Abstract
Bacillus cereus sensu lato is a species complex that includes the anthrax pathogen Bacillus anthracis and other bacterial species of medical, industrial, and ecological importance. Their phenotypes of interest are typically linked to large plasmids that are closely related to the anthrax plasmids pXO1 and pXO2. Here, we present the draft genome sequences of 94 isolates of B. cereus sensu lato, which were chosen for their plasmid content and environmental origins.
GENOME ANNOUNCEMENT
Bacillus cereus sensu lato is a species complex that groups six classically described species of ubiquitous Gram-positive spore-forming bacteria, including the eponymous B. cereus sensu stricto, the entomopathogen Bacillus thuringiensis, the rhizoid-looking Bacillus mycoides and Bacillus pseudomycoides, and Bacillus anthracis, the etiological agent of anthrax.
The members of this group were originally distinguished on the basis of their phenotypic differences, but over the past decade, advances in the understanding of the phylogenomics of this group have largely invalidated this classification. Instead, the members of the B. cereus sensu lato group are more appropriately viewed as forming one single species from which different ecotypes and pathotypes emerge in a dynamic fashion, leading in some cases to the formation of clonal complexes with specific phenotypes (1–6).
Many phenotypic properties that are specific to these ecotypes and pathotypes are directly related to the presence or absence of large plasmids that carry genes associated with those phenotypes. In the case of B. anthracis, the virulence plasmids pXO1 (192 kb) and pXO2 (96 kb) carry the anthrax toxin and capsule genes, respectively, as well as the associated regulatory elements (7). Furthermore, the large plasmids found in a number of previously sequenced B. cereus sensu lato strains of medical or industrial interest were observed to share a backbone with either the pXO1 or pXO2 anthrax plasmids. For example, in strains of B. cereus sensu stricto that are responsible for the B. cereus-associated emetic food poisoning syndrome, the genes encoding the emetic toxin cereulide are carried by a large plasmid that shares a common genetic backbone with the pXO1 anthrax plasmid (8, 9).
We have shown previously that select sequences of the shared pXO1 and pXO2 backbones can be found widely in environmental isolates of B. cereus sensu lato (10). We postulated that these are found in plasmids that are genetically related to the pXO1 and pXO2 plasmids (hence called pXO1-like and pXO2-like, respectively) and may play an important role in the ecotypic and pathotypic differentiation of B. cereus sensu lato organisms.
In order to gain deeper insight into the ecological distribution and genomic diversity of the pXO1-like and pXO2-like plasmids, we sequenced a panel of 94 isolates of B. cereus sensu lato organisms containing a variety of plasmids and having diverse environmental origins. This adds to the ~60 whole or draft genomes of B. cereus of various origins already available in GenBank.
De novo assemblies were generated from Illumina 101-base paired-end reads generated with two libraries, one from 180-bp fragments and one from 3-kb jumping libraries. The assemblies were constructed using AllPaths-LG (11). The protein-coding genes were predicted with Prodigal (12) and filtered to remove genes with ≥70% overlap to the tRNAs or rRNAs. The tRNAs were identified by tRNAscan-SE (13). The rRNA genes were predicted using RNAmmer (14). The gene product names were assigned based on top BLAST hits against the Swiss-Prot protein database (≥70% identity and ≥70% query coverage) and a protein family profile search against the TIGRfam HMMER equivalogs.
Nucleotide sequence accession numbers.
All 94 draft genome sequences have been deposited at GenBank under the accession no. reported in Table 1 (10, 15–22).
Table 1.
Strain characteristics
| Sample source | Strain name | GenBank accession no. | Predicted plasmid(s)d | Reference |
|---|---|---|---|---|
| Soil, Greenland | VD048 | AHEU01000000 | pXO1 | 10 |
| VD078 | AHEV01000000 | pXO1 | 10 | |
| VD045 | AHET01000000 | pXO2 | 10 | |
| VDM022 | AHFP01000000 | pXO2 | 10 | |
| VDM021 | AHFU01000000 | Neither | 10 | |
| VDM019 | AHFO01000000 | Neither | 10 | |
| Soil, Spain | VD014 | AHER01000000 | pXO1 | 10 |
| VDM006 | AHFT01000000 | pXO2 | 10 | |
| VDM034 | AHFQ01000000 | Neither | 10 | |
| Soil, Scotland | VD142 | AHCL01000000 | pXO2 | 10 |
| VD148 | AHFF01000000 | pXO2 | 10 | |
| VDM062 | AHFS01000000 | pXO2 | 10 | |
| VD136 | AHFC01000000 | Neither | 10 | |
| VD140 | AHFD01000000 | Neither | 10 | |
| VD146 | AHFE01000000 | Neither | 10 | |
| Water, Scotland | VD200 | AHFM01000000 | pXO1 | 10 |
| VD214 | AHFN01000000 | Neither | 10 | |
| Soil, Martinique | VD133 | AHFB01000000 | pXO1 | 10 |
| VD131 | AHFA01000000 | Neither | 10 | |
| Soil, Guadeloupe | VD107 | AHEX01000000 | pXO2 | 10 |
| VD115 | AHEY01000000 | pXO2 | 10 | |
| VD102 | AHEW01000000 | Neither | 10 | |
| VD118 | AHEZ01000000 | Neither | 10 | |
| Soil, Abu Dhabi, UAE | VD156 | AHFH01000000 | pXO1 | 10 |
| VD154 | AHFG01000000 | Neither | 10 | |
| Soil, Dubai, UAE | VD169 | AHFJ01000000 | pXO1 | 10 |
| VD196 | AHFL01000000 | pXO1 | 10 | |
| VD166 | AHFI01000000 | Neither | 10 | |
| VD184 | AHFK01000000 | Neither | 10 | |
| Water, Belgium (small pond site) | VD021 | AHES01000000 | pXO2 | 10 |
| VD022 | AHCK01000000 | pXO1, pXO2 | 10 | |
| VDM053 | AHFR01000000 | Neither | 10 | |
| Soil, Belgium (site A) | HuA2-1 | AHDV01000000 | pXO1 | 10 |
| HuA2-3 | AHDW01000000 | Neither | 10 | |
| HuA2-4 | AHDX01000000 | pXO2 | 10 | |
| HuA2-9 | AHDY01000000 | Neither | 10 | |
| HuA3-9 | AHDZ01000000 | pXO1 | 10 | |
| HuA4-10 | AHEA01000000 | pXO2 | 10 | |
| Soil, Belgium (site B) | HuB1-1 | AHEB01000000 | Neither | 10 |
| HuB2-9 | AHED01000000 | pXO2 | 10 | |
| HuB4-4 | AHEF01000000 | Neither | 10 | |
| HuB4-10 | AHEE01000000 | pXO2 | 10 | |
| HuB5-5 | AHEG01000000 | pXO2 | 10 | |
| HuB13-1 | AHEC01000000 | Neither | 10 | |
| Soil, Massachusetts (Boston site AG) | BAG1O-1 | AHCN01000000 | Neither | This study |
| BAG1O-2 | AHCO01000000 | Neither | This study | |
| BAG1O-3 | AHCP01000000 | Neither | This study | |
| BAG1X1-1 | AHCQ01000000 | pXO1 | This study | |
| BAG1X1-2 | AHCR01000000 | pXO1 | This study | |
| BAG1X1-3 | AHCS01000000 | pXO1 | This study | |
| BAG1X2-1 | AHCT01000000 | pXO1, pXO2 | This study | |
| BAG1X2-2 | AHCU01000000 | pXO2 | This study | |
| BAG1X2-3 | AHCV01000000 | pXO2 | This study | |
| BAG2O-1 | AHCW01000000 | Neither | This study | |
| BAG2O-2 | AHCX01000000 | Neither | This study | |
| BAG2O-3 | AHCY01000000 | Neither | This study | |
| BAG2X1-1 | AHCZ01000000 | pXO1 | This study | |
| BAG2X1-2 | AHDA01000000 | pXO1 | This study | |
| BAG2X1-3 | AHDB01000000 | pXO1 | This study | |
| BAG3O-1 | AHFV01000000 | Neither | This study | |
| BAG3O-2 | AHDC01000000 | Neither | This study | |
| BAG3X2-1 | AHDD01000000 | pXO2 | This study | |
| BAG3X2-2 | AHDE01000000 | pXO2 | This study | |
| BAG4O-1 | AHDF01000000 | Neither | This study | |
| BAG4X2-1 | AHDH01000000 | pXO2 | This study | |
| BAG4X12-1 | AHDG01000000 | pXO1, pXO2 | This study | |
| BAG5O-1 | AHDI01000000 | Neither | This study | |
| BAG5X1-1 | AHDJ01000000 | pXO1 | This study | |
| BAG5X2-1 | AHDL01000000 | pXO2 | This study | |
| BAG5X12-1 | AHDK01000000 | pXO1, pXO2 | This study | |
| Soil, Massachusetts (Boston site ES) | BAG6O-1 | AHDM01000000 | Neither | This study |
| BAG6O-2 | AHDN01000000 | Neither | This study | |
| BAG6X1-1 | AHDO01000000 | pXO1 | This study | |
| BAG6X1-2 | AHDP01000000 | pXO1 | This study | |
| Food | AND1407 | AHCM01000000 | pXO1 | 15 |
| K-5975c | AHEL01000000 | pXO1 | 19 | |
| TIAC219 | AHCJ01000000 | pXO1 | 20 | |
| Schrouff | AHCI01000000 | pXO1, pXO2 | Mahillon et al., unpublished | |
| ISP3191 | AHEK01000000 | pXO1, pXO2 | Dierick et al., unpublished | |
| ISP2954 | AHEJ01000000 | pXO1 | Dierick et al., unpublished | |
| Soil, China | B5-2 | AHFW01000000 | pXO1, pXO2 | Sun et al., unpublished |
| Mammals, Poland | IS075 | AHCH01000000 | pXO1, pXO2 | 21 |
| IS195 | AHEH01000000 | pXO1, pXO2 | 21 | |
| IS845/00 | AHEI01000000 | pXO1 | 21 | |
| Insecta | HD73 | AHDU01000000 | pXO2 | 22 |
| Various originsb | MC67 | AHEN01000000 | Other | 17 |
| MC118 | AHEM01000000 | Other | 17 | |
| MSX-A1 | AHEO01000000 | Other | 16 | |
| MSX-A12 | AHEP01000000 | Other | 16 | |
| MSX-D12 | AHEQ01000000 | Other | 16 | |
| BMG1.7 | AHDQ01000000 | Other | 18 | |
| Various originsc | CER057 | AHDS01000000 | 15 | |
| CER074 | AHDT01000000 | 15 | ||
| BtB2-4 | AHDR01000000 | 15 |
Reference strain for the pXO2-like plasmid pAW63.
Strains predicted to carry other plasmids of interest.
Strains predicted to carry plasmid fragments on the chromosome.
Neither, neither pXO1 no pXO2; other, plasmids other than pXO1 or pXO2.
ACKNOWLEDGMENTS
This project was funded in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under contract no. HHSN272200900018C. G.A.V.D.A. was supported by postdoctoral fellowships from the Belgian American Educational Foundation (BAEF) and from the Fonds National pour la Recherche Scientifique (FNRS) of Belgium.
G.A.V.D.A. and J.M. thank the many collaborators at UCL Louvain-la-Neuve, Harvard Medical School, and Huazhong Agricultural University who contributed environmental isolates to this study.
Footnotes
Citation Van der Auwera GA, Feldgarden M, Kolter R, Mahillon J. 2013. Whole-genome sequences of 94 environmental isolates of Bacillus cereus sensu lato. Genome Announc. 1(5):e00380-13. doi:10.1128/genomeA.00380-13.
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