Phylogenetic Analysis of Hepatitis B Virus Genotypes Circulating in Different Risk Groups of Panama, Evidence of the Introduction of Genotype A2 in the Country

<div><p>The Hepatitis B Virus (HBV) can cause acute or chronic infection it is also associated with the development of liver cancer, thousands of new infections occur on a yearly basis, and many of these cases are located in certain areas of the Caribbean and Latin America. In these areas, the HBV prevalence is still high which makes this virus a serious public health concern to the entire region. Studies performed in Panama suggest a complex pattern in the distribution of HBV among the country’s different risk groups. We use phylogenetic analysis in order to determine which HBV genotypes were circulating in these specific groups; for this we used a fragment of the PreS2/2 region of the HBV genome. Subsequently whole HBV genome sequences were used for Bayesian analysis of phylodynamics and phylogeography. Two main genotypes were found: genotype A (54.5%) and genotype F (45.5%). There was a difference in the distribution of genotypes according to risk groups: 72.9% of high risk groups were associated to genotype A, and 55.0% of samples of genotype F were associated to the low risk group (p<0.002). The Bayesian analysis of phylogeny-traits association revealed a statistically significant geographical association (p<0.0001) with both genotypes and different regions of the country. The Bayesian time of most recent common ancestor analysis (tMRCA) revealed a recent tMRCA for genotype A2 circulating in Panama (1997, 95% HPD: 1986—2005), when it is compared with Panamanian genotype F1c sequences (1930, 95% HPD: 1810 – 2005). These results suggest a possible change in the distribution of HBV genotypes in Panama and Latin America as a whole. They also serve to encourage the implementation of vaccination programs in high-risk groups, in order to prevent an increase in the number of new HBV cases in Latin America and worldwide.</p></div

Spatial dissemination of HBV genotypes across the world.

<p>Lines between countries represent sequences in which transition location occurs, and were supported by a significant non-zero rate (Bayes Factor >3). Map (a) is for genotype F (a), and map (b) for genotype A. Lines were colored according to the sub-genotype involved in the migration.</p

Estimated of tMRCA and substitution rate for Genotype A and F, with the HBV Whole genome of the Panamanian samples.

<p>Abbreviations: tMRCA, mean time to the most recent common ancestor; 95% HPD, 95% highest posterior density; subs/site/year, substitutions per site year.</p><p>Estimated of tMRCA and substitution rate for Genotype A and F, with the HBV Whole genome of the Panamanian samples.</p

Bayesian phylogeny-traits association results, according to risk behavior group and geographic location.

<p>Abbreviations: Association index (AI), parsimony score (PS), and monophyletic clade (MC), Highest posterior density (HPD), confidence interval (CI). For the MC statistic the corresponding trait is indicated between parentheses, for p value calculation the desired level of significance was ≤0.05.</p><p>Bayesian phylogeny-traits association results, according to risk behavior group and geographic location.</p

Maximum Likelihood tree of HBV Panamanian sequences.

<p>The tree is based on HBV partial PreS2-S region (~820pb), using reference sequences of each genotype (A-I) (n = 280) and sequences of Panamanian origin (n = 88) which are colored in blue. The place of sample collection is indicated in the taxa name, the symbols represent the behavioral risk groups to which the sample belongs: bold triangle: low risk, bold hexagon: high-risk group (bold circle). Clades in which there are not Panamanian samples are collapsed to simplify, node numbers correspond to αLTR values higher than 0.75.</p

Map of Panama indicating the origin of the HBV sequences analyzed in this study.

<p>The number of sequences according to the genotype is indicated in the top left pie chart, the middle bar plot indicates the number of genotypes according to geographic area. Pie charts within the map are sized by the proportion of sub-genotype found in the corresponding region.</p

Genotype distribution of the HBV sequences evaluated according to risk behavior group, geographic location and the mutations found in BCP, X/PreC and Polymerase.

<p>Data are: Number (%)</p><p>* This number represent the total of samples successfully amplified.</p><p>^&Chiriqui, Bocas del toro, Ngobe Bugle.</p><p>^$$Los Santos, Cocle.</p><p>°Commercial sex workers</p><p>°°Men that have sex with Men.</p><p>^&Exact-Fisher test statistic.</p><p>**Double mutation 1762T/1764.</p><p>*** Any combination of mutations: 1613A, 1653T, 1846T, 1896, 1899A as suggested by Park et al (49).</p><p>****All the subjects with resistance mutation were HIV positive.</p><p>Genotype distribution of the HBV sequences evaluated according to risk behavior group, geographic location and the mutations found in BCP, X/PreC and Polymerase.</p