HSV -1 and HSV -2 are refractory to the antiviral activity of semen exosomes. (A and B) Infectivity of HSV-1 or HSV-2 on HEp-2 cells was determined by counting plaque forming units (PFU) two days post infection with 2 X 105 PFU of HSV-1 or HSV-2 pre incubated for 1 h at 37°C with PBS vehicle (gray bar), 100 μg/ml semen exosomes or 100 μg/ml blood exosomes (white bar). Virus titre is shown as particle forming unit (PFU) per ml. (C and D) HEp-2 cells were preincubated at 37°C for 1 h with PBS vehicle (gray bar), 100 μg/ml SE or 100 μg/ml BE (white bar) followed by challenge with HSV-1 or HSV-2 (MOI =5) at 37°C for 1 h. Viral replication was allowed to ensue for 18 h. Progeny virus was collected and titered on Vero cells for determination of virus titre by plaque assay. Data are mean±SD and are reported as plaque forming units (PFU). Student's t test was used for all samples; significance was taken at P ≤0.05. ns = not significant. Experiments were repeated at least 3 times with similar results. (E) Murine splenocytes are susceptible to SE-mediated inhibition of LP-BM5 replication ex vivo: Naïve splenocytes were infected with LP-BM5 pre-incubated with vehicle or SE. 96 h post challenge, total DNA was isolated and used to evaluate viral load by quantitative PCR detection of proviral DNA. Results were normalized for GAPDH. Data are mean±SD expressed as fold change of vehicle set at 1. *P <0.02, Student's t test was used for all samples. Experiments were repeated at least three times with similar results.