Figure 1. Single and mixed infection of human monocyte derived-macrophages (MDMs) by IAV and SP.

(a) MDMs were infected with IAV at an MOI of 0.01, 0.1 or 1 and total RNA was extracted at 0, 6, 18, 24, 48 and 72 hours post-infection. The IAV A/H1N1 hemagglutinin (HA) gene was amplified by real-time qRT-PCR. (b) Influenza A/H1N1 viral load (TCID₅₀) in the supernatant of single IAV-infected MDMs or IAV-infected MDMs following subsequent challenge with SP. (c) Confocal imaging of IAV and SP in mixed-infected MDMs at 8 hours post-IAV infection and 4 hours post-SP infection. DAPI, nuclear stain, blue. IAV, Influenza A virus nucleoprotein stain, green. SP, S. pneumoniae stain, red. Scale bar = 5 μM. (d) Impact of single and mixed IAV and SP infection on MDM cell viability. Statistical analyses were performed using two-way ANOVA with Tukey’s post-hoc test; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.