Bacterial DNA Extraction from Swabs

Bacterial DNA was extracted from the swabs using a previously described bead beating method^{25 (link)} with the following modifications: The swabs were vortexed in 0.5 ml of sterile ice-cold PBS, of which 175 μL was combined with 235 μL of RBB lysis buffer (500 mM NaCl, 50 mM Tris-HCl (pH 8.0), 50 mM EDTA, 4% SDS) in a bead beating tube. The samples were bead beaten using a FastPrep-24 instrument at 5.5 m/s (MP Biomedicals, Inc., USA) with 0.1 mm zirconium-silica beads (Biospec Products, Bartlesville, OK, USA) for 1 min. Samples were then heated at +95 °C for 15 min with shaking 400 rpm and centrifuged at room temperature for 5 min at 13 000 rpm. The supernatant (200 μL) was used for DNA extraction with KingFisher Flex automated purification system (ThermoFisher Scientific, USA) and Ambion Magma Total Nucleic Acid Isolation Kit (Life Technologies, USA) using MagMAX Pathogen High Vol Duo program. DNA was quantified using Quanti-iT Pico Green dsDNA Assay (Invitrogen, San Diego, CA, USA). An aliquot of the DNA extract was sent to Karolinska Institutet, Sweden for Human papillomavirus (HPV) genotyping^{49 (link)}.