Bowden Ds

Hepatitis C virus genotypes in Australia

Summary. The relative distribution of Australian hepatitis C virus (HCV) genotypes was determined for 500 isolates. Genotyping was performed using a commercial reverse phase hybridization assay after amplification of the 5′ untranslated region of HCV by the polymerase chain reaction. Australian isolates comprised, predominantly, genotype 1 (55%) and genotype 3 (38%) with genotype 2 accounting for only 7%. Genotype 3a was the most common subtype. When the major risk groups of injecting drug users or transfusion‐acquired hepatitis C were compared, there was a significantly higher incidence of genotype 1b in the transfusion‐acquired group (P < 0.03). When the age of the patients was analysed, genotype 3a was more prevalent in the 21–40‐year age group than the 41–60‐year age group (P < 0.05). There was no significant difference in genotype distribution between males and females. HCV genotypes 1, 2 and 3 are most often found in developed countries but the relatively high prevalence of genotype 3a in Australia is unusual.

Short duration of lamivudine for the prevention of hepatitis B virus transmission in pregnancy: lack of potency and selection of resistance mutations

This study sought to assess the antiviral efficacy of lamivudine (LMV) administered during third trimester to reduce maternal viraemia and to identify the emergence of LMV resistance. A prospective observational analysis was performed on 26 mothers with high viral load (>107 IU/mL). Twenty‐one women received LMV (treated group) for an average of 53 days (range 22–88 days), and the remaining five formed the untreated control group. Serum samples from two time points were used to measure HBV DNA levels and antiviral drug resistance. The LMV‐treated women achieved a median HBV DNA reduction of 2.6‐log10 IU/mL. Although end‐of‐treatment (EOT) HBV DNA in four (18%) LMV‐treated women remained at >107 IU/mL (±0.5 log IU/mL), no mother‐to‐baby transmission was observed. In contrast, a baby from the untreated mother was HBsAg positive at 9 months postpartum. Four technologies were used for drug resistance testing. Only ultra‐deep pyrosequencing (UDPS) was sufficiently sensitive to detect minor viral variants down to <1%. UDPS showed that LMV therapy resulted in increased viral quasispecies diversity and positive selection of HBV variants with reverse transcriptase amino acid substitutions at sites associated with primary LMV resistance (rtM204I/V and rtA181T) in four (19%) women. These viral variants were detected mostly at low frequencies (0.63–5.92%) at EOT, but one LMV‐treated mother had an rtA181T variant that increased from 2.2% pretherapy to 25.59% at EOT. This mother was also infected with the vaccine escape variant (sG145R), which was inhibited by LMV treatment. LMV therapy during late pregnancy only reduced maternal viraemia moderately, and drug‐resistant viral variants emerged.

Hepatitis G: viroprevalence and seroconversion in a high‐risk group of children

Hepatitis G virus (HGV), a recently discovered flavivirus, is parenterally transmitted and significantly associated with hepatitis C viraemia. Data on the viroprevalence of this agent in children is scant and its seroprevalence is unknown. The aim of this study was to determine the viroprevalence and seroprevalence of HGV in paediatric patients at risk of parenterally transmitted virus infection. Sera from 35 patients, previously tested for hepatitis C virus (HCV) infection, were analysed for the presence of HGV RNA by reverse transcription–polymerase chain reaction (RT–PCR) and for antibody to the E2 envelope protein (anti‐E2) of HGV using the HGV‐env kit. The mean age of the patients was 9.4 years (range 1–17 years), and risk factors included multiple transfusions and maternal HCV infection. Co‐infection with HCV and HGV was a relatively common occurrence (31%). The prevalence of anti‐E2, a marker of recovery from infection, was low (5%) when compared with overall viroprevalence (20%). This study highlights the significant association of HGV with HCV in children. The novel finding of a low ratio of anti‐E2:HGV RNA contrasts with the pattern seen in adults and may reflect a higher risk of long‐term carriage with acquisition of HGV infection at an early age.

Serum guanase activity in hepatitis C virus infection.

Guanase (guanine aminohydrolase, EC 3.5.4.3), catalyses the hydrolytic deamination of guanine. The reaction is important because of its potential to infuence intracellular guanine nucleotide pools: guanine is directly salvagable to GMP, whereas its product xanthine is not (see Figure 1). Amongst human tissues, liver contains the highest guanase activity; activity is also detectable in brain and kidney [1, 2]. In most assays, guanase activity in other tissues and in normal plasma and serum is close to or below the detection limit [3–5]. Gross increases in serum guanase are considered a specific indication of hepatocellular damage and have been shown to have prognostic and diagnostic value in a variety of clinical situations [2–9]. However, routine guanase measurement is not widely available due to lack of sufficiently facile and accurate assays. We have developed a sensitive assay for guanase using reversed-phase ion-pair HPLC. Using this assay, we found that guanase activity in normal sera was lognormally distributed about a geometric mean of 0.85 units/litre, with no significant gender difference. When sera submitted to us for testing by polymerase chain reaction (PCR) for suspected active hepatitis C virus (HCV) infection were screened in parallel for guanase activity, increases above the 95% percentile of the normal range were found to be predictive of HCV positivity. In two separate groups of patients being treated with interferon-alpha for chronic HCV infection, guanase activity was elevated before treatment and decreased significantly during treatment. We conclude that serum guanase activity is a useful surrogate marker for HCV infection and that serial measurements may have prognostic value.

Mutations in the hepatitis B virus precore/core gene and core promoter in patients with severe recurrent disease following liver transplantation

Recurrent hepatitis B virus (HBV) infection is a major problem in patients undergoing liver transplantation. Previously, we reported that infection with HBV strains containing a mutation in the precore region (G-to-A at nucleotide 1896) was associated with severe recurrent disease posttransplantation. In this study we investigated other mutations in the precore/core gene and core promoter which may be associated with this severe recurrence. The precore/core gene and core promoter of HBV from pre and posttransplantation sera of 15 patients with HBV recurrence were amplified by polymerase chain reaction (PCR) and sequenced. Pre and posttransplant sequences were very similar for each patient. HBV from patients who developed severe recurrence had significantly more mutations in both the nucleotide (P < .05) and predicted amino acid (P < .05) sequences of the precore/core gene, but not in the core promoter, than virus from patients with mild recurrence. There was also an apparent link between severe disease and HBV strains of genotype D (P < .05). The number of nucleotide and amino acid mutations in the precore/core gene was strongly associated with the presence of the precore mutation (P < .01). Mutations were found throughout the entire gene, however, at the amino acid level clustering was observed in the B- and helper T-cell epitopes as well as nuclear localization signals. In the encapsidation signal, nucleotide mutations were found that were predicted to increase the stability of the stem-loop structure. Overall, our data shows that genotype D and accumulated mutations throughout the HBV precore/core gene, but not core promoter, were associated with severe recurrent disease posttransplantation. These mutations were strongly linked to the presence of the precore mutation at nucleotide position 1896 and may contribute to the poor outcome in these patients.