85 Genetic determinants in MRSA carriage 5 were prepared using the Nextera XT v2 kit (Illumina, San Diego, CA, US) according to the manufacturer’s instructions. Short-read sequencing was performed with an Illumina MiSeq System generating paired-end reads of 250 bp. De novo assembly of paired-end reads was performed using CLC Genomics Workbench v12.0.1-v20.0.4 (QIAGEN, Hilden, Germany) after quality trimming (Qs≥20) establishing a word size of 29. Based on next generation sequencing data (ENA project number PRJEB59407), molecular typing was performed using Ridom Seqsphere+v8.3.1 (Ridom, Münster, Germany). Herewith multilocus sequence typing (MLST) ST type was derived and core genome multilocus sequence typing (cgMLST) was performed using a scheme including 1861 alleles [19]. Isolates with a maximum of 24 allelic differences were denominated the same complex type. Antibiotic resistance genes were identified by Resfinder v4.1 (Center for Genomic Epidemiology, Lingby, Denmark). A predefined set of virulence factors were identified using AlereMicroarray schemes in Ridom Seqsphere+v8.3.1 (Ridom, Münster, Germany) [20]. Definitions Uncomplicated MRSA carriage was defined as having all of the following features: (i) the presence of MRSA exclusively located in the nose, (ii) no active infection with MRSA, (iii) in vitro susceptibility for mupirocin, (iv) the absence of active skin lesions, (v) the absence of foreign material that connects an internal body site with the outside (e.g., urine catheter, external fixation material), and (vi) no previously failure of decolonization treatment. All other carriage cases were considered complicated colonization. Uncomplicated carriage is advised to be treated with topical therapy (mupirocin topically applied to the nares, disinfecting shampoo) and hygienic measures. In cases of complicated MRSA carriage, additional systemic antimicrobial therapy with a combination of two antibiotic agents is recommended, according to the national guideline [16]. MRSA infection was defined as a positive culture send to the microbiology laboratory from an infected body site as indicated by the treating physician. Successful decolonization was defined as three consecutive negative MRSA cultures from swabs taken from nose, throat, and perineum, with the cultures obtained at 1-week intervals, without antibiotic usage [16]. For analyses, patients were divided in two groups: patients with failure of eradication treatment (failure group) and patients with successful decolonization with or without preceding treatment (successful decolonization group).
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