| Hepatitis B virus(HBV) CD recombinant was first isolated from Tibet samples in 2002.The HBV/CD recombinant sequence can be classified into genotype C by C gene,while into genotype D by preS/S gene.Then,in the study of the HBV subgenotype in China,we found the existence of CD recombinant in Gansu and Tsinghai provinces.And it can be divided into two forms regarding the size of the genome affected by means of recombination.One of them has a recombination with genotype D over the preS/S region(nt10—799) and another over a large part of the P gene overlapping with the S and X genes(nt 10-1499).But the distribution and characteristics of HBV/CD recombinant are still unclear.Our study aimed at determining the distribution and characteristics of HBV/CD recombinant.To investigate the geographic and ethnic distribution of HBV/CD recombinant, A cohort of 1000 patients with chronic HBV infection,coming from five provinces of northwest China,were analyzed by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP) or nucleotide sequencing.Result of this study showed that CD recombinant was the predominant HBV strain in Tibet and Tsinghai province(64%and 55%respectively),while genotype C was the most prevalent genotype in Gansu and Ningxia provinces(58%and 70%),and CD recombinant was the second prevalent strain(27%and 15%) in these two provinces.Genotype D and C (46%and 43%) were the predominant genotype among uygur in Xinjiang Autonomous Region,while HBV/CD recombinant showed the lowest prevalence in Xinjiang(4%).Comparison of the HBV/CD recombinant prevalence in different ethnics showed that,HBV/CD was the predominant HBV strains in Tibetans(65%), and it was commonly found in Hui nationality(30%) and Han nationality(21%) of northwest china,while rarely found in uygur in Xinjiang Autonomous Region(4%).In the same region,the prevalence of HBV/CD possessed the highest percentage among tibetan in Tibet(79%)and Tsinghai province(67%),HBV/CD was also the most prevalent strains in Hui nationality of Gansu province(61%),while the prevalence was low(15%) in Hui nationality of Ningxia province,we also compared the prevalence of HBV/CD1 with HBV/CD2,founding that different district had different prevalence. The results suggested that C/D recombinants had special ethical and geographical distribution.To investigate the clinical and virological characteristics of HBV/CD recombinants,507 patients with chronic HBV infection from five clinical centers were enrolled in the case control study,.With matched age and gender,no significant differences were found on ALT levels among genotype B,C,D and CD recombinant,while TBiL levels in genotype B(46.3±71.9) were higher than another genotypes(F=3.555,P=0.014),with the HBV/CD recombinant(19.3±28.7)had the lowest TBIL levels.HBeAg status also had significant difference between different genotypes(x~2=30.422,P=0.000).There are more HBeAg positive cases than negative cases in HBV/CD recombinants,while in other genotypes all have more negative cases than positive ones.In the comparison of pre-C and C region mutation rate between HBV/C and HBV/CD,HBV/C was found associated with the highest tendency to develop PC mutation(P=0.003).no significant differences were found on other pre-C and C region mutation rate.The results suggested HBV/CD recombinant had different clinical and virology characteristic from genotype B,C and D.Maybe lower PC mutation rates responsible for the lower HBeAg negative infection of HBV/CD recombinant.Transmission of hepatitis B virus(HBV) in china is thought to be primarily vertical.To find the clues for the transmission pattem of HBV/CD and evidence for recombination,we studied three families with multiple members chronically infected with mixed HBV genotypes and one family with three generations infected with HBV/CD recombinant.The whole genome or S-region of the virus was amplified from serum derived HBV-DNA,and the sequences were subjected to phylogenetic tree analysis.In addition,divergence and mutation rate of HBV/CD recombinant were calculated.The phylogenetic tree analysis provided evidence of intra-familial transmission of the virus and we found that there is no significant genetic distance between C2 and CD1,at the same time,a gradually process from C2 to CD1 can be observed when we were constructing a phylogenetic tree used nt61-1200 with many quasispecies from a mixed genotype infected isolation,the divergence rate showed that the mixed genotype infection of genotype C2 and CD1 more related to diverse in same genotype and less to diverse between genotype C2 and CD1.This results may suggested that HBV/CD recombinant may not the product of recombination of genotype C and D,it may be some divergence quasispecies of C2 and was selected as a major strain in special host.Finally,to measure the differences of replication efficiency and antigen expression of HBV genotype C,D and CD recombinant.We constructed 1.3 copies of HBV genome of these three genotypes(2 strains HBV/C2,2 strains HBV/D and 3 strains HBV/CD),and then cloned them into vector PUC19.After 1.3 copies HBV genome were confirmed with sequencing,Recombinant plasmids of HBV/C,HBV/D,HBV/CD were infected with liposome to cells of HepG2.Then,HBeAg and HBsAg titer were detected in cells medium and cells after 72 hours of transfection. All the 1.3 copies of HBV can replicate and translate themselves in HepG2,using their own regulatory elements to produce HBsAg and HBeAg.But the replication efficiency should be determined in further study. |
