乙型肝炎肝硬化患者与乙型肝炎病毒携带者乙型肝炎病毒X基因序列比较研究

Comparison of X gene Sequences of Hepatitis B Virus Isolates in Hepatitis B Induced Liver Cirrhosis Patients and HBV Carriers

目的研究乙型肝炎肝硬化患者与乙型肝炎病毒（HBV）携带者HBV X基因序列的差异。方法对20例标本的PCR产物进行直接测序;并对3例乙型肝炎肝硬化患者和3例无症状HBV携带者HBV DNA的扩增片段进行克隆测序,并分析比较。结果肝硬化患者中X基因核心启动子双突变T1762/A1764、G1719T、T1727G/A、G1730C、T1753C等变异高于HBV携带者;前者X启动子区变异率明显高于后者;乙型肝炎肝硬化同一患者X区不同克隆之间的碱基序列同源性为91.3%-99.7%,而HBV携带者为96.0%-100.0%。结论乙型肝炎肝硬化患者,HBV DNA X区的CP启动子区以及X启动子区变异程度明显高于HBV携带者,同时乙型肝炎肝硬化患者体内存在变异程度更大的HBV准种群。

OBJECTIVE To discover diversity of X gene sequences of hepatitis B virus isolates in hepatitis B induced liver cirrhosis patients and HBV carriers. METHODS DNA fragment including X gene sequences of hepatitis B virus was amplified, sequencing PCR products was preformed. The PCR products of three liver cirrhosis patients and three HBV symptomless carriers were cloned into pGEMT Easy vectors. Positive clones with target sequences were selected out for sequencing. Sequence comparison was made to find the identity. RESULTS A comparison of T1762/A1764, G1719T, T1727G/A, G1730C and T1753C mutations in a core promoter between the liver cirrhosis patients and the HBV carriers showed that the HBV isolates from the former had higher frequencies of mutation than the latter. The X promoter region of the HBV isolates from the liver cirrhosis patients showed higher frequencies of mutation than the isolates from the HBV carriers. Additionally, the homology between clones of X gene from one individual with liver cirrhosis averages 91.3-99.7%, that of HBV carriers averages 96-100 %. CONCLUSIONS The core and X promoter region of the HBV isolates from the liver cirrhosis patients show the higher frequencies of mutation than the isolates from the HBV carriers. There are HBV quasispecies which possess great variation in the liver cirrhosis patients.