Host reverse transcriptase helps establish and maintain persistent dengue virus infections in C6/36 insect cells

The mechanisms involved in maintenance of persistent Dengue virus-2 (DEN-2) infections in C6/36 cells have not yet been reported. However, the persistence of RNA viruses in Drosophila melanogaster was achieved through the combined action of cellular reverse-transcriptase (RT) activity and RNA-mediated interference (RNAi). RT inhibition in C6/36 cells was achieved using tenofovir. Toxicity tests using tenofovir at 0.1, 0.25 and 0.5 mM revealed some cell death at 0.5 mM but only very limited effects at 0.1 mM. Normally, challenging naive Aedes albopictus C6/36 cells with Dengue virus-2 (DEN-2) results in relatively mild, transitory cytopathic effects (CPE), and 3-5 split passages result in a return to normal morphology and growth despite the presence of persistent, active infections of DEN-2 in all the cells. Here, it is shown that treatment with tenofovir prior to and after challenge with DEN-2 results instead in severe CPE and death (i.e., failure to establish a persistent infection). In addition, treatment of stable, grossly normal C6/36 cell cultures persistently-infected with DEN-2 induces severe CPE and death. The results indicate a key role of host RT in establishment and maintenance of stable, persistent DEN-2 infections. The underlying mechanisms are unknown, but this cell culture system should provide a good model for further studies to reveal them.