Draft Genome Sequence of Bacillus cytotoxicus CVUAS 2833, a Very Close Relative to Type Strain NVH 391-98 Isolated from a Different Location.

We report the draft genome sequence of Bacillus cytotoxicus CVUAS 2833, isolated from potato puree in Germany (2007), which is-despite its clearly different source-very similar to the type strain B. cytotoxicus NVH 391-98 isolated in France (average nucleotide identity, 99.5%).

GENOME ANNOUNCEMENT

The highly enterotoxic Bacillus cereus NVH 391-98 was discovered in vegetable puree, which caused severe food poisoning including fatalities (France, 1998) (1). In this strain, a new cytotoxin CytK-1 was identified. Subsequently, the distantly related homolog CytK-2 was discovered in other strains of the B. cereus group which display considerably attenuated toxicity (2). B. cereus NVH 391-98 was published in 2013 as the type strain of the new species Bacillus cytotoxicus due to the presence of the cytK-1 gene, its thermotolerance (growth at up to 50°C), a distinctive fatty acid profile, DNA-DNA hybridization, and multilocus sequence typing (MLST) (3). B. cytotoxicus CVUAS 2833, which is one of only five known B. cyototoxicus strains, was isolated spatially and chronologically separate, but affiliated to B. cytotoxicus by 16S rRNA gene sequence and MLST comparison (3, 4). To date (June 2015), only the genome sequence of the type strain has been available.

We sequenced B. cytotoxicus CVUAS 2833 on the Illumina MiSeq platform and obtained a library of 3,185,212 reads. About 72.2% of the reads passed quality filtering (length, ≥80%; Phred score, Q ≥ 30) and were assembled to 36 contigs using ABySS version 1.3.7 with an assembly size of 4,127,075 bp and a G+C content of 35.74%. The assembled contigs were submitted to GenBank and annotated by NCBI’s prokaryotic genome annotation pipeline (PGAP). BLAST analysis revealed the presence of the distinct B. cytotoxicus variant of the enterotoxin operon nhe (5) in strain CVUAS 2833 with only a single nucleotide difference to NVH 391-98 nhe. B. cytotoxicus CVUAS 2833 does not contain the ces cluster encoding the emetic toxin or the hbl enterotoxin genes. Whole-genome pairwise average nucleotide identity (ANI) of 99.5% supports the exceptionally high similarity to B. cytotoxicus NVH 391-98. Additionally, Gegenees 2.2.1 was used to compare whole-genome similarity in a fragmented alignment (BLAST 2.2.29+) at a fragment size of 200 bp and a comparison step size of 100 bp (6). In contrast to ANI, this approach applies stricter settings (7) and does not exclude dissimilar sequences (8). B. cytotoxicus NVH 391-98 and CVUAS 2833 showed 93.5% identity in the fragmented all-all comparison.

These genomic and genetic similarities confirm a clonal character of the B. cytotoxicus lineage within B. cereus sensu lato. All known B. cytotoxicus strains originate from different food sources (1, 9), but are similar on a MLST basis (3). The only well-analyzed clonal phylogenetic cluster in B. cereus sensu lato is Bacillus anthracis. The B. anthracis linage is evolutionarily young and demarcation of B. anthracis at a species level is quite debatable (10, 11). In contrast, B. cytotoxicus seems to be a much older and clearly discernible branch within B. cereus sensu lato (12).

Nucleotide sequence accession numbers.

The draft genome sequence of B. cytotoxicus CVUAS 2833 has been deposited at DDBJ/EMBL/GenBank under the accession no. JYPG00000000. The version described in this paper is version JYPG01000000.