Targeted enrichment of Elephant Endotheliotropic Herpesvirus for complete genome sequencing in elephants

Elephant Endotheliotropic Herpesvirus (EEHV) poses a critical threat to young Asian (Elephas maximus) and African elephants (Loxodonta africana), with high mortality rates due to acute haemorrhagic symptoms from vascular endothelial damage. EEHV remains dormant in adult elephants, reactivating under stress or immune suppression. There are multiple genotypes of EEHV have been reported based on genetic heterogeneity. The diversity among EEHV subtypes, such as EEHV1A and EEHV1B, influences disease severity and host susceptibility. EEHV genomes are large and encode over 115 open reading frames, contributing to viral replication and immune evasion. Hence, generation of complete genome is essential to determine the genotypes of EEHV. However, the inability to culture EEHV in-vitro limits the study of its life cycle and pathogenesis, necessitating molecular techniques for direct analysis. Additionally, variable viral load in the infected tissues limits the success of DNA sequencing using direct sequencing. Hence, in this study several methods of viral or DNA enrichment were considered, and ultracentrifugation method was adopted to concentrate viral copies for improved genomic analysis. Ultracentrifugation demonstrates effectiveness in isolating EEHV, significantly enriching viral DNA for detection and characterization. This technique facilitates advancements in diagnostics, therapeutic development, and vaccine research, addressing the challenges of low viral loads and host DNA contamination in clinical samples.