Five Complete Genome Sequences Spanning the Dutch Streptococcus suis Serotype 2 and Serotype 9 Populations

ABSTRACT

ANNOUNCEMENT

Streptococcus suis is an opportunistic pathogen in pigs which can cause zoonotic infections. Human infections are predominantly caused by S. suis serotype 2 (1) and can lead to septicemia and meningitis (2). We recently identified a zoonotic S. suis serotype 2 clone belonging to clonal complex 20 (CC20), which emerged from a nonzoonotic serotype 9 CC16 clone (3) in the Netherlands. To facilitate further research on the zoonotic potential of S. suis, we sequenced the genomes of S. suis serotype 9 CC16 and CC20 strains, isolated from diseased pigs, and three serotype 2 strains, including strain S10 (CC1, pig) and two CC20 strains, one each from human and porcine infections (Table 1). Data were generated using Illumina and Nanopore MinION sequencing technologies.

S. suis was grown overnight in Todd-Hewitt broth supplemented with yeast extract (THY), and genomic DNA was isolated using the Qiagen MagAttract high-molecular-weight (HMW) DNA extraction kit. The sequence library was constructed using the native barcoding (catalog number EXP-NBD114) and ligation sequencing (catalog number SQK-LSK109) kits (Oxford Nanopore). DNA was repaired and A tailed using NEBNext formalin-fixed, paraffin-embedded (FFPE) DNA repair mix and the NEBNext Ultra II end repair/dA-tailing module (New England BioLabs). A barcode was ligated to the A-tailed DNA using blunt/TA ligase master mix (New England Biolabs). Sequence adapters were ligated to barcoded samples pooled by equal mass with Quick T4 DNA ligase (New England BioLabs). The library was loaded on the flow cell (FLO-MIN106D [R9]) and sequenced using MinKNOW fast base calling version 3.5.5. Default parameters were used for all tools except where noted otherwise. Illumina data were available from our previous study (Table 1) (3).

Illumina read filtering was performed using fastp version 0.20.0 (4). MinION reads were filtered for quality and length using Filtlong version 0.2.0 (5), using the filtered Illumina reads as reference. FastQC version 0.11.8 was used for quality control (6). Illumina and MinION reads were used in a hybrid assembly using Unicycler version 0.4.8, which also performs assembly trimming, circularizing, and rotating (7). Assembly statistics were collected using Quast version 4.6.3 (8). Coverage was assessed using Minimap2 version 2.17 (9), SAMtools version 1.9 (10), and BEDTools version 2.29.0 (11). The complete genomes were annotated using Prokka version 1.14.0 (12). Multilocus sequence typing (MLST) was performed using mlst version 2.17.6 (13). For workflow management, Snakemake version 5.7.1 (14) was used. The pipeline is freely available from https://github.com/boasvdp/MRA_Streptococcus_suis.

Genomes of all five strains consisted of a single chromosome ranging from 2,042,889 to 2,292,626 bp with a GC content of 41.10 to 41.43% and a coverage of 23 to 72×, determined using Nanopore data (Table 1).

Draft assemblies of the five strains were 46 to 74 kbp smaller than the complete genomes. Mapping the draft genomes to the complete genomes revealed no missing regions in the draft genomes. The draft genomes are likely smaller than the complete genomes due to the collapse of repeats, which has been described before (15).

ACKNOWLEDGMENTS

REFERENCES