2.3 Whole Genome Sequencing Analysis
Genomic DNA of bacterial isolates was extracted using the DNeasy Blood
& Tissue Kits (Qiagen, Germany) according to the manufacture’s
instruction. Qubit® 3.0 fluorometer (Invitrogen, USA) was used to
measure the DNA concentration. A total amount of 0.2 μg DNA was then
used as input material for the DNA library preparations using NEB
Next® UltraTM DNA Library Prep Kit
for Illumina (NEB, USA), and subsequently sequenced using the 2 × 150 bp
paired-end library on HiSeq 2500 sequencing system (Illumina). Following
trimming and filtering by the NGS_QC Toolkit (v2.3.3), the raw reads
were subjected to de novo assembly by SPAdes 3.6 (Bankevich et
al., 2012). The subsequent annotation of the assembled genome was
performed by Prokka version 1.12 (Seemann, 2014). Plasmid were
identified using Plasmid Finder 2.1
(https://cge.cbs.dtu.dk/services/PlasmidFinder/). The
antimicrobial resistance genes and chromosomal mutations were analyzed
using ResFinder 3.2 (Zankari et al., 2012). CRISPR typing of the
isolates was performed as previously described (Li, 2021).