HCMV infection alters EV biogenesis machinery. Uninfected and HCMV-infected (MOI = 3) fibroblasts at 24, 48, 72, and 96 hpi were harvested for protein (A and C) and RNA (B and D) analyses. (A) Western blot analysis of the ESCRT-III subunits CHMP6, CHMP4B, and CHMP2B. Tubulin was used as a loading control. (B) qPCR of CHMP6, CHMP4B, and CHMP2B. (C) Western blot of EV markers CD63 and CD81. Tubulin was used as a loading control. (D) qPCR analysis of CD63 and CD81 for a high MOI. (E) Western blot of CD63 and CD81 from HCMV-infected (MOI = 3) fibroblasts at 24, 48, 72, and 96 hpi. qPCR results are from three independent experiments. *, P < 0.05 compared to uninfected samples.