Isolation and Characterization of a Novel Gammaherpesvirus from a Microbat Cell Line

Shabman, Reed S.; Shrivastava, Susmita; Tsibane, Tshidi; Attie, Oliver; Jayaprakash, Anitha; Mire, Chad E.; Dilley, Kari E.; Puri, Vinita; Stockwell, Timothy B.; Geisbert, Thomas W.; Sachidanandam, Ravi; Basler, Christopher F.

We obtained a Myotis velifer incautus cell line that was originally derived from an interscapular tumor (MVI-it) and that was generated at the Naval Biosciences Laboratory and deposited at the American Type Culture Collection (ATCC; catalogue no. CRL-6012). The growth properties of this cell line changed over several weeks in culture, from doubling approximately once a week to rapidly proliferating, doubling every 2 to 3 days. We profiled the transcriptome of the cell line after it had begun rapid growth. Total RNA was isolated, and mRNA was purified and subjected to transcriptome sequencing (RNA-seq) followed by an initial de novo assembly that generated approximately 5,000 contigs. Unexpectedly, within this list of contigs 23 that exhibited homology to herpesvirus mRNAs were identified, suggesting the possibility that the cell line might harbor a replicating herpesvirus ( Table 1) . The viral transcripts detected included those for structural proteins, such as the major capsid protein, tegument protein, and glycoprotein B (gB), as well as proteins involved in genome replication, such as DNA polymerase and thymidine kinase, suggesting ongoing replication. A majority of the mRNAs potentially encoding viral proteins had a high level of identity (approximately 42 to 75% amino acid identity) to proteins of equine herpesvirus 2 (EHV-2), a horse gammaherpesvirus. Since each of these viral sequences showed substantial differences from any sequences deposited in NCBI, we refer to the virus as bat gammaherpesvirus 8 (BGHV8) based on suggested International Committee on the Taxonomy of Viruses (ICTV) nomenclature (20) and described bat gammaherpesvirus sequences (3).