Genotyping Hepatitis C Virus by Type-Specific Primers for PCR Based on NS5 Region

Hepatitis C virus (HCV), a flavi-like virus with a positive-sense, single-stranded genome RNA (1-3), shows considerable variation in nucleotide sequences. These variations fall into a series of specific patterns and are the basis for classification of HCV into different types and genotypes (4-9). Typing of HCV is of special interest because it has been suggested that different types or genotypes may cause different disease severities and show different sensitivity to interferon treatment (10-12). The number of types and genotypes of HCV continue to increase with the description of new sequences particularly from southeast Asian countries (13-16). More than 34 genotypes have been reported so far, and there is no doubt that the number will increase in the future with identification of new genotypes. Although there are several methods for determining the genotypes of HCV (5, 17-19), it is impossible to determine all these genotypes by one method. However, the prevalence of HCV genotypes has been well studied in some countries, including Japan and the United States (20-23). Detection of genotypes by polymerase chain reaction (PCR) using type-specific primers or probes is useful in this area where only a limited number of genotypes are known to exist. In this chapter, I describe a simple one-step PCR method to detect six major HCV genotypes (la, lb, 2a, 2b, 3a, and 3b) using genotype-specific primers based on the nucleotide sequences of the NS5 region.