Thymidine kinase-expressing yellow fever 17D reporter virus facilitates prodrug activation and bioorthogonal labelling of infected cells

Tracking of viral replication and tissue tropism by in vivo imaging can help to unveil how live-attenuated vaccines such as the yellow fever 17D (17D) work, and likewise, to understand how adverse effects develop. Here we validate 17D-TK, a reporter virus derived from 17D that expresses herpes virus thymidine kinase (TK) that specifically converts nucleoside analogues such as Ganciclovir (GCV) to induce cell death, or difluoro-EdU (dF-EdU) for bioorthogonal labelling of infected cells by Click chemistry. 17D-TK induces a cytopathic effect in infected cell cultures, as well as mortality in intracranially inoculated mouse pups in a GCV dependent manner. Preferential phosphorylation of difluoro-EdU (dF-EdU) in 17D-TK infected cells allows to selectively stain cells that support 17D replication. Prospectively, 17D-TK can be used in combination with radiolabeled tracers for real-time detection and localization of sites of active viral replication in living animals using positron emission tomography (PET).