Genomic characterization of serial-passaged Ebola virus in a boa constrictor cell line

Ebola virus disease (EVD) is a viral hemorrhagic fever with a high case-fatality rate in humans. EVD is caused by four members of the filoviral genus Ebolavirus, with Ebola virus (EBOV) being the most notorious one. Although bats are discussed as potential ebolavirus reservoirs, limited data actually support this hypothesis. Glycoprotein 2 (GP2) of reptarenaviruses, known to infect only boa constrictors and pythons, are similar in sequence and structure to ebolaviral glycoprotein 2 (GP2), suggesting that EBOV may be able to infect snake cells. We therefore serially passaged EBOV and a distantly related filovirus, Marburg virus (MARV), in the boa constrictor kidney cell line, JK, and characterized viral growth and mutational frequency by sequencing. We observed that EBOV efficiently infected and replicated in JK cells, but MARV did not. In contrast to most cell lines, EBOV infected JK cells did not result in obvious cytopathic effect (CPE). Genomic characterization of serial-passaged EBOV in JK cells revealed that genomic adaptation was not required for infection. Deep sequencing coverage (>10,000x) demonstrated the existence of only a single non-synonymous variant (EBOV glycoprotein precursor preGP T544I) of unknown significance within the viral population that exhibited a shift in frequency of at least 10% over six passages. Our data suggest that boid snake derived cells are competent for filovirus infection without appreciable genomic adaptation; that cellular filovirus infection without CPE may be more common than currently appreciated; and that there may be significant differences between the natural host spectra of ebolaviruses and marburgviruses.\n\nIMPORTANCEEbola virus (EBOV) causes a high case-fatality form of viral hemorrhagic fever. The natural reservoir of EBOV remains unknown. EBOV is distantly related to Marburg virus (MARV), which has been found in bats in the wild. The glycoprotein of a reptarenavirus known to infect boid snakes (pythons and boas) shows similarity in sequence and structure to these viruses, suggesting that EBOV and MARV may be able to infect and replicate in snake cells. We demonstrate that JK, a boa constrictor cell line, does not support MARV infection, but does support EBOV infection without causing overt cytopathic effect or the need for appreciable adaptation. These findings suggest different filoviruses may have a more diverse natural host spectra than previously thought.