The sequence variability of the Epstein–Barr virus has been extensively studied throughout previous years in isolates from various geographic regions and consequent variations at both genetic and genomic levels have been described. However, isolates from South America were underrepresented in these studies. Here, we sequenced 15 complete EBV genomes that we analyzed together with publicly available raw NGS data for 199 EBV isolates from other parts of the globe by means of a custom-built bioinformatic pipeline. The phylogenetic relations of the genomes, the geographic structure and variability of the data set, and the evolution rates for the whole genome and each gene were assessed. The present work contributes to overcoming the scarcity of complete EBV genomes from South America and is the most comprehensive geography-related variability study, which involved determining the actual contribution of each EBV gene to the geographic segregation of the entire genome. Moreover, to the best of our knowledge, we established for the first time the evolution rate for the entire EBV genome based on a host–virus codivergence-independent assumption and assessed their evolution rates on a gene-by-gene basis, which were related to the encoded protein function. Considering the evolution of dsDNA viruses with a codivergence-independent approach may lay the basis for future research on EBV evolution. The exhaustive bioinformatic analysis performed on this new dataset allowed us to draw a novel set of conclusions regarding the genome evolution of EBV.

Epstein Barr virus sequence variability has been deeply studied throughout the past years in isolates from various geographic regions and consequent geographic variation at both genetic and genomic levels has been described. However, isolates from South America have been underrepresented in these studies. Here, we sequenced 15 complete EBV genomes that we analyzed, by means of a custom-built bioinformatic pipeline, together with publicly available raw NGS data for 199 EBV isolates from other parts of the globe. Phylogenetic relations of the genomes, geographic structure and variability of the data set, and evolution rates for the whole genome and each gene were assessed. The present study contributes to overcome the scarcity of EBV complete genomes from South America, and hence, achieves the most comprehensive geography-related variability study bydetermining the actual contribution of each EBV gene to the geographic segregation of the entire genomes. Moreover, to the best of our knowledge, we established for the first time the evolution rate for the entire EBV genome, on a host-virus codivergence-independent assumption, and statistically demonstrate that evolution rates, on a gene-by-gene basis, are related to the encoded protein function. Considering evolution of dsDNA viruses with a codivergence-independent approach, may lay the basis for future research on EBV evolution. Additionally, this work also expands the sampling-time lapse of available complete genomes derived from different EBV-related conditions, a matter that until today, prevents for detailed phylogeographic analysis.