特异性引物聚合酶链反应乙型肝炎病毒基因分型法的建立及应用  

Establishment and application of a type-specific polymerase chain reaction(PCR) for HBV genotyping

作　　者：李雅娟[1] 庄辉[1] 李杰[1] 董庆鸣[2] 牛俊奇[3] 马为民[4] 赵伟[5] 王磊[6] 赵保安 陈雅洁 钟金群 

机构地区：[1]北京大学医学部病原生物学系,100083 [2]北京地坛医院 [3]吉林大学第一附属医院 [4]北京大学深圳医院 [5]南京市第二医院 [6]济南传染病医院 [7]湖北汉川疾病预防控制中心 [8]长春北方肝胆医院 [9]广东省清远市疾病预防控制中心

出　　处：《中华微生物学和免疫学杂志》2006年第9期859-862,共4页Chinese Journal of Microbiology and Immunology

基　　金：国家"十五"攻关课题(2004BA718802);北京市科委课题(No.H020920020190)

摘　　要：目的应用型特异性引物聚合酶链反应法(PCR)进行乙型肝炎病毒基因分型并分析该法的可靠性。方法应用型特异性引物PCR和INNO-LiPA分别对深圳、长春、北京152份HBV DNA阳性慢性乙型肝炎患者的血清标本进行了基因型分型,对该两种分型法不一致的血清标本再进行S区基因测序分型,以确定该两法的可靠性。结果型特异性引物PCR和INNO-LiPA的总符合率为86．8％(132／152),不一致率为13．2％(20／152)。型特异性引物PCR检测到81份(53．3％)B型;58份(38．2％)C型;13份(8．5％)B+C型混合感染,未检出其他基因型或混合感染的基因型。INNO-LiPA检测到74份(48．7％)B型;61份(40．1％)C型;5份(3．3％)B+C型混合感染;另检出3份(2．0％)A +B型混合感染;1份(0．7％)B+E型混合感染;1份(0．7％)C型与D型,1份(0．7％)D型感染,3份(2．0％)B／C／D型及3份未能分型。20份两法分型不一致的标本中,6份无剩余血清,对其余14份进行了S区基因测序分型,结果型特异性引物PCR与S区基因测序分型法的符合率为71．4％(10/14),而INNO-LiPA与S区测序法的符合率仅为7．1％(1／14),前者明显高于后者(P<0．05)。结论型特异性引物PCR和INNO-LiPA均可鉴定HBV基因型,但前者较为简便和可靠,且费用较低,可用于临床标本的检测和流行病学调查。Objective To determine the accuracy of a type-specific primer polymerase chain reaction （PCR） for identifying HBV genotypes. Methods A type-specific primer PCR and INNO-LiPA assay were used to identify HBV genotype of 152 serum samples of patients with chronic hepatitis B from Shenzhen, Changchun and Beijing. Fourteen of 20 serum samples with discrepant genotype were analyzed by the sequencing of HBV S gene to determine their accuracy. Results The total coincidence rate between the type-specific primer PCR and INNO-LiPA was 86.8% （132/152）, with the discrepancy of 13.2% （20/152）. Of 152 samples from patients with HBV infection determined by the type-specific PCR, the proportion of genotype B, C, and B ＋ C were 53.3% （81/152）, 38.2% （58/152） and 8.5% （13/152）, respectively. The figures for INNO-LiPA were 48.7% （74/152）, 40.1% （61/152） and 3.3%（5/152）, respectively. In addition, 3（2.0%） genotype A ＋ B, 1（0.7%） B ＋ E, 1（0.7%） C ＋ D, 1（0.7%） D, 3（2.0%） B ＋ C ＋ D, and 3（2.0%） were unidentified by INNO-LiPA. Of 20 sera with discrepant genotype results by the type-specific primer PCR and INNO-LiPA, 6 had no serum left, and the remaining 14 samples were further analyzed by the sequencing of HBV S gene for genotyping. The total coincidence rate between the type-specific primer PCR and the sequencing of HBV S gene was 71.4% （10/14）, which was significantly higher than that of INNO-LiPA（7.1%, 1/14）, P〈0.05. Conclusion Type-specific primer PCR and INNO-LiPA can be used for HBV genotyping. However, the type-specific PCR is more simple and accurate, with low cost. It may be used for test of clinical specimens and for epidemiological investigations.

关 键 词：乙型肝炎病毒 基因型 型特异性引物PCR INNO-LiPA 

分 类 号：R450[医药卫生—治疗学]