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目的 了解我国西部新发现的两种HBV C/D基因型重组体的基因型特点.方法 采用PCR对来自我国西部地区的17株HBV全基因组序列进行扩增,并利用生物信息学软件对其全基因组结构、遗传距离及重组位点进行分析.结果 两种HBV C/D基因型重组体与HBV基因型A、B、D、E、F的异质性>8%,而与C基因型的异质性为3.8%~8.0%.进化树分析发现,所有C/D重组体与基因型C形成一个大分支,但又独立于C基因型的其他5种哑型而形成明确的另外两个分支,即两个新的HBV/C基因亚型.结论 两种HBVC/D重组体从遗传距离分析属于与C1~C5亚型不同的新的HBV/C基因亚型. 相似文献
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《世界华人消化杂志》2016,(26)
目的区分D基因型重组类型,了解青海地区乙型肝炎病毒(hepatitis B virus,HBV)感染者C/D基因型重组情况.方法收集217份青海地区慢性乙型肝炎感染者样品,PCR扩增535-1460 nt和1779-2400 nt2个片段的乙型肝炎病毒基因序列,分2区(593-799 nt、799-1450 nt和1799-2400 nt)构建进化树,确定青海地区HBV基因重组类型.利用INNO-Li PA HBV分型试剂检测青海地区的C/D基因型重组样品,计算符合率.结果青海地区利用HBV不同区域进化树分型,结果显示各基因型比例为CD1(61.9%)、CD2(8.4%)、C(27.0%)和B(2.8%).基因型分布在藏族和汉族乙型肝炎感染者中的差异具有显著的统计学意义(?2=17.9,P0.01),藏族乙型肝炎感染者中C/D基因型重组和C基因型比例为9 1.5%和8.5%;在汉族中的比例为68.5%和32.3%.C/D基因型重组和C基因型HBV e抗原阴阳性的差异(?2=0.28,P0.05)以及病毒载量的差异(t=1.125,P0.05)均没有统计学意义.C/D重组样品INNO-Li PA分型试剂检测结果为D基因型的理论符合率为87.8%.结论青海地区乙型肝炎感染者以CD1(10-799 nt)基因型重组以主,对其临床意义应进一步的研究分析.该地区的HBV分型检测应注意基因重组对分型试剂的影响. 相似文献
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目的研究乙型肝炎病毒(HBV)e抗原阴性患者HBV核心基因启动子变异方式。方法自5例慢性HBV感染者血清中分别提取HBVDNA,应用多重PCR法鉴定其基因型。PCR扩增X基因序列,克隆入pMD19T载体,共挑选23个克隆测序,与已知HBV基因相应序列比较该患者体内HBV基因变异程度。结果 5例患者中3例基因型分别为B、C和B/C混合型,2例患者通过现行多引物方法无法分型。16个(16/23,69.6%)克隆在X基因下游出现大段缺失突变,其中15例缺失长度为234bp,1例为245bp。缺失突变区域包括C基因启动子区,并导致前-C区起始密码子ATG编码缺失,这种缺失突变在5例患者中均被检出。结论 CP/HBeAg起始密码子缺失突变可能是HBeAg阴性慢性乙型肝炎的一种变异模式。 相似文献
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目的:扩增克隆乙型肝炎病毒(HBV)B,C,D/C及A基因型的全基因组,构建不同基因型的全基因序列克隆.方法:从中国慢性乙型肝炎患者血清提取DNA以L-PCR技术对3.2 kb的HBV DNA进行扩增.纯化后克隆到TA载体.对于HBV不同基因型的全基因序列利用Simplot、DNAStar(Clustal W方法的MegAlign和Phylogenetic tree)等生物软件进行分析.结果:克隆出我国主要存在的几种HBV DNA全基因序列:B、C、D/C和A基因型.包括:adr、adw、ayw血清型.B基因型(adw血清型)全基因序列长度为3215 bp,c基因型(adrq+血清型)为3215 bp,D/C基因型(ayw2血清型)为3215 bp,A基因型(adw血清型)为3182 bp,均递交GenBank.在H3样品中的4个全基因克隆中存在一个同时含有2853-2885 nt缺失突变和核心区的1762A-T、1764G-A双位点突变的变异株,表明同一个体中同时存在突变株和野生株,可能是耐抗病毒药物的分子基础.结论:本研究克隆了我国有报道存在的主要基因型、血清型的HBV全基因序列,为全面了解我国HBV基因型和血清型提供了序列资料. 相似文献
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目的 进行一株新疆维吾尔族D型HBV的全基因克隆.方法应用PCR扩增HBV全基因,进行全基因克隆,挑取阳性克隆进行序列测定,使用BioEdit和美国国立生物技术信息中心(NCBI)-BLAST工具进行生物信息学分析.结果获得1例新疆维吾尔族慢性HBV感染者D型HBV病毒株的全基因序列,其基因组全长为3174 bp,与目前D基因型参照序列比较,同源性为92%~98%,与1株来自中国的DC重组基因型(AY862865)比较,同源性为91%.进一步分析发现,该新疆病毒株与目前D基因型基因库参照序列比较,有9个碱基缺失,位于核苷酸第1760~1768位(ATTAAAGGT),即可能发生了缺失突变.氨基酸分析发现,其血清型为ayw2型.重组位点分析发现,该新疆病毒株未与其他基因型重组.该新疆株与2株来自土耳其的HBV株(AY796032和AY721605)遗传距离最近.结论成功克隆了一例维吾尔族慢性HBV感染者D型HBV病毒株的全基因序列,新疆维吾尔族D基因型HBV病毒株有其自身特点. 相似文献
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乙型肝炎病毒基因型S基因PCR-RFLP分型方法的建立 总被引:57,自引:1,他引:56
目的 建立乙型肝炎病毒(HBV)S基因聚合酶链反应-限制性片段长度多态性(PCR-RFLP)的基因型分型方法并验证其准确性。方法 比较GenBank中HBV223株各基因型全序列的S基因氨基酸及核苷酸序列,设计限制性内切酶BsrⅠ、StyⅠ、DpnⅠ和HpaⅡ鉴别HBVA-F基因型的方法,选择经前SPCR-PFLP的方法已鉴定出的HBVB、C和D基因型标本各30例,用此方法鉴定验证,并随机对HBVB、C和D基因型各3株标本进行S基因测序证实。结果 两种PCR-RFLP及S基因测序鉴定HBV基因型结果一致。结论 S基因PCR-RFLP的方法能够准确鉴定HBV基因型,此方法灵敏性高,特异性强,且酶切图谱简明单一,易于识别,适宜于HBV基因型的流行病学调查。 相似文献
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乙型肝炎病毒S基因限制性片段长度多态性分型方法的建立及应用 总被引:10,自引:1,他引:10
目的 建立乙型肝炎病毒(HBV)S基因片段聚合酶链反应-限制性片段长度多态性(RFLP)的基因分型方法并用于基因型与临床疾病谱关系的研究。 方法 比较GenBank中124株各基因型HBV全序列的S基因核苷酸序列及13株单独S基因序列,设计利用限制性内切酶Mbo Ⅰ、BsTN Ⅰ、BsmA Ⅰ、HpaⅡ酶切S基因片段鉴定HBV基因型(A-F)的方法。对贵州地区176份乙型肝炎患者血清进行基因分型并分析基因型与疾病谱的关系。直接测序2份B型和3份C型毒株的PCR扩增产物,以验证本酶切分型方法的正确性。 结果 测序结果证明本方法能够准确鉴定基因型。在176份标本中,B型100份(56.8%),C型76份(43.2%),未发现其他基因型。B型中无症状携带者(ASC)比例(40.0%)高于C型(15.8%),x2=12.16,P<0.005;慢性中度比例(14.0%)低于C型(31.6%),x2=7.88,P<0,005。 结论 本S基因片段PCR-RFLP基因分型方法简便、准确。贵州地区存在HBV B和C基因型。 相似文献
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目的 对江苏省启东市肝癌高发区的HBV流行株进行基因型别鉴定,并分析其与肝癌的相关性. 方法自启东市肝癌高发区的182份血清标本中抽提HBV DNA,以PCR方法扩增HBV全基因或X基因,测序后与基因库中已知基因型的参照序列共同构建基因进化树,根据序列在进化树中的位置进行型别鉴定.率的比较采用χ 2检验或Fisher精确概率法. 结果对包含24例HBV携带者、11例慢性乙型肝炎患者及13例肝癌患者在内的48例患者的血清HBV全基因测序结果显示,44例(91.7%)为C2基因型,4例(8.3%)为B2基因型,未见A、D、E、F、G、H等型别,也未见B/C重组病毒.以X基因测序方法对182例患者血清标本的鉴定结果显示,C2和B2型分别占92.9%(169例)和5.5%(10例),另有1.6%(3例)为B2/C2混合感染.启东地区C基因型的感染率明显高于邻近上海地区的78.9%(χ2=12.252,P<0.01).在81例肝癌和77例肝炎患者中,B、C基因型HBV的分布差异并无统计学意义(P>0.05).核心启动子区T1762/A1764联合突变在C基因型中的发生率为70.3%,明显高于B基因型中的30.8%(P<0.05);T1766和A1768突变则仅见于C基因型病毒中. 结论启东地区HBV感染以C2基因型为主,但其与肝癌的发生未见明显相关性.C基因型中较易出现核心启动子突变. 相似文献
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Two retroviral vectors carrying an antisense gene from the hepatitis B virus (HBV) preS/S or preC/C were constructed and used to infect the human hepatoblastoma cell line 2.2.15, which expresses HBV surface antigen (HBsAg), HBV e antigen (HBeAg) and releases HBV particles. The results showed that the inhibitory effects of antisense gene transfer, mediated by retroviral vectors on the expression of HBV antigens, appeared as early as day 3 after transduction, reached a maximum on day 5 and persisted for at least 11 days. Our data indicate that, on day 5 after introduction, antisense preS/S inhibited HBsAg and HBeAg expression by 71% and 23%, and the antisense preC/C inhibited HBsAg and HBeAg expression by 23% and 59%. HBV DNA production, in the supernatant of the 2.2.15 cells transduced with either antisense preS/S or preC/C, was also reduced on day 5, but the viability of the 2.2.15 cells was not affected. Our results demonstrate that the replication and expression of HBV can be inhibited through antisense gene transfer mediated by retroviral vectors and that the antisense-preC/C or antisense-preS/S may be potentially useful for clinical gene therapy against HBV. 相似文献
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A trans-activator function is generated by integration of hepatitis B virus preS/S sequences in human hepatocellular carcinoma DNA. 总被引:11,自引:1,他引:10
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W H Caselmann M Meyer A S Kekulé U Lauer P H Hofschneider R Koshy 《Proceedings of the National Academy of Sciences of the United States of America》1990,87(8):2970-2974
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Chauhan R Kazim SN Kumar M Bhattacharjee J Krishnamoorthy N Sarin SK 《World journal of gastroenterology : WJG》2008,14(40):6228-6236
AIM: To confirm the presence of recombination, fulllength hepatitis B virus (HBV) from chronic patients was sequenced and analyzed. METHODS: Full-length HBV genomes from 12 patients were amplified and sequenced in an automated sequencer. Phylogenetic analysis was carried out on full-length, Core and preS2/Surface regions using MEGA software. SimPIot Boot Scanning and amino acid sequence analysis were performed for confirmation of recombination. RESULTS: Eight patients were infected with genotype D strain; one patient with genotype A and three patients had genotype A and D recombination; two of them had cirrhosis and one had hepatocellular carcinoma. Phylogenetic analysis of core and preS2/surface regions separately showed evidence of genotype A and D recombination. The breakpoints of recombination were found to be at the start of preS2 and at the endof surface coding regions. CONCLUSION: We identified and characterized recombinant A and D genotype HBV in hepatitis B surface antigen (HBsAg)-positive patients. 相似文献
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慢性乙型肝炎病毒基因型与BCP区变异的临床研究 总被引:7,自引:0,他引:7
为了研究乙型肝炎病毒 (HBV)基因型与C基因启动子 (BCP)基因变异的关系 ,对 6 9例慢性乙型肝炎患者分别用聚合酶链反应 (PCR)—限制性片段长度多态性 (RFLP)技术和PCR微板核酸分子杂交技术 ,进行基因分型及HBVBCP基因变异检测。在 6 9例慢性乙型肝炎患者中 ,各基因型发生的BCP区A176 2T/G176 4A双突变分别为C型 18例 (4 3 9% )B型 3例 (16 7% )D型 2例 (2 0 % )。C型的BCP双突变率明显高于B型 ,两者相比有显著性差异(P <0 0 5 )。故可以认为乙型肝炎病毒基因型与BCP区双突变存在有一定的相关性。推测A176 2T/G176 4A双突变可能是造成C型患者比B型存在更严重肝损害的原因之一 相似文献
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Masaharu Takeuchi Jiro Fujimoto Hirofumi Niwamoto Yoshihiro Yamamoto Eizo Okamoto 《Digestive diseases and sciences》1997,42(11):2264-2269
Hepatitis B virus is associated with humanhepatocellular carcinoma. We performed polymerase chainreaction for the X, C, S, and preS2/S regions of theviral genome in 23 hepatitis B surface antigen-negative hepatocellular carcinomas and adjacent liver.Hepatitis B viral genomes were detected in 17 of 23tumors and adjacent tissues (73.9%). Among recognizedtransactivators, the X gene was present in 16 (69.6%) cases of hepatocellular carcinoma, but preS2/Swas detected in only 7 (30.4%). Hepatitis B virus C andS regions were detected in 3 (13.0%) and 9 (39.1%)hepatocellular carcinomas, respectively. Serologic study revealed antibodies to hepatitis Bsurface antigen, hepatitis B core antigen, and hepatitisB e antigen in 14 patients; among these, X-gene DNA wasdetected in 12 of 14 tumors (85.7%). The X gene was also detected in 4 of 9 tumors ofseronegative patients. The X gene, present in manyhepatocellular carcinomas, may promote hepatocellularcarcinoma in hepatitis B surface antigen-negativepatients. 相似文献
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Yoshihiko Yano Bui X. Truong Yasushi Seo Hirotaka Kato Akira Miki Yasuhito Tanaka Masashi Mizokami Ayako Kagawa Hiroyuki Miyazaki Masato Kasuga Takeshi Azuma Yoshitake Hayashi 《Hepatology research》2007,37(12):1095-1099
Hepatitis B virus (HBV) is classified into eight genotypes based on complete genome sequence. Each genotype is related to geographic distribution and race. In Japan, most of the genotypes are B and C. In the present study, we report the first Japanese strain of HBV having a recombination between genotypes C and D. A 30-year-old woman was admitted to Kobe Medical Center because of liver dysfunction. She was diagnosed with spontaneous reactivation of chronic hepatitis B. She had no history of blood transfusion and her parents were negative for HBV. The phylogenetic analysis based on the complete genome sequences revealed that this strain was classified into genotype C, whereas the analysis based on Sgene sequence showed that this strain was genotype D. By using a SimPlot program, this strain was confirmed as a recombinant strain between genotypes C and D. Compared with previous recombinant strains in China, the breakpoint was the same and the difference was only 0.8% of the complete genome sequence. It was unclear whether or not this strain was transmitted from China, but the recombinant strains and intergenotypes of HBV have already existed in Japan. 相似文献
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表达HBV preS2S基因重组MVA假病毒颗粒的构建及抗原性鉴定 总被引:2,自引:0,他引:2
目的构建携带HBV preS2S基因的重组MVA假病毒颗粒,评价其抗原性。方法将HBV preS2S基因通过基因重组构建入穿梭载体pSC11,得到质粒pSC11-preS2S,将此质粒转染MVA病毒通过同源重组得到MVA—preS2S,通过免疫印迹法鉴定其抗原性。结果利用基因测序,PCR鉴定证实所得假病毒颗粒可以表达HBV preS2S基因,并且具有良好的抗原性,经过9次传代得到单克隆重组病毒。结论本试验成功地构建了表达HBV preS2S基因的重组MVA假病毒颗粒MVA-preS2S,为基因治疗HBV慢性感染做了一项实验性奠基工作。 相似文献