乙型肝炎病毒基本核心启动子及前C区突变与基因型的关系

目的　探讨乙型肝炎病毒 (HBV)基本核心启动子 (BCP)及前C区突变与基因型之间的关系。方法　随机选取 113例慢性HBV感染者外周血 ,采用INNO LiPA法测定BCPT176 2 A176 4双突变及前C区A1896突变 ,测定HBVS基因序列明确基因型。结果　在C基因型感染者中BCPT176 2 A176 4双突变率明显高于B基因型感染者 ,差异有显著性 (34.2 %∶10 % ,χ2 =6 .74 ,P <0 .0 1) ,而前C区A1896突变率在B基因型和C基因型感染者中差异无显著性 (2 .5 %∶4 .1% ,χ2 =0 .0 0 ,P >0 .0 5 )。结论　与B基因型相比 ,C基因型感染者更易发生BCPT176 2 A176 4双突变。     

重型肝炎时乙型肝炎病毒基因型与基本核心启动子及前C区突变关系的研究

目的探讨重型肝炎（重肝）乙型肝炎病毒（HBV）基因型与基本核心启动子（BCP）及前C区突变的关系。方法采用聚合酶链反应（PCR）-限制性片段长度多态性分析技术（PCR-RFLP）对52例重肝和52例慢性乙肝（CHB）进行HBV基因分型。采用PCR产物直接测序技术，随机对15例B型和15例C型重肝患者的BCP区和前C区进行序列测定，分析HBV基因型与BCPT1762／A1764及前C区A1896突变的关系。结果泉州地区重肝的基因型以B型为主（48．08％），其次为C型（30．77％）和B／C混合型（17．31％），无A、E、F型存在。与CHB组比较，重肝组B型检出率明显降低，而C型和BIC混合型检出率明显升高。C型重肝患者BCPT1762／A1764双突变率显著高于B型（P〈0．05），而前C区A1896突变率在B、C型感染者中差异无统计学意义（P〉0．05）。结论C型感染易引起较重肝损伤，而B／C型混合感染可能是导致重肝发生的重要原因之一。C型重肝患者BCP T1762／A1764双突变率显著高于B型。     

乙型肝炎病毒基因型与国产阿德福韦酯片疗效相关性的研究

目的 探讨我国患者HBV基因B和C型与国产阿德福韦酯片(ADV)疗效间的关系.方法 采用随机、有限双盲、安慰剂(PLB)对照、多中心临床研究方法.筛选合格的HBeAg阳性的慢性乙型肝炎(慢乙肝)患者按照2:1的比例,随机分为试验组(ADV+ADV组:ADV 10 mg/d,治疗48周,疗程48周,共145例)、对照组(PLB+ADV组:PLB 10 mg/d,24周后改用ADV 10 mg/d,治疗24周,共71例).用PCR-RFLP(PCR-限制性片段长度多态性)的方法检测患者血清中乙肝病毒基因型.结果 HBV基因型主要为C型,占66.7%,B型占25.2%.B型多见于广州患者,较C型患者明显年轻(26.98岁 vs 31.71岁,P=0.001),既往接受抗病毒治疗的明显较少.感染B基因型患者48周血清中HBV DNA与基线相比平均下降(-3.86±1.44)log10拷贝/ml(ADV+ADV组)和(-3.63±2.10)log10拷贝/ml(PLB+ADV组),与C基因型患者的(-3.87±1.24)log10拷贝/ml(ADV+ADV组)和(-3.41±1.82)log10拷贝/ml(PLB+ADV组)相比,无明显差别,P＞0.05.B和C基因型患者的血清学应答(HBeAg阴转率和HBeAg/HBeAb血清转换率)间也无统计学意义.结论 感染乙肝病毒B基因型和C基因型的HBeAg阳性慢性乙肝患者,应用国产阿德福韦酯片10 mg/d治疗均可取得显著的抗乙肝病毒复制作用,疗效在不同基因型间差异无统计学意义.     

乙型肝炎病毒核心启动子及前C区突变检测基因芯片的制备及临床应用

目的 建立检测乙型肝炎病毒核心启动子及前C区突变基因芯片并探讨其临床应用的价值.方法 设计并合成针对核心启动子1762/1764、1814及前C区1896位点突变的特异性探针,制备寡核苷酸芯片.采用不对称PCR对该区域进行扩增,扩增产物与芯片杂交后分析结果,并评价该方法的特异性、灵敏度.采用该方法检测138例HBV DNA阳性血清标本.结果 该方法能够特异地检测乙型肝炎病毒核心启动子1762/1764、1814及前C区1896位点,灵敏度达1×101拷贝/μl.138例HBV DNA阳性血清标本中,T1762/A1764突变40例（28.99%）,C1814突变11例（7.97%）,A1896突变16例（11.59%）.前C区A1896突变在高拷贝组中明显高于低拷贝组（P＜0.01）.结论 本研究建立的基因芯片能够准确同时检测HBVV核心启动子区突变和前C区突变,前C区A1896突变可能与HBV复制状态有关.     

乙型肝炎病毒基因型C亚型和核心启动子、前C/核心区基因变异的关系

目的 研究慢性乙型肝炎病毒（HBV）感染者中HBV基因型C亚型（HBV／C）的核心启动子、前C／核心区基因变异情况，分析HBV／C亚型的病毒学特征。方法 用酶联免疫法（ELISA）筛选出79例HBV／C，再用聚合酶链反应．限制性酶长度多态性分析方法（PCR-RFLP）进行HBV／C亚型分析；同时针对HBV核心启动子、DreC／核心区基因进行半巢式PCR及PCR产物直接测序。结果 ①79例HBV／C中，33例（41．8％）为HBV／C1亚型，46例（58．2％）为HBV／C2亚型。②HBV／C1亚型仅见于来自中国南方的患者（P〈0．0001）。③A1898位点变异仅见于HBV／C1亚型（P=0．056），V1753位点变异在HBV／C1亚型中多见（P〈0．05）；HBV／C2以T1858（90％）、A1896（40％）位点变异多见（P〈0．008）。T1762／A1764位点变异在HBV／C两种亚型中均常见。④肝细胞癌（HCC）患者中，V1753和T1762／A1764变异最常见（P〈0．05）。结论 HBV／CI和HBV／C2在中国有明显的地区差异；V1753合并T1762／A1764双变异与发展为HCC相关，尤其在HBV／C1患者。     

乙型肝炎病毒前C区1 896位点突变的PCR分析法

近年研究表明,乙型肝炎病毒（ＨＢＶ）感染者体内,可能存在ＨＢＶ前Ｃ区突变。我们根据ＨＢＶ前Ｃ区ＤＮＡ序列,建立了３′碱基特异性聚合酶链反应（３′ＢＳＰＣＲ）法,检测ＨＢＶ前Ｃ区第１８９６位核苷酸的突变,并检测了７２例ＨＢＶ感染者、１４例甲型肝炎患者的血清ＨＢＶ前Ｃ区１８９６位的突变。病例选择为１９９６年１１月～１９９７年８月期间,在安徽医科大学附属医院传染科就诊的门诊和住院病人。７２例ＨＢＶ感染者中,抗ＨＢｅ阳性７２例,其中慢性乙型肝炎４７例、慢重肝９例、肝炎后肝硬化１６例、另有甲型肝炎患…     

乙型肝炎病毒前S1蛋白抗原检测的临床意义

目的通过分析HBV前S1（PreS1）蛋白抗原和血清标志物之间的关系,探讨PreS1蛋白抗原检测在临床上的应用价值。 方法收集382例HBV患者血清样本,采用ELISA法检测HBVPreS1蛋白抗原和血清标志物,采用荧光定量PCR法检测HBV—DNA,并比较所有检测结果。 结果382例HBV患者血清样本PreS1蛋白抗原和HBV-DNA的检出率分别为33．8％和40．8％;其中HBeAg阳性的144例中,PreS1抗原阳性为112例,HBV-DNA阳性为134例,阳性率分别为77．8％和93．1％;HBeAg阴性的102例中,PreS1抗原阳性为17例,HBV-DNA阳性为22例,阳性率分别为7．1％和9．2％。HBV-DNA检出率稍高于PreS1抗原的检出率,但两者间检出率差异均无统计学意义（P〉0．05）。 结论HBVPreS1蛋白可反映HBV复制的情况,与HBV-DNA结果有较好的一致性,可作为临床上HBV感染诊断或监测的指标。     

山东省人群乙型肝炎病毒＂a＂抗原决定簇突变分析

目的 了解目前山东省社区人群乙型肝炎病毒（HBV）＂a＂抗原决定簇突变率和突变形式,探讨乙肝疫苗（HepB）接种对＂a＂抗原决定簇突变的影响.方法 在全省1～59岁社区人群中通过多阶段随机抽样确定调查对象,通过询问（15岁以上）或查阅接种记录（14岁以下）了解调查对象HepB免疫史;采集血标本,酶联免疫吸附方法 检测血清乙肝表面抗原（HBsAg）,阳性者提取血清DNA,采用巢式PCR方法 扩增HBV S基因,测序后与标准序列进行比较.结果 共对7601人进行调查和血标本采集,得到HBsAg阳性标本239份（3.14%）,可用于HBV DNA提取206份,扩增HBV S基因并成功测序102份.15份血清标本检测到13种HBV＂a＂抗原决定簇突变,突变率为14.70%（15/102）.新生儿普种HepB前、后出生调查对象间,以及有、无HepB免疫史调查对象间＂a＂抗原决定簇突变率差异均无统计学意义.结论 目前山东社区人群中＂a＂抗原决定簇突变率较低且突变位点比较分散;未发现HepB接种对人群＂a＂抗原决定簇突变产生影响.     

乙型肝炎病毒前C区和BCP区突变株复制质粒的构建

目的　构建乙型肝炎病毒 (HBV)前C区和基本核心启动子区 (BCP)突变株的复制质粒。方法　以含 1 2倍拷贝HBVDNA全基因的质粒 (pHBV1 2 )为工具 ,采用分子克隆、PCR定点诱变、限制性酶切长度多态性和序列分析等技术构建目的质粒。转染肝癌细胞株Huh7后 ,测定培养上清HBsAg、HBVDNA来了解病毒抗原的表达和病毒复制。结果　成功构建了 10种HBV全基因前C区 /BCP区突变的表达质粒 ,转染肝癌细胞株 ,获得病毒的表达和病毒颗粒的分泌。其中 ,pUC HBVT176 2、A176 4双突变以及pUC HBVT175 3突变株复制效率较高 ,转染细胞培养上清的HBVDNA为3 16× 10 5拷贝 ml。野生型复制子培养上清HBVDNA含量稍低于BCP突变复制子。结论　获得 10株含有不同前C区和BCP区突变的HBV全基因复制质粒 ,为体外进一步研究上述变异的生物学意义提供基本模型。     

河北正定县某镇人群乙型肝炎病毒"α"抗原决定簇突变初步分析

目的 了解河北正定县某镇自然人群感染乙型肝炎病毒(HBV)的"α"抗原决定簇突变情况,分析新生儿乙肝疫苗计划免疫对其的影响.方法 为正定县某镇全体村民整群采集静脉血标本,用放射免疫法检测血清HBV-M,选取乙肝表面抗原(HBsAg)阳性者提取血清HBV DNA,扩增S基因,测序后与标准序列对比.结果 11 478份血清样品中,HBsAg检测阳性率4.78%.其中可用于HBV DNA提取的样品443份,扩增HBV S基因并成功测序434例.检测到该地区HBV"α"抗原决定簇氨基酸突变率6.7%,1986年前后出生人群HBV"α"抗原决定簇两个茎环的氨基酸突变率差异无统计学意义(P=1.0000,P=0.0877).结论 该镇存在HBV"α"抗原决定簇突变株,其在乙肝疫苗免疫出生后人群中感染率较低,尚不能说明该镇新生儿乙肝疫苗接种会引发突变株的流行.     

Prevalence of HBV core promoter/precore/core mutations in Gambian chronic carriers.

One hundred forty-two precore/core sequences were obtained from Gambian chronic hepatitis B virus (HBV) carriers and the predominant variants defined. The two point mutations, from A to T and G to A at nt positions 1762 and 1764 in the basic core promoter region, were found in only 7/99 (7%) of the samples where this region was sequenced. These mutations were found in both HBeAg-positive and -negative patients. The precore stop-codon mutation at nt position 1896 was found in 14/51 (27%) of HBeAg-negative samples, which is a lower prevalence rate in comparison with other parts of the world with high carrier rates. In HBeAg-positive patients the core amino acid sequences were conserved, but after seroconversion to anti-HBe significantly more changes were apparent. Several of the amino acid substitutions found have been described previously been in wild-type viruses of other genotypes.     

Hepatitis B e antigen‐negative mutations in the precore and core promoter regions in Korean patients

Most patients with hepatitis B e antigen (HBeAg)‐negative chronic hepatitis B have variants of the hepatitis B virus (HBV) that include mutations in the precore or core promoter regions of the HBV genome. The aim of this study was to investigate the patterns of precore and core promoter mutations and their relationship to HBeAg expression in Korean patients. Four hundred seventy‐five Korean patients with chronic HBV infection between February 1995 and December 2003 were enrolled in this study. There were 236 HBeAg‐positive and 239 HBeAg‐negative patients. Blood samples were tested for HBsAg, anti‐HBs, HBeAg, hepatitis B e antibody (anti‐HBe), liver function tests, and serum HBV DNA. Mutations in the precore and core promoter regions were determined by direct sequencing. In the core promoter region, the C1740, C1753, T1762/A1764, and T1766 mutations were associated with HBeAg escape (all; P < 0.05). In the precore region, a higher frequency of the C1802, A1828, T1846, A1850, C1858, T1862, and A1896 mutations was found in HBeAg‐negative patients (all; P < 0.05). In particular, the A1896 mutation was associated with high serum levels of ALT and HBV DNA in HBeAg‐negative patients (P = 0.014 and 0.026, respectively). Mutations around the Kozak sequence (nucleotides 1809–1812) were found in 6.7% of patients and were not associated with undetectable HBeAg (P = 0.13). In Korean patients, various mutations in the precore and core promoter regions were associated with HBeAg escape and amelioration of hepatic inflammation in HBeAg‐ negative patients. Only the A1896 mutation contributed to HBeAg‐negative chronic hepatitis B. J. Med. Virol. 81:594–601, 2009 © 2009 Wiley‐Liss, Inc.     

HBV基因型与HBV感染慢性化、重症化的关系

目的 探讨HBV基因型与HBV感染后慢性化、重症化的关系.方法 应用型特异性引物聚合酶链反应法,对中国2922例HBV感染者进行HBV基因型检测,比较各临床类型HBV感染者基因型分布差异及各基因型HBV感染者肝功能和病毒学差异.结果 2922例HBV感染者中,基因型B、C、B/C、D分别占15.9%、83.5%、0.41%、0.21%.与慢性肝炎比较急性肝炎B基因型所占比例较高（P=0.003）,肝硬化和肝细胞性肝癌C基因型所占比例较高（P值均为0.000）,慢加急性肝衰竭与慢性肝炎比较基因型分布差异无统计学意义.急性肝炎和慢性肝炎B、C基因型患者HBeAg 阳性率、HBV DNA病毒载量、肝功能生化指标差异无统计学意义.慢加急性肝衰竭、肝硬化、肝细胞性肝癌组C基因型较B基因型患者HBeAg阳性率更高（P值分别为0.000、0.024、0.003）,肝细胞性肝癌C基因型患者HBV DNA病毒载量高于B基因型患者（P=0.025）,慢加急性肝衰竭和肝细胞性肝癌组C基因型较B基因型患者胆碱酯酶更低（P值为0.0004、0.02）.结论 中国HBV感染者的HBV基因型以B、C基因型为主,少量的B/C、D基因型;C基因型较B基因型HBV感染者更易发生慢性化和进展为肝硬化和肝细胞性肝癌,但未观察到基因型对慢加急性肝衰竭发生的差异.急性和轻症HBV感染者B、C基因型未显示对病情的明显影响,但重症和终末期HBV感染者C基因型较B基因型患者HBeAg阳性率、HBV DNA病毒载量更高,肝功能损害更严重.     

Frequency and clinical significance of core promoter and precore region mutations in Tunisian patients infected chronically with hepatitis B

Genetic variability of hepatitis B virus (HBV) in the C gene and its association with the different stages of chronic liver disease has been studied inadequately with controversial results. The objectives of the current study were to determine the frequency of core promoter and precore mutations in chronic hepatitis B in Tunisia and to evaluate their impact on viral replication and disease progression. Sequencing was performed in upstream regulatory sequence (URS), pre‐core (PreC) and basal core promoter (BCP) regions for 123 chronic infected patients by HBV genotype D at different status of disease. Mutations were detected in 98.4% of cases, affecting URS, BCP and Pre‐C in 95.1%, 95.9% and 87.8% respectively. Multi‐mutations increased significantly from asymptomatic carrier to advanced liver disease status. G1896A (74.8%), G1764A/T/C (71.5%), G1899A (54.4%) and T1678C (52%) were the most common. Special attention should be paid to A1703T, T1678C/G‐A1703T, and A1652G‐A1679G mutations probably specific of Tunisians sequences; they were observed in 40.6%, 41.5% and 30.1% respectively. A1679G/C, T1753C/G/A, A1762T/G and A1762T‐G1764A were more prevalent in older patients. High DNA levels were associated with G1899A or G1764T/C‐C1766G‐C1799G and advanced liver disease with mutations at positions 1762, 1764 and/or 1899 alone or in double or triple mutations. It was also shown that substitutions at nucleotides 1762, 1764 and 1899 have an impact on the disease progression. It is the first report for specific mutations in the URS region for genotype D. It should be completed by studying eventual correlation with clinical progression and the response to treatment. J. Med. Virol. 84:1719–1726, 2012. © 2012 Wiley Periodicals, Inc.     

HBV PC区和BCP区变异与Peg-IFNα-2b疗效的关系及治疗前后的变化

目的:探讨乙型肝炎病毒(HBV)前核心区(PC区)G1896A和基本核心启动子区(BCP区)A1762T/G1764A突变与聚乙二醇化干扰素α-2b(Peg-IFNα-2b)治疗应答的关系及相关变异在治疗前后的变化。方法:69例HBeAg阳性慢性乙型肝炎(CHB)患者,接受48周Peg-IFNα-2b治疗并随访24周。PCR扩增每位患者第0周和第72周HBV PC和BCP区并测序分析突变情况,同时监测患者第0、4、8、12、24、36、48、60和72周HBsAg、HBeAg、丙氨酸转氨酶(ALT)和HBV的DNA水平。结果:共14例患者检测为野生型(20.29%),55例患者检测为突变型(79.71%)。野生型较突变型HBeAg基线水平更高(P=0.024)。患者基线和72周时野生型、PC突变型、BCP突变型和PC+BCP突变型所占构成比发生明显变化(P=0.004)。野生型、PC突变型、BCP突变型和PC+BCP突变型患者在72周的HBeAg血清转换率和联合应答率差异均无统计学意义。结论:PC区和BCP区突变对HBeAg阳性B/C基因型CHB患者Peg-IFNα-2b治疗应答无明显影响,但治疗前后各突变所占构成比发生明显变化。     

HBV基因型与HBV感染慢性化、重症化的关系

Objective To explore the association between HBV genotype and chronic/severe liver disease with HBV infection in Chinese patients.Methods Serum samples were collected from 2922 patients with HBV infection.HBV genotyping was performed with type-specific primers polymerase chain reaction,and the virological and biochemical markers were detected,which differences in the genotypes between various clinical types of HBV infection and liver function and virological markers between various HBV genotyping were analyzed.Results The genotype B,C,BC combinations,D of 2922 patients with HBV infection accounted for 15.9%,83.5%,0.41%,0.21% respectively.The ratio of genotype B in acute hepatitis group was higher(P=0.003),which the ratio of genotype C in the cirrhosis group and the hepatocellular carcinoma group was higher(P=0.000,0.000).The difference in ratio of genotype C was not statistically significant between acute-on-chronic liver failure group and chronic hepatitis group.HBeAg-positive rate,viral load and liver function markers of B,C genotype group in acute hepatitis group and chronic hepatitis group were not significant different.HBeAg-positive rates of genotype C in acute-on-chronic liver failure group,cirrhosis group,hepatoeellular carcinoma group were higher than that of genotype B(P=0.000,0.024,0.003).Viral load of genotype C in hepatocellular carcinoma group was higher than that of genotype B(P=0.025).Cholinesterase levels of genotype C in the acute-on-chronic liver failure group and the hepatocellular carcinoma group was lower than that of genotype B(P=0.0004、0.02).Conclusion There were HBV genotype B,C,B/C combinations and D in Chinese patients with HBV infection,with genotype B and C being the major ones.Compared with HBV genotype B,genotype C in Chinese patients with HBV infection was more likely to chronic infection,evolved to cirrhosis and hepatocellular carcinoma, but genotype difference was not observed in occurrence of acute-on-chronic liver failure.Genotype was not significant effect in acute and chronic hepatitis B,but HBeAg-positive rate/viral load was higher and liver damage was more severe in severe and end-stage genotype C HBV infection patients.     

HBV基因型与HBV感染慢性化、重症化的关系

Objective To explore the association between HBV genotype and chronic/severe liver disease with HBV infection in Chinese patients.Methods Serum samples were collected from 2922 patients with HBV infection.HBV genotyping was performed with type-specific primers polymerase chain reaction,and the virological and biochemical markers were detected,which differences in the genotypes between various clinical types of HBV infection and liver function and virological markers between various HBV genotyping were analyzed.Results The genotype B,C,BC combinations,D of 2922 patients with HBV infection accounted for 15.9%,83.5%,0.41%,0.21% respectively.The ratio of genotype B in acute hepatitis group was higher(P=0.003),which the ratio of genotype C in the cirrhosis group and the hepatocellular carcinoma group was higher(P=0.000,0.000).The difference in ratio of genotype C was not statistically significant between acute-on-chronic liver failure group and chronic hepatitis group.HBeAg-positive rate,viral load and liver function markers of B,C genotype group in acute hepatitis group and chronic hepatitis group were not significant different.HBeAg-positive rates of genotype C in acute-on-chronic liver failure group,cirrhosis group,hepatoeellular carcinoma group were higher than that of genotype B(P=0.000,0.024,0.003).Viral load of genotype C in hepatocellular carcinoma group was higher than that of genotype B(P=0.025).Cholinesterase levels of genotype C in the acute-on-chronic liver failure group and the hepatocellular carcinoma group was lower than that of genotype B(P=0.0004、0.02).Conclusion There were HBV genotype B,C,B/C combinations and D in Chinese patients with HBV infection,with genotype B and C being the major ones.Compared with HBV genotype B,genotype C in Chinese patients with HBV infection was more likely to chronic infection,evolved to cirrhosis and hepatocellular carcinoma, but genotype difference was not observed in occurrence of acute-on-chronic liver failure.Genotype was not significant effect in acute and chronic hepatitis B,but HBeAg-positive rate/viral load was higher and liver damage was more severe in severe and end-stage genotype C HBV infection patients.