Illumina short read sequencing was performed for all INF isolates (2–10 per infection, 23 infections, n = 194), CloNo isolates (n = 30) and nCloNo isolates (n = 62).

Strains were inoculated into 3mL of TSB from blood agar plates and incubated at 37°C under vigorous shaking for 4-6h. Cells were pelleted, washed with phosphate-buffered saline and mechanically lysed with zirconia beads 3x20s on a tissue homogenizer (Precellys 12, Bertin, Montigny-le-Bretonneux, France). DNA was then extracted with the QIAamp Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. DNA was then fragmented on a Bioruptor Pico instrument (Diagenode, Seraing, Belgium) to a fragment length of approximately 300-400nt. DNA libraries were constructed with the NEB Next Ultra DNA library Prep Kit for Illumina (New England Biolabs, Ipswich, MA, USA) and whole genome sequencing was conducted on a NextSeq 500 sequencing system and a 300-cycle mid-output kit (Illumina, San Diego, CA, USA). A mean of 4.3 million paired-end, 150 nt reads were generated. Bases less than Q30, as well as adapter sequences of the reads, were trimmed and any reads shorter than 35 nt were removed using Trimmomatic v0.36 [102].Retained high-quality NextSeq reads were used as input for the SPAdes assembler (version 3.7.1) [103], resulting in a mean depth of 250.3 (max: 1487.9, min: 10.6) and an N50 contig length of 123 kb.

Long-read Nanopore sequencing (Oxford Nanopore technologies, Oxford, UK) was performed for at least one isolate per infection and all isolates with SCCmec or ACME rearrangements (HD04-1, HD05-1, HD12-1, HD17-1, HD21-2, HD21N4, HD25-1, HD26-1, HD26-2, HD27-2, HD27N1, HD29-1, HD31-1, HD33-1, HD33-3, HD39-1, HD40-1, HD46-1, HD47-1, HD59-1, HD66-1, HD66-6, HD75-1, HD99-1, HD99-4, HD104-2). High-molecular weight DNA was extracted with the QIAamp Mini Kit (Qiagen, Hilden, Germany). Concentration was assessed on a Qubit fluorometer (Thermo Fisher Scientific, Waltham, MD, USA) and purity was assessed on a NanoDrop photometer (Thermo Fisher Scientific, Waltham, MD, USA). Native barcoding for multiplexing and PCR free ligation based ONT library preparation was performed according to manufactures protocol (Oxford Nanopore Technologies, Oxford, UK). Sequencing was done on ONT GridIOn X5 R9.4.1 flowcells. Base-calling was done with guppy version version 3.3.0 (https://community.nanoporetech.com, Oxford Nanopore Technologies, Oxford, UK). A hybrid assembly of the Illumina short-read sequencing data and Nanopore data was conducted with Unicycler version 0.4.7 [104]. All genomes with long- and short read data were successfully closed and submitted to NCBI Genbank (S3 Table for Accession numbers). Annotation of all genomes was conducted with the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) [105] upon submission to Genbank.

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