| Popis: |
It has been long known that C6/36 cells challenged with Dengue virus serotype 2 (DENV-2) show initial cytopathic effects (CPE) but overcome these within 3 split passages to resume normal growth and morphology. It is also known that establishment of persistent infections requires combined action of host reverse-transcriptase (RT) activity and RNA-mediated interference (RNAi). We hypothesized that RT inhibition in C6/36 cells would also prevent persistent infections with DENV-2 and that treatment of stable, grossly normal C6/36 cells persistently infected with DENV-2, even after long passaging, would revert to CPE and die under continuous exposure to the RT inhibitor tenofovir disoproxil fumarate (TDF). Toxicity tests using TDF with naive C6/36 cells revealed limited effects at 0.1 mM but challenge with DENV-2 in its presence resulted in death, as expected based on previous reports. However, TDF treatment of stable, grossly normal C6/36 cell cultures persistently infected with DENV-2 for up to 30 split passages also induced severe CPE and death. The results indicate a key role of host RT not only in the establishment of persistent infections but also in their long-term maintenance. Persistent infections in shrimp and insects are sometimes characterized with low to very low levels of viral infection that are not detectable by routine histological analysis, in situ hybridization, immunohistochemistry or even by very sensitive PCR. Our results suggest that treatment of grossly normal insects or crustaceans with TDF may be a simple approach to induce higher replication by accommodated viruses. This could improve detection by histological or molecular methods and provide a way to invoke a disease state. It could also be used to increase viral quantities for purification purposes. |
