Draft genome sequences of three Carnobacterium maltaromaticum isolates from two low pH bogs in Denmark

TL;DR

This paper presents draft genomes of three Carnobacterium maltaromaticum bacteria isolated from acidic bogs in Denmark.

Contribution

The novelty is the sequencing and reporting of three new draft genomes from C. maltaromaticum isolates in low pH bogs.

Findings

Three draft genome sequences of C. maltaromaticum were obtained from low pH bogs in Denmark.

The Illumina MiSeq platform was used for genome sequencing.

Abstract

Carnobacterium maltaromaticum is a lactic acid bacterium that is widely distributed in the environment, including freshwater. Here, we report the draft genome sequences of three C. maltaromaticum isolates from low pH Danish bogs, using the Illumina MiSeq platform.

Figures

A maximum-likelihood phylogenetic tree based on SNP variants in the whole core genomes of the Carnobacterium spp. strains. C. maltaromaticum ATCC 35586 was used as the reference genome to map and screen the SNPs.

Tables

TABLE 1. Sampling location description and genome characteristics for the three C. maltaromaticum isolates

Variable	s2-1	s3-1	s6-2
Location	Site 1	Site 2	Site 2
Type	Forest bog	Forest bog	Forest bog
pH	4.35	4.40	4.30
Geographic coordinates	55°45'12.0"N 12°30'17.3"E	55°49'35.0"N 12°33'40.5"E	55°49'34.8"N 12°33'40.4"E
Genome characteristics
Assembly size (bp)	3,728,548	3,498,272	3,930,617
N50 (bp)	181,003	188,278	246,871
L50 (bp)	7	6	8
GC (%)	34.10	34.34	34.14
Number of contigs	73	47	76
Number of CDS	3,372	3,141	3,566
Number of tRNAs	57	63	62
Number of rRNAs	8	7	12
Number of sequences	645,632	449,296	222,660
Total number of reads (bp)	193,689,600	134,788,800	66,798,000
Genome coverage	55×	38×	19×
Bacteriocins a	BM1	BM1	None
BioProject accession	PRJNA990564	PRJNA990564	PRJNA990564
BioSample accession	SAMN37661217	SAMN37661256	SAMN37661257
SRA accession for raw reads	SRR26265303	SRR26265302	SRR26265301
GenBank accession	JAWLUX000000000	JAWLUY000000000	JAWLUZ000000000

Full text

ANNOUNCEMENT

The lactic acid bacterium Carnobacterium maltaromaticum is widely distributed in Danish freshwater environments during the winter season but underrepresented in bogs and poor fens (<pH 5.15) (1). Here, we show that C. maltaromaticum could be readily isolated from such habitats when culture-dependent sampling was done during the summer season and by using an enrichment procedure including an increased volume of sample relative to enrichment medium.

Ten milliliters of water samples were obtained on 30 May 2023, from two sites (Table 1) (1). One or three milliliters of samples were mixed 1:1 with all purpose Tween (APT) broth (Difco, Sparks, MD, USA) and incubated anaerobically at 15°C for 3 d before plating on de Man, Rogosa, Sharpe (MRS) agar (Oxoid, Thermo Fisher Scientific, Waltham, USA) anaerobically at 15°C for 3–5 d. Single colonies of Gram-positive and catalase-negative isolates (s2-1 from site 1; s3-1 and s6-2 from site 2) were cultivated in 3-mL APT broth under anaerobic and static conditions at 15°C for 3 d. Bacterial cells were harvested by centrifugation, and DNA extraction was performed using a DNeasy Blood and Tissue Kit, according to the manufacturer’s instructions (Qiagen GmbH, Hilden, Germany). DNA concentrations were measured by NanoDrop One Spectrophotometer (Thermo Fisher Scientific). Library preparation was done using the Nextera DNA Flex Library Prep Kit (Illumina, San Diego, USA). Sequencing on an Illumina MiSeq platform was performed as described by Li et al. (2), generating paired-end reads of 2 × 300 bp. An initial quality control involved FastQC v.0.11.9 (3) and trimming low-quality bases (Q <20) and adapter sequences with Cutadapt v.4.4 (4). Trimmed reads were assembled using SPAdes v.3.13.1 (5) with assembly quality assessed by QUAST 5.2.0 (6). Genome annotation was performed with Prokka v. 1.13 (7). Within the assembled contigs, the 16S rRNA gene regions were identified using Barrnap v.0.9 (8). The corresponding sequences were extracted using Bedtools v2.30.0 (9). Identification of the isolates was performed by comparing their 16S rRNA gene sequences via BLAST against the NCBI 16S rRNA database. A phylogenetic tree based on single-nucleotide polymorphism (SNP) variants in the whole core genomes was constructed using sequences of the 3 isolates from this study, 28 freshwater isolates of C. maltaromaticum from our previous study (BioProject: PRJNA990564, BioSample: SAMN36274541 to SAMN36274568) (1) and 19 published genome sequences of C. maltaromaticum, Carnobacterium gallinarum, and Carnobacterium sp. retrieved from the NCBI database. SNPs were identified by mapping reads against the reference C. maltaromaticum ATCC 35586 genome (GCF_000238575.1) via CSI Phylogeny v1.4 (10) and visualized in ChiPlot (11). Finally, we probed the presence of genes encoding bacteriocins by AntiSMASH 6.0 (12) and BAGEL 4 (13). All bioinformatic tools were run with default parameters.

Sequence statistics of the C. maltaromaticum genomes are summarized in Table 1, and a phylogenetic tree shows the close relatedness of the three isolates from this study to other C. maltaromaticum strains (Fig. 1). Genes encoding carnobacteriocin BM1 and the corresponding immunity protein were found in the s2-1 and s3-1 genomes. No genes encoding structures of other bacteriocins were detected.

A maximum-likelihood phylogenetic tree based on SNP variants in the whole core genomes of the Carnobacterium spp. strains. C. maltaromaticum ATCC 35586 was used as the reference genome to map and screen the SNPs.