Fig 1: Verification of acyclovir-resistant (ACVR) HSV-1 infection in Vero cells. (A) Flow chart of the methodology used to create ACV-resistant (ACVR) mutants, adapted from Sarisky et al. [33]. Verification of acyclovir-resistant (ACVR)-GFP-HSV-1 17+ in Vero cells. (B) Fluorescent images of 0.1 MOI GFP-HSV-1 infection in Vero cells with or without 45 µM ACV. FITC filter with excitation of 490 nm and emission of 525 nm was used to visualize viral plaques at 40× magnification; Scale bar represents 200 µm. (C) Fluorescent images of 0.1 MOI ACVR-GFP-HSV-1 infection in Vero cells with or without 45 µM ACV. FITC filter with excitation of 490 nm and emission of 525 nm was used to visualize viral plaques at 40× magnification. Scale bar represents 200 µm. (D) Plaque Assays of viral stocks (1 MOI) with or without 45 µM ACV in Vero cells in viral dilutions of 10−2 to 10−4, shown as qualitative image and in a table with averaged viral titers. (E) Western Blot analysis using antibodies directed against HSV-1 infected cellular protein 0 (ICP0; Santa-Cruz, sc-53070), glycoprotein D (gD; Santa-Cruz, sc-69802), thymidine kinase (TK; Invitrogen, PA5-67984), and GAPDH (0411; Santa-Cruz, sc-47724). M: Chameleon 800 Pre-stained Protein Ladder (Li-Cor, 928-80000). Signaling intensities of bands for each antibody have been normalized (as a percentage) to GFP-HSV-1. ACVR: ACVR-HSV-1. (F) Plaque Assays of GFP-HSV-1 17+ (0.1 MOI) and ACVR- HSV-1 (0.1 MOI) with various treatments in Vero cells such as DMSO (1:2000), 10 µM GA, and 10 µM ACV in viral dilutions of 10−2 to 10−4, shown as qualitative image and in a table with averaged viral titers.