Genome Sequencing and Comparative Analysis

Genomic DNA was extracted using the EZ1 biorobot and the EZ1 DNA tissue kit (Qiagen) and then sequenced on a MiSeq sequencer (Illumina, San Diego, France) with the Nextera Mate Pair sample prep kit (Illumina) and Nextera XT Paired End (Illumina), as previously described [46 (link)]. The assembly was performed using Spades V. 3.15 [47 (link)] and trimmed (using Trimmomatic v. 0.36) [48 (link)]. Scaffolds of < 800 bp and scaffolds with a depth value < 25% of the mean depth were removed. The best assembly was selected using criteria such as the number of scaffolds, N50, and number of N. All assembled genomes were annotated using Prokka v 1.14.5 (Fontainebleau, France) [49 (link)]. The gff3 output files were used to construct a core genome alignment using Roary v3.13.0 (USA) using default parameters [50 (link)]. Comparative analysis was performed for the five placental isolates and reference strains such as RSA493 (Genbank: NC_002971.4), Z3055 9Genbank: NZ_LK937696), NL3262 (Genbank: NZ_CP013667), and Guiana (Genbank: HG825990.30). The phylogenetic tree was reconstructed by using FastTree (Berkeley, CA, USA) [51 (link)].