Culturing Oral Pathogenic Bacteria

Streptococcus oralis (ATCC 9811, American Type Culture Collection, Manassas, VA, USA), Aggregatibacter actinomycetemcomitans (DSM 11123, German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany), and Porphyromonas gingivalis (DSM 20709) were cultured on tryptone soy agar (TSA) or fastidious anaerobe agar (FAA) plates supplemented with 5% sheep blood at 37 °C under anaerobic conditions (80% N₂, 10% H₂, 10% CO₂). Few colonies of S. oralis were inoculated into tryptone soy broth (TSB) (Oxoid Limited, Hamsphire, United Kingdom) supplemented with 10% yeast extract (Carl Roth GmbH + Co. KG, Karlsruhe, Germany) and 50 mM glucose (Carl Roth GmbH + Co. KG, Karlsruhe, Germany) at 37 °C under constant shaking. A few colonies of A. actinomycetemcomitans or P. gingivalis were inoculated overnight into brain heart infusion medium (BHI; Oxoid, Wesl, Germany) supplemented with 10 μg/mL vitamin K (Roth, Karlsruhe, Germany) under anaerobic conditions [21 (link)]. Treponema denticola (DSM 14222) cultures were prepared in new oral spirochete (NOS) medium at 37 °C for 72 h under static anaerobic conditions [22 (link)].