The miR-196a2 rs11614913 T > C, miR-146a rs2910164 C > G, miR-499 rs3746444 T > C, miR-26a-1 rs7372209 C > T and miR-27a rs895819 T > C genotypes were determined using a custom-by-design 48-Plex SNPscan™ Kit and matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS).
All supernatants were diluted 1 1 with the kit serum matrix diluent, following optimization experiments.
Plasma samples (25 μl) were diluted 1 1 with the kit serum matrix and added to the plate.
DNA was extracted by using a Chelex resin-based kit (InstaGene Matrix, Bio-Rad, Laboratories, Hercules, CA, USA) (21 ).
DNA for mecA-PCR, SCC mec and MLST-typing was extracted using a commercial kit (InstaGene Matrix, Bio-Rad, USA) as previously described [ 30].
C. difficile DNA was extracted from blood agar sub-cultured colonies, using a Chelex resin-based DNA extraction kit (InstaGene Matrix; Bio-Rad, USA).
Extraction of DNA was done on pure cultures after they were obtained from blood agar with a Chelex resin-based DNA extraction commercial kit (InstaGene Matrix, Bio-Rad Laboratories, USA), following the manufacturer's instructions.
At 80% confluence, cells were trypsinized (Trypsin/EDTA, Toyobo) and subcultured onto culture plates coated with human Type-1 collagen (Coating Matrix Kit, Cascade Biologics).
DNA for PCR and multilocus sequence typing analysis was prepared from 200 µl of overnight TGY culture with the InstaGene matrix kit [Bio-Rad] according to the manufacture's instructions.
