Seqsphere software v 7

A total of 3,678 targets were used to establish the core genome multilocus sequence typing (cgMLST) scheme of Cronobacter spp. using strain ATCC BAA-894 as a reference using Ridom SeqSphere+ software v. 7.8.0 (Ridom, Münster, Germany; Jünemann et al., 2013 (link)). For Salmonella, the cgMLST scheme was performed based on the profile of 2,969 S. enterica target gene loci task template of the Ridom SeqSphere+ software v. 7.8.0 (Ridom, Münster, Germany). According to the cgMLST scheme, isolates were visualized with a minimum spanning tree (MST) to establish their genotypic relationships (Lepuschitz et al., 2019 (link)). In addition, the sequences of the seven housekeeping genes of the conventional MLST for Cronobacter spp. and Salmonella were extracted and cross-checked against the Cronobacter MLST database¹ (Baldwin et al., 2009 (link)) and Salmonella MLST² (Achtman et al., 2012 (link)), respectively. The Cronobacter strains are ID 3409–3413 in the Cronobacter PubMLST database and Salmonella are ID RID389119–RID389124 in the cgMLST database.

Before WGS, all the Cronobacter spp. and Salmonella spp. strains were cultured in Columbia blood agar plates (bioMérieux, Marcy-l’Étoile, France) at 37°C for 24 h. DNA was isolated from bacterial cultures with the MagAttract HMW DNA Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. The amount of 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 2 × 300 bp paired-end sequencing run on an Illumina MiSeq sequencer. Samples were sequenced to achieve a minimum of 80-fold coverage using 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 5-fold coverage and 200 bp minimum length with Ridom SeqSphere+ software v. 7.8.0 (Ridom, Münster, Germany; Jünemann et al., 2013 (link)).