Direct polymerase chain reaction for detection of hepatitis B, C and G virus genomes from serum without nucleic acid extraction—simple, rapid and highly sensitive method

Abstract

Although hepatitis C virus (HCV) detection by polymerase chain reaction (PCR) assay is now the standard, extensive clinical application has been thwarted by the troublesome procedure, long reaction time and potential for contamination. To overcome these problems, we carried out a PCR assay for the detection of HCV, hepatitis G virus (HGV) and hepatitis B virus (HBV) genomes directly from serum samples without any nucleic acid extraction (direct PCR). The sensitivity of this assay was one chimpanzee-infectious dose of HCV and a 10 −1 chimpanzee-infectious dose of HBV. This result was similar to the sensitivity determined by the conventional PCR. Furthermore, when the detection rate of these genomes in serum samples from chimpanzees and humans is compared, the results matched completely between two different PCR assays. The whole process, including the reverse transcriptase reaction and second round PCR, can be completed within 6 h by the combination of the direct PCR and one-step PCR assay. Our findings indicate that this method is simple, rapid and highly sensitive and could be useful for the screening of blood-borne hepatitis virus infections using serum samples.