S. pneumoniae strains used in this study are listed in Supplementary Table 1 . Thirty of them (PG01 to PG30) belong to a surveillance study done in a Nijmegen hospital, the Netherlands23 (link), while the other six are lab strains available at our lab.
Different strains were grown on tryptic soy agar or blood agar base no. 2 (Sigma-Aldrich) plates supplemented with 5% defibrinated sheep’s blood at 37 °C in a 5% CO2 atmosphere. Liquid cultures were grown statically in THY, C+Y or semi-defined minimal media (SDMM) at pH 7.3, with 5 µl ml−1 oxyrase (Oxyrase)2 (link) at the same incubation conditions as plates. For growth curve assays, strains were grown in THY until an optical density (OD)600 of ~0.5, pelleted and resuspended in the same volume of phosphate saline buffer (PBS). OD600 was adjusted to 0.05 in PBS and 20 μl of this suspension were diluted in 180 μl of different media conditions in wells of flat-bottom 96-well plates. OD600 measurements were taken on a BioSpa 8 plate reader (BioTek) and experiments were repeated at least three times. Different modifications to SDMM were assayed as described in figure legends and text.
Different strains were grown on tryptic soy agar or blood agar base no. 2 (Sigma-Aldrich) plates supplemented with 5% defibrinated sheep’s blood at 37 °C in a 5% CO2 atmosphere. Liquid cultures were grown statically in THY, C+Y or semi-defined minimal media (SDMM) at pH 7.3, with 5 µl ml−1 oxyrase (Oxyrase)2 (link) at the same incubation conditions as plates. For growth curve assays, strains were grown in THY until an optical density (OD)600 of ~0.5, pelleted and resuspended in the same volume of phosphate saline buffer (PBS). OD600 was adjusted to 0.05 in PBS and 20 μl of this suspension were diluted in 180 μl of different media conditions in wells of flat-bottom 96-well plates. OD600 measurements were taken on a BioSpa 8 plate reader (BioTek) and experiments were repeated at least three times. Different modifications to SDMM were assayed as described in figure legends and text.