Study On Molecular Virology And Epidemiology Of Sen-Virus

To date, five hepatitis viruses from HAV to HEV which accounts for eighty to ninety percent hepatitis cases had been discovered .But we couldn't find any hepatitis viruses in the left ten to twenty percent cases which had typical virual hepatitis manifestation. Because of the discovery of HCV in 1989 and HEV in 1990, it was believed that all virual hepatitis had their own pathogens. So researchers had been working hard to look for the pathogens behind the no causes hepatitis cases. In July 1999, a new family of DNA viruses was isolated and was designated as SEN virus (SENV) after the initials of the infected patient, a human immunodeficiency virus-infected injection drug user, by a researcher group leading by Dr. Priini in Diasorin Research Centre. Now we had a little to know about its etiology. In this project we wanted to study on the molecular biology characteristic and epidemiology of two subtype SENV-D and SENV-H which were related to transfusion-associated non-A to E hepatitis, and hope to obtain some useful information on its etiology.1. The gene cloning and sequence analysis of SENV-D and SENV-H isolated from ChinaAccording to the published nucleotide sequences of SEN viruses, specific primers were designed and synthesized. From the serum of two Chinese patients with non-A-E hepatitis, one SENV-D isolate named SENV-D-BJ1 spanning the complete coding region was amplified by semi-nested PCR, another isolate named SENV-D-BJ2 spanning the partial coding region (including ORF1 and ORF2) was amplified too. From one blood donor serum, two SENV-H isolates named SENV-H12-1 and SENV-H 12-2 spanning the complete coding region were amplified by nested PCR respectively. Analyzing gene organization, these two SENV-D Chinese isolates were analogous to that of the reported SENV-D (AX025730) isolate, two SENV-H Chinese isolates were a little different from the reported SENV-H (AX025838) isolate. The ORF1 protein of SENV-H12-1 isolate was consisted of 675 amino acids and 92 amino-terminal amino acids less than SENV-H (AX025838), it had insertion in two sites also. The ORF1 protein of SENV-H 12-2 isolate was 7 amino acids less than SENV-H (AX025838) because of deletion. Because the first start code was mutated, the ORF2 protein of SENV-H 12-2 isolate started from the second ATG code, so that it was 7 amino acids less than SENV-H (AX025838) from its N-terminus. The ORF1 proteins of all four Chinese SENV isolates had two motifs (FTL and YXXK) which were conserved in replication-associated proteins. In all four Chinese SENV isolates, the putative non-structure ORF2 protein had a CAV-like region. The putative ORF3 protein had a serine-rich tract preceded by a cluster of basic amino acids(R and K).Database scaning suggested this region had high a percentage of homology with DNA topoisomerase I protein of D.melanogaster, and might play an important role in the replication of such single-stranded DNA viruses.2. Establishment and comparison of polymerase chain reaction for detecting SENV-D and SENV-H and prevalence of SEN virus in ChinaAccording to the published articles and gene sequences, two nested-PCR methods for SENV-D and SENV-H were built. We had estimated the specificity and sensitivity of two nested oligonucleotide primer pairs. The sensitivity of these primer sets was lOOcopies. In order to investigate the prevalence of SENV-D and SENV-H in China, some kinds of hepatitis patients serum and blood donors serum were collected and tested. The prevalence rate of SENV-D was equivalent to that of SENV-H (26% vs. 24%). We detected SENV-D and /or SENV-H DNA in 31% of blood donors which was higher than that of America, Italy and Thailand, and equivalent to 15% to 20% of Japan, Taiwan and Germany. The prevalence of SENV-D/H viremia was significantly greater among patients with hepatitis B and hepatitis C than among blood donors and patients with hepatitis A(59%-85% vs. 31%-36%, P<0.05).This result coincided with that in Japan and Taiwan.We also found that the prevalence among patients with non-A-E hepatitis was sign...