Bacterial Species Identification via MALDI-TOF MS and WGS

Prior to WGS, a primary species identification from single colonies was performed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) (Bruker, Billerica, MA, United States) and MBT Compass IVD software 4.1.60 (Bruker) as described by Halbedel et al. (2020) (link).
As for WGS, DNA was isolated from bacterial cultures with the MagAttract HMW DNA Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions for Gram-positive bacteria. The amount of input DNA was quantified on a Lunatic instrument (Unchained Labs, Pleasanton, CA, United States). Nextera XT chemistry (Illumina Inc., San Diego, CA, United States) was used to prepare sequencing libraries for a 300 bp paired-end sequencing run on an Illumina MiSeq sequencer. Samples were sequenced to achieve a minimum 80-fold coverage using recommended standard protocols by Illumina. The resulting FASTQ files were quality trimmed and de novo assembled with the SPAdes version 3.9.0. Contigs were filtered for a minimum of fivefold coverage and 200 bp minimum length with SeqSphere+ software v. 7.8.0 (Ridom, Münster, Germany) (Jünemann et al., 2013 (link)).