Figure 8
Measurement of KOS and 0--GFP viral genome loads in the trigeminal ganglia of HSV-1 latently infected mice. A. Dotblot of HSV-1 VP16 PCR products. Each "dot" contains VP16 PCR product amplified from the TG DNA of a single mouse, and the n-values indicate numbers of mice per group. TG harvested from uninfected (UI) mice served as negative controls for the PCR. TG harvested from mice dying of encephalitis (Day 9 p.i.) belonged to one of the following groups: rag2-/- ifnar-/- mice, ifnar-/- ifngr-/- mice, or stat1-/- mice inoculated with 2 × 104 pfu per eye of 0--GFP. TG harvested from mice that were latently infected with HSV-1 (Day 40 p.i.) belonged to one of the following groups: strain 129 mice inoculated with 2 × 105 pfu per eye of KOS; strain 129 mice, ifngr-/- mice, or ifnar-/- mice inoculated with 2 × 105 pfu per eye of 0--GFP; or stat1-/- mice inoculated with 2 × 104 pfu per eye of 0--GFP. The standard curve on the right consists of PCR products amplified from a two-fold dilution series of VP16 plasmid DNA. B. The ratio of yields of VP16 to competitor PCR product yields (competitor dotblot not shown) was used to estimate viral genome copy number per PCR. The logarithm of viral genomes per TG, y, was plotted as a function of the mean logarithm of the ratio of VP16 PCR product yield: competitor PCR product yield, x, amplified from duplicate PCRs of each dilution of VP16 plasmid (error bars indicate the standard deviation between duplicate PCRs). The relationship between viral genome load and PCR product yields was described by the equation, y = 0.2556•x3 + 0.1055•x2 + 1.2079•x + 5.9309 (r2 = 0.99). The number of HSV-1 genomes per TG in each sample was derived from fitting the data shown in panel A to the standard curve shown in panel B. C. Number of HSV-1 genomes per TG in mice that were uninfected or were latently infected with KOS or 0--GFP. The dashed line indicates the lower limit of detection of the PCR assay. Asterisks denote significant differences in viral genome load per TG relative to strain 129 mice latently infected with KOS (p < 0.05, two-way t-test).