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. 2015 Sep 21;10(9):e0137568.
doi: 10.1371/journal.pone.0137568. eCollection 2015.

Detection of Hepatitis B Virus (HBV) Genomes and HBV Drug Resistant Variants by Deep Sequencing Analysis of HBV Genomes in Immune Cell Subsets of HBV Mono-Infected and/or Human Immunodeficiency Virus Type-1 (HIV-1) and HBV Co-Infected Individuals

Z Lee  1 , S Nishikawa  1 , S Gao  1 , J B Eksteen  2 , M Czub  3 , M J Gill  4 , C Osiowy  5 , F van der Meer  6 , G van Marle  4 , C S Coffin  1
Affiliations

Detection of Hepatitis B Virus (HBV) Genomes and HBV Drug Resistant Variants by Deep Sequencing Analysis of HBV Genomes in Immune Cell Subsets of HBV Mono-Infected and/or Human Immunodeficiency Virus Type-1 (HIV-1) and HBV Co-Infected Individuals

Z Lee et al. PLoS One. .

Abstract

The hepatitis B virus (HBV) and the human immunodeficiency virus type 1 (HIV-1) can infect cells of the lymphatic system. It is unknown whether HIV-1 co-infection impacts infection of peripheral blood mononuclear cell (PBMC) subsets by the HBV.

Aims: To compare the detection of HBV genomes and HBV sequences in unsorted PBMCs and subsets (i.e., CD4+ T, CD8+ T, CD14+ monocytes, CD19+ B, CD56+ NK cells) in HBV mono-infected vs. HBV/HIV-1 co-infected individuals.

Methods: Total PBMC and subsets isolated from 14 HBV mono-infected (4/14 before and after anti-HBV therapy) and 6 HBV/HIV-1 co-infected individuals (5/6 consistently on dual active anti-HBV/HIV therapy) were tested for HBV genomes, including replication indicative HBV covalently closed circular (ccc)-DNA, by nested PCR/nucleic hybridization and/or quantitative PCR. In CD4+, and/or CD56+ subsets from two HBV monoinfected cases, the HBV polymerase/overlapping surface region was analyzed by next generation sequencing.

Results: All analyzed whole PBMC from HBV monoinfected and HBV/HIV coinfected individuals were HBV genome positive. Similarly, HBV DNA was detected in all target PBMC subsets regardless of antiviral therapy, but was absent from the CD4+ T cell subset from all HBV/HIV-1 positive cases (P<0.04). In the CD4+ and CD56+ subset of 2 HBV monoinfected cases on tenofovir therapy, mutations at residues associated with drug resistance and/or immune escape (i.e., G145R) were detected in a minor percentage of the population.

Summary: HBV genomes and drug resistant variants were detectable in PBMC subsets from HBV mono-infected individuals. The HBV replicates in PBMC subsets of HBV/HIV-1 patients except the CD4+ T cell subpopulation.

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Conflict of interest statement

Competing Interests: Dr. Coffin has received speaker, advisory board and/or consulting fees from Boehringer ingelheim, Janssen Pharmaceuticals, Bristol Myers Squibb, Roche Pharmaceuticals and Gilead Sciences. The other co-authors have no disclosures relating to this body of work. There are no other financial, personal, or professional interests that could be construed to have influenced the work. This does not alter the authors' adherence to all PLOS policies on sharing data and materials.

Figures

Fig 1
Fig 1. Comparison of median HBV covalently closed circular (ccc) DNA copies in peripheral blood mononuclear cells in HBV mono-infected (before and after antiviral treatment) and HBV/HIV-1 co-infected patients.
The HBV covalently closed circular (ccc)—DNA copies/peripheral blood mononuclear cell (PBMC) were determined by quantitative PCR using a TaqMan probe and normalized to a housekeeping gene (i.e., β-globin, β-glo). The HBV cccDNA copies/PBMC did not significantly differ between groups, i.e, HBV treatment naïve mono-infected group (n = 11): median 4.2, range 3.4–4.7 log10 copies/105 PBMC; HBV mono-infected on antiviral therapy (n = 4): median 3.8, range 3.6–3.9 log10 copies/105 PBMC; and HBV/HIV-1 co-infected (n = 2) mean 3.8 copies/105 PBMC.
Fig 2
Fig 2. Frequency (% of total reads) of HBV drug-resistant and immune escape (i.e., G145R) mutations detected in CD4+ and/or CD56+ immune cell subset by deep sequencing analysis in 2 HBV monoinfected cases (i.e., #3B and #8B) on tenofovir anti-HBV therapy.
See this image and copyright information in PMC

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