Complete Genome Sequences of Elephant Endotheliotropic Herpesviruses 1A and 1B Determined Directly from Fatal Cases

Abstract

e A highly lethal hemorrhagic disease associated with infection by elephant endotheliotropic herpesvirus (EEHV) poses a severe threat to Asian elephant husbandry. We have used high-throughput methods to sequence the genomes of the two genotypes that are involved in most fatalities, namely, EEHV1A and EEHV1B (species Elephantid herpesvirus 1, genus Proboscivirus, subfamily Betaherpesvirinae, family Herpesviridae). The sequences were determined from postmortem tissue samples, despite the data containing tiny proportions of viral reads among reads from a host for which the genome sequence was not available. The EEHV1A genome is 180,421 bp in size and consists of a unique sequence (174,601 bp) flanked by a terminal direct repeat (2,910 bp). The genome contains 116 predicted protein-coding genes, of which six are fragmented, and seven paralogous gene families are present. The EEHV1B genome is very similar to that of EEHV1A in structure, size, and gene layout. Half of the EEHV1A genes lack orthologs in other members of subfamily Betaherpesvirinae, such as human cytomegalovirus (genus Cytomegalovirus) and human herpesvirus 6A (genus Roseolovirus). Notable among these are 23 genes encoding type 3 membrane proteins containing seven transmembrane domains (the 7TM family) and seven genes encoding related type 2 membrane proteins (the EE50 family). The EE50 family appears to be under intense evolutionary selection, as it is highly diverged between the two genotypes, exhibits evidence of sequence duplications or deletions, and contains several fragmented genes. The availability of the genome sequences will facilitate future research on the epidemiology, pathogenesis, diagnosis, and treatment of EEHV-associated disease.

Comparative Genome Analysis of Four Elephant Endotheliotropic Herpesviruses, EEHV3, EEHV4, EEHV5, and EEHV6, from Cases of Hemorrhagic Disease or Viremia

“…On the basis of phylogenetic and genome organization analyses, we proposed that EEHV1 and EEHV2 represented distinct species within the Proboscivirus genus and that they might be better considered the prototypes of a newly proposed Deltaherpesvirinae subfamily of mammalian herpesviruses rather than their previously assigned status as outliers of the Betaherpesvirinae subfamily (10). The recently completed next-generation sequencing-based genome analysis of very high quality necropsy DNA samples from three more strains of EEHV1 (13,14) has also revealed another 40 or so novel genes in both EEHV1A and EEHV1B, but these genes lie outside the areas covered in the current studies. To further evaluate the four other known major types of EEHV genomes, we carried out selective PCR DNA sequencing to generate between 4 and 32 kb each from six more viremic disease cases (Table 1) representing the prototypes of EEHV3, EEHV4, EEHV5, and EEHV6.…”

“…For both virus types, this should represent close to 15% of their total genomes, considering the observed nearly180-kb size for EEHV1 (13,14). A schematic summary diagram showing the map locations of all of these sequenced loci relative to the intact genome of EEHV1A(Kimba) is presented in Fig.…”

“…Sanger PCR-based DNA sequence data for between 20 to 70 kb each from seven representative strains of either EEHV1A and EEHV1B, as well as for 59 kb from a single prototype strain of EEHV2, have been generated and described in the preceding accompanying paper (10), adding to two earlier less extensive studies (11,12). Furthermore, the complete intact 180-kb genomes of three more EEHV1 strains were determined recently by de novo next-generation sequencing of necropsy tissue samples (13,14). The genomes of EEHV1A, EEHV1B, and EEHV2 together form a very distinctive family of herpesviruses with unique genomic organization characteristics.…”

Commentary: Trunkloads of Viruses

“…As a group, EEHV display forms and levels of sequence diversity that are collectively unique with respect to forms of variation seen for other groups of herpesviruses. Somewhat analogous to human herpesviruses 6A and 6B (16), most of the 64 genes that span the center of the EEHV1A and EEHV1B genomes are Ͼ99% identical, with three interspersed sharply bounded regions of much greater sequence divergence (referred to as chimeric domains, or CD), as well as some sequence rearrangements in the vicinity of the genomic termini (4,8). Although less sequence is available, EEHV5A and EEHV5B appear to have a similar relationship (9).…”

Bioinformatics tools for analysing viral genomic data

“…As an example, this approach was used to construct the genome sequence of elephant endotheliotropic herpesvirus from samples in which as little as 0.04% of the DNA was viral (45). Sequencing errors disrupt the assembly process, so it is essential to trim poor quality bases and remove adapter sequences from the read datasets before running assemblies.…”