Detection of serum hepatitis B virus DNA in patients with chronic hepatitis using the polymerase chain reaction assay.

Abstract

We compared the sensitivity of the polymerase chain-reaction (PCR) assay to that of slot-blot hybridization for detecting hepatitis B virus (HBV) DNA in the serum of 31 patients with chronic hepatitis. Of 14 chronic hepatitis patients positive for both HBV surface and HBV e antigens, 9 were positive for HBV DNA by slot-blot hybridization and all 14 by PCR. Also, of 9 patients positive for HBV surface antigen and antibody against HBV e antigen, 2 were positive for HBV DNA by slot-blot analysis and 8 by PCR. Finally, in 8 patients positive for antibody against HBV surface antigen, none were positive for HBV DNA by slot-blot hybridization, but 4 were positive by PCR. We find that analysis by the PCR technique provides a >104-fold increase in sensitivity over the slot-blot hybridization assay. This result represents an important breakthrough in sensitivity because it is now possible to detect as few as three HBV DNA genomes per sample of serum.The polymerase chain reaction (PCR) technique is a method for amplifying nucleic acid by repeated cycles of hightemperature template denaturation, oligonucleotide primer hybridization, and polymerase extension. The thermostable polymerase of Thermus aquaticus (Taq) is often used because it can withstand the high denaturation temperatures without loss of enzymatic activity. Previous investigations with this technique have shown amplification of >105-fold over the input quantity of nucleic acid (1, 2). We have used PCR amplification, combined with agarose gel electrophoresis and Southern blot hybridization analysis, to detect extremely small quantities of hepatitis B virus (HBV) DNA in the serum of chronic hepatitis patients.

Methodology of a nationwide cross-sectional survey of prevalence and epidemiological patterns of hepatitis A, B and C infection in Brazil

“…Nested PCR was carried out as described by Kaneko et al 23 , with some modifications. The S region was amplified according to a standard technique 24 for identification of the HBV genotype.…”

Clinical and Serological Variation between Patients Infected with Different Hepatitis B Virus Genotypes

“…Conversely, it was believed for several years that patients who had cleared HBeAg and seroconverted to anti-HBe positivity were noninfectious. Clinical studies in which HBV DNA was detected in HBsAg-positive patients with anti-HBe, initially by hybridization and in subsequent studies by PCR, demonstrated that a proportion of HBsAg-positive patients who clear HBeAg still show signs of replicative infection (14). Carman et al (9) put forward a molecular explanation for this.…”

Genotypes of hepatitis B virus identified in patients tested prior to endoscopy from a Teaching Hospital in the Central Province of Sri Lanka

“…This might be due to the high number of HBsAg particles but lack of HBV in the sera [5]. Another reason for this finding would be that the blood samples from the corresponding patients would have been obtained in the latter rather than early phases of chronic HBV carriage where the levels of HBV DNA are likely to be Brief report low in the serum due to clearance by the immune system resulting in negative PCR [6].…”