Genome-wide comparative study of epstein-barr virus genomic recombination in endemic burkitt lymphoma and healthy children from jootrh, western Kenya

Abstract

Endemic Burkitt lymphoma (eBL), the most prevalent pediatric cancer in Western Kenya, is augmented by the interplay between Epstein-Barr Virus (EBV) and holoendemic malaria infection. Despite the relevance of genomic recombination on EBV genetic diversity, its genome-wide occurrence has not been characterized in genomic sequences from western Kenya hence the association with age, gender, and EBV type and eBL pathogenesis in children from western Kenya is unknown. This study, therefore, sought to:1) characterize genome-wide occurrence of EBV genomic recombination events and breakpoints; 2) establish relationship between EBV genomic recombination events with age and gender; 3) establish relationship between genomic recombination events with EBV type and EBV genetic diversity; 4) determine the association of genomic recombination events and breakpoints with eBL. This study employed a case-control design using 86 archival samples involving 54 children diagnosed with eBL and 32 geographically matched healthy children previously collected from Jaramogi Oginga Odinga Teaching and Referral Hospital (JOORTH) that met the inclusion criteria. DNA was extracted and sequenced in the Illumina Sequencing Kit. Whole-genome multiple sequence alignment, recombination analyses, and phylogenetic inferencing were done using MAFFT, RDP4, and MEGA X software respectively. Wilcoxon rank test compared the occurrence of genomic recombination between genomic sequences of the; males and females, type 1 and type 2, and between the eBLs and the healthy. Univariate and multivariate logistic regression modeled eBL association with genomic recombination events and their breakpoints. This study identified 28 genomic recombination events present in 82.6% of the EBV genomes analyzed with most breakpoints reported in genes of the lytic phase. There was no significant difference across the age groups (p=0.68) and between males and females (p=0.59). Type 1 genomic sequences reported more genomic recombination events (p=6.4e-06). The EBV genomic sequences clustered on the neighbor-joining (NJ) phylogenetic tree by genomic recombination events suggesting an association with EBV genetic diversity. Genomic sequences from the eBLs reported more genomic recombination events compared to the healthy (p=0.037). Further, recombination breakpoints cutting through; BRLF1, BZLF1, BDLF3.5, BDLF4, LMP2A, LMP2B, and EBNA2 genes were found enriched in genomic sequences from the eBLs. Evidence from this study suggests that there is minimal accrual of genomic recombination events with infections over time hence these genomic recombination events are most likely transferred vertically down EBV genome lineages. Type 1 EBV genomic sequences and those from the eBLs have accumulated more genomic recombination events pointing to the availability of factors that increases the propensity for genomic recombination in these set of genomic sequences. For future studies, long-read sequencing and improved EBV DNA enrichment methods should be employed to generate complete EBV sequences to allow the analysis of whole EBV genomes. In summary, this study addresses the complexities that underlie genomic recombination as a source of genetic variation in EBV, findings that contribute significantly to the pool of knowledge on EBV genetic diversity and its contribution to disease.