Distribution Of Hbv Genotypes And Subgenotypes Among Lc And Hcc Patients In Northern China

Background and Objective Hepatitis B virus(HBV) is a significant public health problem that may lead to chronic hepatitis,cirrhosis and hepatocellular carcinoma(HCC).Approximately 350 to 400 million(5-6%) people in the world are persistent carders.And China is the middle/high epidemic area.The serological investigation in 2006 suggested that there are 7.18%Chinese people are HBV carriers,and there are nearly 30,000,000 chronic hepatitis patients in China.At present,according to the molecular evolutionary analysis of genomic DNA sequence, HBV strains isolated in various countries are classified into eight main genotypes, designated A-H,and arbitrarily defined by an inter-group sequence divergence of more than 8%based on complete genomes or 4%based on S gene.There are many subgenotypes including Aa,Ae,Ba,Bj,C1,C2.The recent researches indicated that the genotypes and subgenotypes are associated with the mode of transmission, progression and prognosis of disease and the effect of the antiviral therapy.Our aim is to investigate the distribution of hepatitis B virus(HBV) genotypes and subgenotypes in northern China,and determine the relationship between the subgenotypes of hepatitis B virus and serious hepatitis B.HBV Genotypes and subgenotypes of 101 cirrhosis and 178 hepatocellular carcinoma cases were determined with nested PCR with type specificity primers,and the relevance between hepatitis B virus subgenotypes and serious hepatitis B was statistically analyzed.Combining with the clinical data we analyse the relationship between the genotypes/subgenotypes and the final stage liver disease patients to identify the differences of the injuries resulted by different genotypes/subgenotypes HBV.Methods1.The 279 specimen from northern China were collected between 2007 and 2008. There were 155 fresh tissue specimen and 124 serum specimen.2.DNA extracting Extract the DNA of the blood serum and tissues by phenol and chloroform.3.Determine the HBV DNA's genotypes and subgenotypes with nested PCR.4.Do electrophoresis with 2%agarose gel and use 2000DL as the marker to compare the molecular weight.5.Observe the gel image,judge the genotypes/subgenotypes of the products by the length of the fragment.(Some of them are sequenced(pre-S/S and X) to make sure the genotypes/subgenotypes.)6.Data Analysis Use SPSS 13.0.Do the analysis with t test,x~2 test, non-parametric test.The results will make sense when P<0.05.Results1.We can determine the HBV DNA's genotypes/subgenotypes clearly with the length of the fragments of the products after the electrophoresis.And the results of the sequencing of preS/S and X were exactly coincident with the results of the nested PCR.So the nested PCR were reliable because of its accuratissime and specificity.2.Of the 101 cirrohsis cases,1 was Ba subgenotypes(0.99%),94 were C2 subgenotypes(93.07%),and 6 are Ba and C2 subgenotypes co-infection (5.94%).And of the 178 hepatocellular carcinoma cases,6 were Ba subgenotypes(3.37%),159 are C2 subgenotypes(89.33%) and 13 were Ba and C2 subgenotypes co-infection(7.30%).No other genotypes or subgenotypes were found in both of them.And no difference between Ba and C2 subgenotypes was found between cirrhosis and HCC cases(P>0.05).3.Difference between gender was found(P<0.05),the percent of the male patients in HCC was higher than the percent in cirrohsis,the percent of the male patient was higher than the female in each subgenotype,and difference between gender of the C2 subgenotype was found(P<0.05).No significant difference between age was found(P>0.05) of the patients with different genotypes or subgenotypes with Kruskal-Walls test.4.There were no significant difference between HBeAg positive rate of the patients with different genotypes or subgenotypes(P>0.05).But the HBeAg positive rate of the HCC patients was apperently lower than the cirrohsis patients.5.There were difference between TBIL level of the cirrohsis and HCC patient (P<0.05).There were no difference between TBIL level of the patients with different genotypes or subgenotypes(P>0.05).There were no difference between ALT level of the cirrohsis and HCC patient(P>0.05).There were no difference between TBIL level of the patients with different genotypes or subgenotypes(P>0.05).6.We detected the subgenotypes of the tumor and paratumor tissue were different in 3 matched-pairs.And the tumor were C2 subgenotypes and the paratumor were Ba and C2 subgenotypes co-infection.Conclusions1.Subgenotype C2 is dominant in northern China,and then is subgenotype Ba.2.No significant distribution difference of patients with cirrhosis and HCC infected with different HBV genotypes and/or subgenotypes is found.But the cirrohsis and HCC patients with C genotype are more than chronic hepatitis B patients.So the condition of patient with C genotype are serious and will develop rapidly.And this indicate that the HBV genotypes are associated with the development and prognosis of the disease.3.The gender has no relationship with the development of the cirrohsis and HCC. 4.The difference between the tumor and paratumor tissue may indicate that HCC may be associated with the unicell clone.