Lilly Yuen

Circulation of Type 1 Vaccine-Derived Poliovirus in the Philippines in 2001

ABSTRACT In 2001, highly evolved type 1 circulating vaccine-derived poliovirus (cVDPV) was isolated from three acute flaccid paralysis patients and one contact from three separate communities in the Philippines. Complete genomic sequencing of these four cVDPV isolates revealed that the capsid region was derived from the Sabin 1 vaccine strain but most of the noncapsid region was derived from an unidentified enterovirus unrelated to the oral poliovirus vaccine (OPV) strains. The sequences of the cVDPV isolates were closely related to each other, and the isolates had a common recombination site. Most of the genetic and biological properties of the cVDPV isolates were indistinguishable from those of wild polioviruses. However, the most recently identified cVDPV isolate from a healthy contact retained the temperature sensitivity and partial attenuation phenotypes. The sequence relationships among the isolates and Sabin 1 suggested that cVDPV originated from an OPV dose given in 1998 to 1999 and that cVDPV circulated along a narrow chain of transmission. Type 1 cVDPV was last detected in the Philippines in September 2001, and population immunity to polio was raised by extensive OPV campaigns in late 2001 and early 2002.

The L80I Substitution in the Reverse Transcriptase Domain of the Hepatitis B Virus Polymerase Is Associated with Lamivudine Resistance and Enhanced Viral Replication In Vitro

ABSTRACT Long-term lamivudine (LMV) treatment of chronic hepatitis B almost inevitably engenders viral resistance. Mutations that result in the replacement of the methionine at position 204 of the deoxynucleoside triphosphate-binding site of the hepatitis B virus (HBV) reverse transcriptase (rt) by isoleucine, valine, or (rarely) serine (rtM204I/V/S) confer high-level resistance to LMV but reduce replication efficiency. The subsequent selection or coselection of secondary mutations that partially restore replication efficiency is common and may influence drug resistance. Genotyping has shown that LMV treatment can select for HBV rtL80V/I mutants, but their prevalence and phenotype have not been documented. Analysis of a large sequence database revealed that rtL80V/I occurred almost exclusively in association with LMV resistance, and 85% of these isolates encoded rtL80I. Coselection of rtL80V/I occurred in 46% of isolates in which LMV resistance was attributable to rtM204I but only 9% of those in which resistance was attributable to rtM204V. Moreover, rtL80V/I did not occur in HBV genotype A isolates but occurred at similar frequencies in genotype B, C, and D isolates. In vitro phenotyping showed that although the rtL80I mutant by itself replicated less efficiently and was hypersensitive to LMV compared to the replication efficiency and sensitivity of its wild-type parent, the presence of rtL80I enhanced the replication efficiency of rt204I/V mutants without significantly affecting LMV resistance. Molecular modeling revealed that rt80 does not interact directly with the enzymes substrates. Collectively, these results suggest that coselection of rtL80V/I and rtM204I/V occurs because the former compensates for the loss of replication efficiency associated with the acquisition of LMV resistance, particularly in the case of rtM204I.

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.

Possible origins and evolution of the hepatitis B virus (HBV)

All members of the family Hepadnaviridae are primarily viruses which contain double-stranded DNA genomes that are replicated via reverse transcription of a pregenomic RNA template. There are two subgroups within this family: mammalian and avian. The avian members include the duck hepatitis B virus (DHBV), heron hepatitis B virus, Ross goose hepatitis B virus, stork hepatitis B virus and the recently identified parrot hepatitis B virus. More recently, the detection of endogenous avian hepadnavirus DNA integrated into the genomes of zebra finches has revealed a deep evolutionary origin of hepadnaviruses that was not previously recognised, dating back over 40 million years ago. The non-primate mammalian members of the Hepadnaviridae include the woodchuck hepatitis virus (WHV), the ground squirrel hepatitis virus and arctic squirrel virus, as well as the recently described bat hepatitis virus. The identification of hepatitis B virus (HBV) in higher primates such as chimpanzee, gorilla, orangutan, and gibbons that cluster with the human genotypes further implies a more complex origin of this virus. By studying the molecular epidemiology of HBV in indigenous and relict populations in Asia-Pacific we propose a model for the origin and evolution of HBV that involves multiple cross-species transmissions and subsequent recombination events on a background of genotype C HBV infection.

Lamivudine resistance in patients with chronic hepatitis B: Role of clinical and virological factors

Background:  Lamivudine resistance is associated with long‐term monotherapy for chronic hepatitis B and can lead to potentially serious clinical consequences. Scant information exists regarding the influence of hepatitis B virus variants in the development of resistance. The present study was designed to identify factors predictive of lamivudine resistance, with a particular focus on the role of precore and basal core promoter variants in the setting of hepatitis B e antigen‐negative disease.

Identification of a novel hepatitis B virus precore/core deletion mutant in HIV/hepatitis B virus co-infected individuals.

Objectives:Although HAART has resulted in improved health outcomes for most HIV-infected individuals, liver failure has emerged as a major cause of morbidity and mortality in people co-infected with hepatitis B virus (HBV). In HBV mono-infected individuals, core deletion mutants are associated with more aggressive liver disease. As HIV accelerates HBV liver disease progression, we hypothesized that HIV-HBV co-infected individuals have increased frequency of core mutations including deletions. To test this hypothesis, we have analysed genome-length sequences of HBV DNA from patients both prior to and during antiviral therapy. Setting:Prospective HIV/HBV co-infected cohort study. Methods:Genomic length HBV DNA was amplified by PCR from the serum samples of ten HIV/HBV co-infected individuals and five HBV mono-infected individuals prior to the commencement of lamivudine therapy and again after nine to 74 months of treatment. The complete genomes were sequenced and in order to further analyse some mutations, their frequency was determined in additional HIV/HBV co-infected and HBV mono-infected individuals. Results:A novel –1G mutation was identified in the HBV precore and overlapping core genes that truncated the deduced precore/core proteins. The mutant genome was the dominant species in some HIV/HBV co-infected individuals and was more prevalent in HIV/HBV co-infected individuals than HBV mono-infected individuals. The mutation was also associated with high HBV DNA concentrations in HIV/HBV co-infected individuals. Additional mutations were identified in the core/precore and polymerase genes and regulatory regions. Conclusion:Mutations in the HBV core and precore genes may be contributing to disease pathogenesis in HIV/HBV co-infected individuals.

Mutation pattern of lamivudine resistance in relation to hepatitis B genotypes: Hepatitis B genotypes differ in their lamivudine resistance associated mutation pattern†

Little is known about differences between individual hepatitis B genotypes and mutation patterns associated with lamivudine resistance. This study analyses the lamivudine‐associated mutation pattern in relation to the four major HBV genotypes A–D. The PubMed database was screened for keywords “HBV OR Hepatitis B,” “YMDD,” “genotype,” and “lamivudine”; all identified publications published till June 2009 were analyzed for differences in mutation pattern. To confirm the literature‐based findings the databases of two reference laboratories in Tübingen (Germany), and Melbourne (Australia) were analyzed. Twenty‐nine studies were identified reporting 827 patients with known hepatitis B genotype who underwent lamivudine treatment and developed resistance mutations. The literature data revealed that genotype A favors the rtM204V mutation unlike the other major genotypes (P < 0.001), which corresponds to a significant difference in the mutation pattern of genotypes endemic in Asian countries and those found in the rest of the world. These significant findings of the literature‐review could be reproduced in the analysis of the databases from Tübingen and Melbourne. Furthermore, the rtL180M mutation is significantly connected to the rtM204V mutation in genotypes A, B, and C, respectively. It is concluded that there is proof that HBV genotypes differ in their mutation pattern of lamivudine resistance. Future studies will need to evaluate whether this will translate into genotype‐specific differences in resistance emergence on either entecavir or telbivudine as these antivirals differ in their mutation profile, rtM204V for entecavir and rtM204I for telbivudine. J. Med. Virol. 82:1850–1858, 2010.

Molecular and clinical characteristics of hepatitis B virus in Korea

Korea is an endemic area of hepatitis B virus (HBV) infection but very little is known about the molecular characteristics of HBV isolates from Korean patients or the association with disease progression. The complete HBV genome sequences from 53 Korean patients with chronic hepatitis B, advanced cirrhosis, or hepatocellular carcinoma (HCC) were analyzed to identify (i) subgenotype distribution and genetic diversity and (ii) signature mutations associated with liver disease progression. With the exception of 1 patient infected with HBV/B, all 52 patients (98.1%) were infected with HBV/C, subgenotype C2. These strains were 98.4% identical and the frequency of amino acid substitutions occurring within key immunological epitopes increased with disease severity. A number of amino acid/nucleotide substitutions were associated with HCC, namely sR24K (HBsAg), SI126T (HBsAg), and pcA1846T (precore gene) mutations (P = 0.029, 0.001, and 0.008, respectively). HBV harboring deletions in the pre‐S region were also associated with increased liver disease severity (chronic hepatitis B vs. cirrhosis, P = 0.040; chronic hepatitis B vs. HCC, P = 0.040). Despite the high degree of sequence conservation, several key HBV mutations were associated with disease progression. Prospective studies with larger cohorts of patients are required to evaluate further the clinical manifestation of HBV/C2 in Korea. J. Med. Virol. 82: 1126–1134, 2010.

Prolonged use of tenofovir in HIV/hepatitis B virus (HBV)-coinfected individuals does not lead to HBV polymerase mutations and is associated with persistence of lamivudine HBV polymerase mutations.

The aim of the study was to identify and characterize hepatitis B virus (HBV) polymerase gene mutations associated with ongoing HBV replication in HIV/HBV‐coinfected individuals receiving tenofovir (TDF).

Origins and Evolution of Hepatitis B Virus and Hepatitis D Virus

Members of the family Hepadnaviridae fall into two subgroups: mammalian and avian. The detection of endogenous avian hepadnavirus DNA integrated into the genomes of zebra finches has revealed a deep evolutionary origin of hepadnaviruses that was not previously recognized, dating back at least 40 million and possibly >80 million years ago. The nonprimate mammalian members of the Hepadnaviridae include the woodchuck hepatitis virus (WHV), the ground squirrel hepatitis virus, and arctic squirrel hepatitis virus, as well as a number of members of the recently described bat hepatitis virus. The identification of hepatitis B viruses (HBVs) in higher primates, such as chimpanzee, gorilla, orangutan, and gibbons that cluster with the human HBV, as well as a number of recombinant forms between humans and primates, further implies a more complex origin of this virus. We discuss the current theories of the origin and evolution of HBV and propose a model that includes cross-species transmissions and subsequent recombination events on a genetic backbone of genotype C HBV infection. The hepatitis delta virus (HDV) is a defective RNA virus requiring the presence of the HBV for the completion of its life cycle. The origins of this virus remain unknown, although some recent studies have suggested an ancient African radiation. The age of the association between HDV and HBV is also unknown.