Molecular epidemiology of hepatitis B and Delta virus strains that spread in the Mediterranean North East Coast of Tunisia

International audienceBackground: Tunisia is classified as an area of middle endemic for hepatitis B virus (HBV) infection, however little is known about hepatitis Delta virus (HDV) infection. Objectives: This study aimed to address the prevalence of HDV infection, to identify possible risks factors, and to analyze the genetic diversity of HDV strains that are spreading in Tunisia. Study design: A retrospective large-scale study including 1615 HBsAg positive patients, native of the North East coast of Tunisia, recruited from Gastroenterology departments, was conducted. Demographic, epidemiological, ethnical, clinical and biological data were recorded. HBV and HDV serological analyses and DNA and RNA viral load quantification were performed. Genotyping of HBV and HDV strains was performed using nucleotide sequencing followed by phylogenetic analyses. Results: The study population included 819(50.7%) men and 796(49.3%) women; aged 12-90 years (mean age 41 + 13 years). A very low prevalence of HDV infection, 2% was observed. No risk factor, except a history of hospitalization for surgery was found. All HDV strains belonged to genotype 1, with a wide distribution within the HDV-1 group. They all share the African amino acid marker, a serine at position 202 of the large Delta protein. HBV genotypes were distributed as follows: HBV/D1 (56.8%), HBV/D7 (40.9%), and HBV/A2 (2.3%). Conclusion: Tunisia is a low endemic region for HDV infection, due to an efficient policy of HBV infection control. HDV-1 is the sole genotype found, with a high diversity within this group. Further studies are ongoing in order to better characterize and manage the HBV/HDV-infected patients according to the genetic variability of the viral strains. (C) 2015 Elsevier B.V. All rights reserved

Phylogenetic Analysis and Epidemic History of Hepatitis C Virus Genotype 2 in Tunisia, North Africa.

HCV genotype 2 (HCV-2) has a worldwide distribution with prevalence rates that vary from country to country. High genetic diversity and long-term endemicity were suggested in West African countries. A global dispersal of HCV-2 would have occurred during the 20th century, especially in European countries. In Tunisia, genotype 2 was the second prevalent genotype after genotype 1 and most isolates belong to subtypes 2c and 2k. In this study, phylogenetic analyses based on the NS5B genomic sequences of 113 Tunisian HCV isolates from subtypes 2c and 2k were carried out. A Bayesian coalescent-based framework was used to estimate the origin and the spread of these subtypes circulating in Tunisia. Phylogenetic analyses of HCV-2c sequences suggest the absence of country-specific or time-specific variants. In contrast, the phylogenetic grouping of HCV-2k sequences shows the existence of two major genetic clusters that may represent two distinct circulating variants. Coalescent analysis indicated a most recent common ancestor (tMRCA) of Tunisian HCV-2c around 1886 (1869-1902) before the introduction of HCV-2k in 1901 (1867-1931). Our findings suggest that the introduction of HCV-2c in Tunisia is possibly a result of population movements between Tunisia and European population following the French colonization

Phylogenetic analysis of isolates from subtype 2c.

<p>Phylogenetic analyses were performed using the Maximum Composite Likelihood method included in the MEGA package (version 5.1). The reliability of the phylogenetic constructions was estimated by bootstrap analysis with 1000 pseudo replicate data sets. The tree is based on the analysis of a 222-nucleotide-long fragment in the NS5B region (nucleotides 8316 to 8537 according to the H77-1a prototype strain, AF009606). It includes the 102 studied Tunisian sequences (in red) and 53 HCV-2c field sequences from other countries (in black). All the sequences were indicated by their Accession Numbers followed by the country code and the collection date. The country codes according to the standard abbreviation are: France (FRA), Martinique (MTQ), Canada (CAN), Italy (ITA), Venezuela (VEN), Ghana (GHA), Russia (RUS), Netherlands (NLD), Spain (ESP), Japan (JPN), Germany (DEU), Argentina (ARG) and United Kingdom (GBR).</p

Bayesian Skyline Plots for Demographic Reconstruction using NS5B sequences.

<p>A) subtype 2c, B) subtype 2k; X-axis: Date in Years; Y-axis: Estimated effective number of infections; Bold Line: Mean Effective Number of viral population. Upper and lower Lines: Upper and Lower HPD95% (High Population Density) of Effective Number of viral population.</p

The timeline of HCV subtype 2c introduction in Tunisia.

<p>The timeline of HCV subtype 2c introduction in Tunisia.</p

_tMRCA estimations using the NS5B region for the HCV- 2c and HCV-2k in Tunisia.

<p>_tMRCA estimations using the NS5B region for the HCV- 2c and HCV-2k in Tunisia.</p

Phylogenetic analysis of isolates from Subtype 2k.

<p>Phylogenetic analyses were performed using the Maximum Composite Likelihood method included in the MEGA package (version 5.1). The reliability of the phylogenetic constructions was estimated by bootstrap analysis with 1000 pseudo replicate data sets. The tree is based on the analysis of a 222-nucleotide long fragment in the NS5B region (nucleotides 8346 to 8568 according to the H77-1a prototype strain, AF009606). It includes the 11 studied Tunisian sequences (in red) and 39 HCV-2k field sequences from other countries (in black). All the sequences were indicated by their Accession Numbers followed by the country code and the collection date. The country codes according to the standard abbreviation are: France (FRA), Martinique (MTQ), Canada (CAN), Russia (RUS), United Kingdom (GBR), Uzbekistan (UZB), Azerbaijan (AZE), Morocco (MAR), Iran (IRN) and Moldova (MDA).</p