European genotype PRRSV downregulates PAM phagocytic capacity, depending on the number of internalized virions. Flow cytometric analysis of the effect of European genotype PRRSV entry on PAM phagocytosis and confocal microscopical analysis of the number of internalized PRRSV virions per PAM for Marc-145- and macrophage-grown PRRSV (LV) (Black circle) or PRRSV (Lena) (Grey circle): (A–B) The effect of both PRRSV strains on phagocytosis of beads by viable macrophages, expressed as the percentage of viable cells that have beads associated with them (A) or the MFI per viable cell (B), which is a measure for the amount of beads per cell. PAM were inoculated with both PRRSV strains at different moi as indicated and infection was allowed for 1 h, after which a phagocytosis assay was performed. Mock infected cells were included as a positive control. In addition, a phagocytosis assay was performed at 4°C (dotted line) after mock infection, to assess the percentage of macrophages that have beads bound to their surface and their MFI. Data represent the mean ± SEM of 3 independent experiments; (C) PAM were inoculated with both PRRSV strains (moi = 1) and further incubated for 1 h, after which the number of virions per PAM was determined by an immunofluorescence staining. At least 20 cells were analyzed per sample. Data represent the mean ± SEM of 1 representative experiment. *p < 0.05; **p < 0.01; ***p < 0.001.