(yīng)研究
發(fā)布時(shí)間：2018-07-28 18:15
【摘要】：輪狀病毒(Rotavirus, RV)是世界范圍內(nèi)導(dǎo)致嬰幼兒嚴(yán)重腹瀉的主要原因。每年大約有60萬(wàn)左右的嬰幼兒因輪狀病毒的感染重癥發(fā)病而死亡,其中80%以上發(fā)生在亞洲南部、非洲南部等經(jīng)濟(jì)及衛(wèi)生條件相對(duì)落后的發(fā)展中國(guó)家和地區(qū),輪狀病毒造成的胃腸炎在世界各國(guó)都有不同程度的經(jīng)濟(jì)與疾病負(fù)擔(dān)。目前尚無(wú)治療輪狀病毒感染的特效藥物,使用疫苗是預(yù)防和控制輪狀病毒感染,降低重癥死亡率的最經(jīng)濟(jì)有效的手段。輪狀病毒基因組由11條片段組成,分別編碼6個(gè)結(jié)構(gòu)蛋白(VP1-VP4與VP6-VP7)和6個(gè)非結(jié)構(gòu)蛋白(NSP1-NSP6)。到目前為止,根據(jù)結(jié)構(gòu)蛋白VP6(組抗原)可以分為A-H 8個(gè)不同的組,其中引起嬰幼兒感染的主要是A組輪狀病毒。另外,依據(jù)結(jié)構(gòu)蛋白VP7(G型抗原或糖蛋白)和VP4(P型抗原或蛋白水解酶敏感蛋白)間的基因差異分為27個(gè)G基因型(G1-G27)和35個(gè)P基因型(P[1]-P[35])。疫苗研發(fā)的型別選擇主要是依據(jù)流行型而定。已有3種輪狀病毒減毒活疫苗用于預(yù)防輪狀病毒感染,分別是葛蘭素史克公司生產(chǎn)的單價(jià)人輪狀病毒疫苗Rotarix(基因型為G1P[8]),默克公司生產(chǎn)的五價(jià)人-牛重組輪狀病毒疫苗RotaTeq(基因型為G1,G2, G3, G4, G6和P[8])以及由蘭州生物制品研究所研發(fā)的單價(jià)輪狀病毒口服活疫苗羅特威(基因型為G10P[12])。現(xiàn)有的輪狀病毒疫苗有單價(jià)和多價(jià),這兩種疫苗在制備工藝及質(zhì)控方面都有所不同,多價(jià)疫苗相對(duì)困難。因此,單型能覆蓋多型的盡可能用單價(jià)疫苗。而問(wèn)題在于能否用單價(jià)疫苗獲得多價(jià)疫苗的效果,從目前研究的結(jié)果表明,尚不能確定其否。盡管最終的效果取決于人體免疫后的資料,但人體資料的獲得周期長(zhǎng),評(píng)價(jià)體系復(fù)雜。因此,建立實(shí)驗(yàn)室評(píng)價(jià)方法對(duì)考慮上述問(wèn)題具有重要的指導(dǎo)意見和前瞻性。本課題研究旨在對(duì)單價(jià)和多價(jià)輪狀病毒免疫原的免疫原性進(jìn)行比較,評(píng)價(jià)單價(jià)免疫原引起的交叉保護(hù)作用與多價(jià)免疫原間的差異。本課題研究選取三種不同基因型的輪狀病毒標(biāo)準(zhǔn)株：Wa(基因型為G1P[8]), SA11(基因型為G3P[1])和Gottfried(基因型為G4P[6]),這三株輪狀病毒標(biāo)準(zhǔn)株單獨(dú)作為單價(jià)輪狀病毒免疫原,將這三株輪狀病毒標(biāo)準(zhǔn)株按照不同的比例進(jìn)行混合制備成多價(jià)輪狀病毒免疫原。實(shí)驗(yàn)動(dòng)物ICR小鼠隨機(jī)分組為8個(gè)組,分別是3個(gè)單價(jià)輪狀病毒免疫原組(Wa組,SA11組,Gottfried組),3個(gè)二價(jià)輪狀病毒免疫原組(Wa+SA11組,Wa+Gottfried組,SA11+Gottfried組),1個(gè)三價(jià)輪狀病毒免疫原組(Wa+SA11+Gottfried組)以及1個(gè)對(duì)照組(PBS組),每組按照相同的總劑量免疫輪狀病毒免疫原。免疫后利用ELISA,中和試驗(yàn)以及IgA ELISA試劑盒等方法分別對(duì)各組血清樣本中的輪狀病毒特異性IgG水平,輪狀病毒特異性中和抗體滴度以及血清IgA濃度進(jìn)行檢測(cè)與評(píng)價(jià),從而對(duì)單價(jià)和多價(jià)輪狀病毒免疫原的免疫原性進(jìn)行比較分析。實(shí)驗(yàn)結(jié)果顯示,單價(jià)輪狀病毒免疫原免疫后,不同基因型之間能產(chǎn)生低于親本株的血清交叉中和抗體；二價(jià)輪狀病毒免疫原誘導(dǎo)的免疫反應(yīng)要強(qiáng)于單價(jià)輪狀病毒免疫原,但在二價(jià)輪狀病毒免疫原的三種組合中,對(duì)Wa株及SA11株的免疫原性強(qiáng)于Gottfried株；三價(jià)輪狀病毒免疫原誘導(dǎo)的免疫反應(yīng)略高于單價(jià)輪狀病毒免疫原,而略低于二價(jià)輪狀病毒免疫原,其中免疫原性仍以Wa株最強(qiáng),SA11次之,Gottfried較低。通過(guò)以上實(shí)驗(yàn)結(jié)果得出結(jié)論,多價(jià)輪狀病毒免疫原比單價(jià)輪狀病毒免疫原能夠誘導(dǎo)產(chǎn)生更多的血清輪狀病毒特異性抗體,產(chǎn)生的血清IgA抗體也高于單價(jià)輪狀病毒免疫原,而且,多價(jià)輪狀病毒免疫原比單價(jià)輪狀病毒免疫原能夠誘導(dǎo)交叉免疫反應(yīng)的速度更快。鑒于以上實(shí)驗(yàn)結(jié)果,目前還不能考慮用單價(jià)輪狀病毒疫苗代替多價(jià)輪狀病毒疫苗。
[Abstract]:Rotavirus (RV) is the main cause of severe diarrhoea in infants worldwide. About 600 thousand of infants and young children die from the severe incidence of rotavirus infection every year. More than 80% of them occur in southern Asia, South Africa and other developing countries and regions with relatively backward economic and health conditions, rotavirus At present, there are no special drugs for the treatment of rotavirus infection, and the use of vaccines is the most economical and effective means to prevent and control rotavirus infection and reduce severe mortality. The rotavirus genome consists of 11 fragments, which encode 6 structural proteins (VP 1-VP4 and VP6-VP7) and 6 non structural proteins (NSP1-NSP6). Up to now, the structural protein VP6 (group antigen) can be divided into 8 different groups of A-H, among which, the main cause of infantile infection is group A rotavirus. In addition, the gene between structural protein VP7 (G type antigen or sugar egg white) and VP4 (P type antigen or protein hydrolase sensitive protein) The differences are divided into 27 G genotypes (G1-G27) and 35 P genotypes (P[1]-P[35]). The type selection of the vaccine development is mainly based on the epidemic type. There are 3 rotavirus attenuated live vaccines used in the prevention of rotavirus infection, respectively, the monovalent human rotavirus vaccine Rotarix (genotype G1P[8]) produced by GlaxoSmithKline Co, Merck Co. The production of the pentavalant bovine rotavirus vaccine RotaTeq (genotype G1, G2, G3, G4, G6 and P[8]) and the orally active rotavirus (genotype of G10P[12]) of the monovalent rotavirus developed by the Lanzhou Institute of biological products. The existing rotavirus vaccines have univalent and polyvalent, and these two vaccines have the preparation process and quality control. As a result, polyvalent vaccines are relatively difficult. Therefore, the single type can cover the multiple type of vaccine as much as possible. The problem is whether a single valent vaccine can be used to obtain the effect of a multivalent vaccine. The results from the present study show that it is not sure. Although the final effect depends on the data of the human body after the epidemic, the cycle of human data is long, The evaluation system is complex. Therefore, the establishment of laboratory evaluation method has important guidance and foresight to consider the above problems. The aim of this study is to compare the immunogenicity of the monovalent and polyvalent rotavirus immunogen, and to evaluate the difference between the cross protection and the multivalent immunogen caused by the monovalent immunogen. Three rotavirus standard strains of different genotypes: Wa (genotype G1P[8]), SA11 (genotype G3P[1]) and Gottfried (genotype G4P[6]), the three rotavirus standard strains were used individually as monovalent rotavirus immunogens, and the three rotavirus standard strains were mixed in different proportions to prepare polyvalent rotavirus free virus. The experimental animal ICR mice were randomly divided into 8 groups, which were 3 monovalent rotavirus immunogen groups (group Wa, group SA11, Gottfried), 3 two valence rotavirus immunogen groups (group Wa+SA11, Wa+Gottfried group, SA11+Gottfried group), 1 trivalent rotavirus immunogen group (Wa+SA11+Gottfried group) and 1 control groups (PBS group), each group was treated by the same group. The same total dose of the immune rotavirus immunogen was taken. After immunization, ELISA, neutralization test and IgA ELISA kit were used to detect and evaluate the specific IgG level of rotavirus, the titer of rotavirus specific neutralization antibody and the concentration of IgA in serum, so that the monovalent and polyvalent rotavirus was used. The immunogenicity of the immunogen was compared and analyzed. The results showed that after immunization of the monovalent rotavirus, different genotypes could produce serum cross neutralization antibody below the parent strain, and the immune response induced by the two rotavirus immunogen was stronger than the monovalent rotavirus immunogen, but three of the two valence rotavirus immunogen. In the combination, the immunogenicity of Wa strain and SA11 strain was stronger than that of the Gottfried strain; the immune response induced by the trivalent rotavirus immunogen was slightly higher than that of the monovalent rotavirus immunogen, but slightly lower than the two rotavirus immunogen, in which the immunogenicity was still the strongest in Wa, SA11 times and low in Gottfried. The immunogen of rotavirus immunogen can induce more serum rotavirus specific antibodies than the monovalent rotavirus immunogen, and the produced serum IgA antibody is also higher than the monovalent rotavirus immunogen. Moreover, the immunogen of the polyvalent rotavirus immunogen can induce the cross immune response faster than the monovalent rotavirus immunogen. It is not possible to replace the rotavirus vaccine with single valent rotavirus vaccine.
【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R725.1
本文編號(hào)：2151188
[Abstract]:Rotavirus (RV) is the main cause of severe diarrhoea in infants worldwide. About 600 thousand of infants and young children die from the severe incidence of rotavirus infection every year. More than 80% of them occur in southern Asia, South Africa and other developing countries and regions with relatively backward economic and health conditions, rotavirus At present, there are no special drugs for the treatment of rotavirus infection, and the use of vaccines is the most economical and effective means to prevent and control rotavirus infection and reduce severe mortality. The rotavirus genome consists of 11 fragments, which encode 6 structural proteins (VP 1-VP4 and VP6-VP7) and 6 non structural proteins (NSP1-NSP6). Up to now, the structural protein VP6 (group antigen) can be divided into 8 different groups of A-H, among which, the main cause of infantile infection is group A rotavirus. In addition, the gene between structural protein VP7 (G type antigen or sugar egg white) and VP4 (P type antigen or protein hydrolase sensitive protein) The differences are divided into 27 G genotypes (G1-G27) and 35 P genotypes (P[1]-P[35]). The type selection of the vaccine development is mainly based on the epidemic type. There are 3 rotavirus attenuated live vaccines used in the prevention of rotavirus infection, respectively, the monovalent human rotavirus vaccine Rotarix (genotype G1P[8]) produced by GlaxoSmithKline Co, Merck Co. The production of the pentavalant bovine rotavirus vaccine RotaTeq (genotype G1, G2, G3, G4, G6 and P[8]) and the orally active rotavirus (genotype of G10P[12]) of the monovalent rotavirus developed by the Lanzhou Institute of biological products. The existing rotavirus vaccines have univalent and polyvalent, and these two vaccines have the preparation process and quality control. As a result, polyvalent vaccines are relatively difficult. Therefore, the single type can cover the multiple type of vaccine as much as possible. The problem is whether a single valent vaccine can be used to obtain the effect of a multivalent vaccine. The results from the present study show that it is not sure. Although the final effect depends on the data of the human body after the epidemic, the cycle of human data is long, The evaluation system is complex. Therefore, the establishment of laboratory evaluation method has important guidance and foresight to consider the above problems. The aim of this study is to compare the immunogenicity of the monovalent and polyvalent rotavirus immunogen, and to evaluate the difference between the cross protection and the multivalent immunogen caused by the monovalent immunogen. Three rotavirus standard strains of different genotypes: Wa (genotype G1P[8]), SA11 (genotype G3P[1]) and Gottfried (genotype G4P[6]), the three rotavirus standard strains were used individually as monovalent rotavirus immunogens, and the three rotavirus standard strains were mixed in different proportions to prepare polyvalent rotavirus free virus. The experimental animal ICR mice were randomly divided into 8 groups, which were 3 monovalent rotavirus immunogen groups (group Wa, group SA11, Gottfried), 3 two valence rotavirus immunogen groups (group Wa+SA11, Wa+Gottfried group, SA11+Gottfried group), 1 trivalent rotavirus immunogen group (Wa+SA11+Gottfried group) and 1 control groups (PBS group), each group was treated by the same group. The same total dose of the immune rotavirus immunogen was taken. After immunization, ELISA, neutralization test and IgA ELISA kit were used to detect and evaluate the specific IgG level of rotavirus, the titer of rotavirus specific neutralization antibody and the concentration of IgA in serum, so that the monovalent and polyvalent rotavirus was used. The immunogenicity of the immunogen was compared and analyzed. The results showed that after immunization of the monovalent rotavirus, different genotypes could produce serum cross neutralization antibody below the parent strain, and the immune response induced by the two rotavirus immunogen was stronger than the monovalent rotavirus immunogen, but three of the two valence rotavirus immunogen. In the combination, the immunogenicity of Wa strain and SA11 strain was stronger than that of the Gottfried strain; the immune response induced by the trivalent rotavirus immunogen was slightly higher than that of the monovalent rotavirus immunogen, but slightly lower than the two rotavirus immunogen, in which the immunogenicity was still the strongest in Wa, SA11 times and low in Gottfried. The immunogen of rotavirus immunogen can induce more serum rotavirus specific antibodies than the monovalent rotavirus immunogen, and the produced serum IgA antibody is also higher than the monovalent rotavirus immunogen. Moreover, the immunogen of the polyvalent rotavirus immunogen can induce the cross immune response faster than the monovalent rotavirus immunogen. It is not possible to replace the rotavirus vaccine with single valent rotavirus vaccine.
【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R725.1
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