Identibac microarray

We extracted genomic DNA from S. aureus isolate UKM4229 using the QIAamp DNA Mini Kit (QIAGEN, Hilden, Germany) according to the manufacturer’s instructions. We isolated plasmid DNA with the PrepEase MiniSpin Plasmid Kit (Affymetrix USB, Santa Clara, CA, USA) following the protocol standards. For both plasmid and genomic DNA, we applied lysostaphin (20 µg/mL) (Wakchemie, Steinbach, Germany) for bacterial cell lysis. We performed multilocus sequence typing and spa gene typing initially as described elsewhere (25 (link),26 (link)) and confirmed our results later by analysis of whole-genome sequencing (WGS) and DNA microarray data (discussed later in this article). We analyzed DNA sequences using RidomStaphType and SeqSphere+ (Ridom GmbH, Münster, Germany). Applying DNA microarray analysis (IdentiBAC Microarray; Alere Technologies GmbH, Jena, Germany), we identified resistance and virulence determinants and checked genotyping results.

MLST was done based on the WGS data applying the PubMLST database and software available on PubMLST.org/saureus/ (Jolley and Maiden, 2010 (link)). Typing of the spa gene was performed with spaTyper 1.0 available from the Center for Genomic Epidemiology homepage (Bartels et al., 2014 (link)) also using whole-genome data. Verification of MLST and spa typing and determination of regulatory and hemolysin genes were performed at the genetic level using DNA microarray analyses (IdentiBAC Microarray, Alere, Jena, Germany).