Quantification of intrahepatic hepatitis B virus (HBV) DNA in patients with chronic HBV infection

Abstract

No data are available about the amount of hepatitis B virus (HBV) genomes in liver of patients with chronic HBV infection. The aim of this study was to quantify the intrahepatic HBV DNA in hepatitis B surface antigen (HBsAg)-positive patients with either active or suppressed viral replication and in HBsAg-negative subjects with occult HBV infection. We optimized the Roche ''Amplicor HBV Monitor'' kit for quantifying liver HBV DNA and analyzed hepatic DNA extracts and serum samples from 19 HBs-Ag-positive and 43 HBsAg-negative individuals. Eight of the HBsAg carriers had active HBV replication, and for 3 of them we analyzed samples obtained before and at the end of 1 year of lamivudine treatment. Five hepatitis Delta virus (HDV) coinfected patients and 6 healthy HBsAg carriers had inhibited HBV activity. Among the HBsAg-negative subjects 21 had occult HBV infection and 22 had no evidence of HBV infection. The median of HBV genomes per microgram of liver DNA milliliter of serum was 34,500 to 2,620,000 in patients with active viral replication, 20,000 to 3,900,000 before and 10,000 to 2,800 at the end of therapy in lamivudine-treated individuals, 9,800 to 600 in HDVinfected individuals, and 7,450 to 17,400 in healthy HBsAg carriers. These data indicate that cases with suppressed HBV activity, despite the very low levels of viremia, maintain a relatively high amount of intrahepatic viral genomes. This virus reservoir is likely involved in HBV reactivation, which is usually observed after stopping lamivudine treatment. Finally, the analysis of cases with occult HBV infection showed that the assay we used was able to specifically detect and quantify as few as 100 copies of viral genomes per microgram of liver DNA. (HEPATOLOGY 2000;31: 507-512.)Hepatitis B virus (HBV) is a major health problem worldwide and it is estimated that 350 million individuals are chronically infected with this virus. 1 Chronic HBV infection may be associated with a large spectrum of clinical forms ranging from the status of healthy carrier of hepatitis B surface antigen (HBsAg) to a moderate or severe chronic hepatitis susceptible of evolution to cirrhosis and development of hepatocellular carcinoma (HCC). 1 HBV-induced liver damage is strictly related with virus active replication, which is usually investigated detecting serum viral DNA by direct hybridization techniques. Consequently, HBsAg-positive patients with suppressed viral replication and undetectable levels of viremia have usually low grade or absence of hepatocellular injury. 2-4 Nevertheless, several observations suggest that an undefined amount of free viral genomes may persist into the hepatocytes of these subjects, 5,6 and such episomal HBV forms might be implicated in the reactivation of silent infections that may occur either spontaneously 7-9 or in conditions of immunosuppression, [10][11][12] or, in patients treated with antiviral drugs, after stopping therapy. [13][14][15][16][17] Thus, analysis and quantification of intrahepatic HBV genomes appear to be of biological inter...

COBAS AmpliPrep-COBAS TaqMan Hepatitis B Virus (HBV) Test: a Novel Automated Real-Time PCR Assay for Quantification of HBV DNA in Plasma

“…Two independent qualitative PCRs for the core and X HBV genes and CAP-CTM were used to assess HBV DNA in plasma and liver biopsy samples from 26 HBsAg-negative, anti-HBcAg-positive liver donors (16 of 19 were also reactive for antibody to HBsAg). The two in-house PCRs detected HBV DNA with a higher sensitivity (10 copies of DNA/g) than CAHBM, as demonstrated by previously published studies (5,10). Plasma from all donors was negative for HBV DNA with CAP-CTM at the time of organ donation and with the two nested PCRs at a retrospective analysis.…”

Persistent hepatitis B virus infection in subjects without hepatitis B surface antigen: Clinically significant or purely “occult”?

“…Liver HBV-DNA levels were also low in HBsAg-negative HBV-DNA-positive subjects with chronic hepatitis and cirrhosis. 2,3 Investigations performed in hepatocellular carcinoma (HCC) tissues also showed a low copy number per cell of HBV genomes (around 0.1 to 0.01). 2 Moreover, we have observed some heterogeneity in the distribution of the viral genome within the liver that may further complicate the interpretation of negative results (Bréchot et al, personal observations).…”

Anti-HBc Screening of Blood Donors in Bangladesh: Relevance to Containment of HBV Propagation

“…With the advent of new technologies with high sensitivity and specificity for detection of HBV DNA, it became evident that a group of people do not express HBsAg, but harbor HBV DNA. 14,15 Most of the developed and advanced countries of the world modified blood transfusion procedures on the basis of new information and donor blood have been checked for antiHBc and ALT. Recently, nucleic acid testing (NAT) has been adopted by many of those countries to avoid transfusion-induced hepatitis.…”