|Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1995: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1994: ¥1,000,000 (Direct Cost: ¥1,000,000)
We analyzed nucleic and amino acid sequences of the hypervariable region (HVR) of the hepatitis C virus (HCV) in the 4 kinds of sera, for the study of the next step of the titration experiments using chimpanzees as follows. One ml each of 10^<-5>,10^<-3> and 10^<-2> dilutions of human serum #4 were subsequently inoculated to chimpanzees nos.364,346, and 345, respectively, and one ml of 10^<-4>,10^<-5>, and 10^<-6> dilutions of human serum #6 were subsequently inoculated to chimpanzee nos.353,215, and 340, respectively. Only chimpanzee nos.353 and 215 were positive for HCV RNA in the sera, while both of inocula #4 and #6 contained high titer of HCV RNA (2x10^6 copies/ml, each, by semi-quantitative competitive assay). RNA extraction, transcription to cDNA,and amplification were done, 20 colonies were picked up, and nucleic and amino acid sequences were determined. Genetic variations were limited mostly to the HVR region in clones from #4 and #6. There were 6 and 4 clones from #4 and #6, respectively, and clones from #4 were different from those from #6 in amino acid sequences (table 1 & 2). On the other hand, clones of chimpanzee nos.353 and 215 were completely same in amino acid sequences. Homology of all clones from both chimpanzees was identical to the major type (6a-1) of human inoculum #6. The results of titration experiment suggested that HCV RNA titer in sera was not always correlate with in vivo infectivity, and the analytical study of amino acid sequences suggested that major group clone in the diluted inocula were mainly transmitted and replicated.