Complex Genotype Mixtures Analyzed by Deep Sequencing in Two Different Regions of Hepatitis B Virus

This study assesses the presence and outcome of genotype mixtures in the polymerase/surface and X/preCore regions of the HBV genome in patients with chronic hepatitis B virus (HBV) infection. Thirty samples from ten chronic hepatitis B patients were included. The polymerase/surface and X/preCore regions were analyzed by deep sequencing (UDPS) in the first available sample at diagnosis, a pre-treatment sample, and a sample while under treatment. HBV genotype was determined by phylogenesis. Quasispecies complexity was evaluated by mutation frequency and nucleotide diversity. The polymerase/surface and X/preCore regions were validated for genotyping from 113 GenBank reference sequences. UDPS yielded a median of 10,960 sequences per sample (IQR 16,645) in the polymerase/surface region and 11_,595 sequences per sample (IQR 14,682) in X/preCore. Genotype mixtures were more common in X/preCore (90%) than in polymerase/surface (30%) (p<0.001). On X/preCore genotyping, all samples were genotype A, whereas polymerase/surface yielded genotypes A (80%), D (16.7%), and F (3.3%) (p = 0.036). Genotype changes in polymerase/surface were observed in four patients during natural quasispecies dynamics and in two patients during treatment. There were no genotype changes in X/preCore. Quasispecies complexity was higher in X/preCore than in polymerase/surface (p = 0.004).

The results provide evidence of genotype mixtures and differential genotype proportions in the polymerase/surface and X/preCore regions. The genotype dynamics in HBV infection and the different patterns of quasispecies complexity in the HBV genome suggest a new paradigm for HBV genotype classification.

Below:  Regions of the HBV genome amplified for the present study: P/S is indicated in green and X/preCore in pink

Full article at:   http://goo.gl/8JD0CH

By:   

Andrea Caballero, Maria Homs, David Tabernero, Carolina Gonzalez, Maria Blasi, Leonardo Nieto, Francisco Rodriguez-Frias

Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain

Josep Gregori, Maria Homs, David Tabernero, Josep Quer, Maria Blasi, Mar Riveiro-Barciela, Rafael Esteban, Maria Buti, Francisco Rodriguez-Frias

Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain

Josep Gregori, Josep Quer, Rosario Casillas

Liver Diseases Unit, Vall d’Hebron Research Institute, Barcelona, Spain

Josep Gregori

Roche Diagnostics S.L., Sant Cugat del Vallès, Spain

Mar Riveiro-Barciela, Rafael Esteban, Maria Buti

Liver Unit, Hospital Universitari Vall d'Hebron, Universitat Autònoma de Barcelona (UAB), Barcelona, Spain

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