Four subtypes of hepatitis C virus (HCV), Pt(I), K1(II), K2a(III) and K2b(IV), have been suggested based on the nucleotide sequences of the non-structural (NS) 5 region. A fifth subtype from Japanese patients, Tr(V), which shows a less than 68% homology in nucleotide sequence when compared with other subtypes has been identified. A one-step method which enables a quick determination of subtype using polymerase chain reaction with a mixed primer set deduced from the sequence of each subtype has been developed. Using this technique, the subtypes of 418 out of 478 Japanese patients (87.4%) were determined. The incidence of each subtype in Japan was as follows: K1(II), 307 (73.4%); K2a(III), 74 (17.7%); K2b(IV), 28 (6.7%); and Tr(V), 3 (0.7%). This one-step subtyping technique should be useful for studying the epidemiology or biology of the HCV.
To determine how various factors influence the response to interferon (IFN) therapy, we retrospectively studied 157 consecutive Japanese patients with chronic hepatitis C who received various treatment schedules of IFN. They were divided into two groups on the bases of outcome. One group was comprised of 65 patients who achieved a sustained normalization of serum alanine aminotransferase (ALT) levels for at least 6 months after treatment, while the other group was comprised of 84 patients with persistent elevation of serum ALT levels, despite treatment. Genotyping of hepatitis C virus (HCV) was done by polymerase chain reaction (PCR) with genotype specific primers, analysing the variations in nucleotide sequence within the NS 5 region of the HCV genome, namely genotypes PT, K1, K2a and K2b. We then used a multivariate analysis to determine the factors related to mode of treatment, patient characteristics and HCV genotype in relation to the response to IFN therapy. Of the 16 factors analysed, the HCV genotype (genotype K2a or K2b, P < 0.0008), treatment schedule (intermittent administration following a daily schedule, designated as combined schedule, P > 0.0014) and liver histology just before treatment (chronic persistent hepatitis or mild chronic aggressive hepatitis, P < 0.0324) were the most strongly correlated with a normalizing response to IFN therapy. These results suggest that not only are the IFN treatment schedule and patient profile significant, but the properties of the virus also influences the response. However, as the IFN treatment schedule is the only changeable factor, it should be designed to maximize the benefit of IFN therapy.
A method that allows the quantitation of hepatitis C virus (HCV) RNA is described. The RNA was extracted from serum samples and reverse transcribed. Target cDNA was then co‐amplified by nested polymerase chain reaction with a known amount of competitive template at various concentrations. Since this internal control DNA uses the same primers as those of the target and is distinguishable from the target cDNA after amplification by size, the initial concentration of the target could be estimated by comparing the intensity of the two bands of amplified DNA fragments. That is, if the starting amount of the cDNA and the competitive template are equal, the intensity of the two bands should also be equal. Using this method the amount of HCV RNA in serum samples obtained from 85 patients with chronic hepatitis type C was determined. There was as much as a 100 000‐fold difference in the levels of HCV RNA from patient to patient. A rapid decrease of HCV RNA in a patient treated with interferon is also described.
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