Bacterial strains and culture conditions.

The bacterial strains and plasmids used in these studies are listed in Table S1 in the supplemental material. E. coli strains were grown in LB broth or agar with or without 100 μg/ml ampicillin, 250 μg/ml erythromycin, 20 μg/ml chloramphenicol, 30 μg/ml of kanamycin, or 12 μg/ml tetracycline, as required. NTHi strains were grown in brain heart infusion broth or agar supplemented with 10 μg/ml heme-histidine and 10 μg/ml β-NAD (sBHI) or on chocolate agar with 5 U/ml bacitracin. NTHi was routinely cultured at 37°C with 5% CO₂. To construct the ectopic strains, transformants were selected on chocolate agar plates with 5 μg/ml erythromycin, and single colonies were passaged for 3 days on selection plates before being confirmed by DNA sequencing of PCR-amplified purified genomic DNA.