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. 2013 Oct 3;1(5):e00380-13. doi: 10.1128/genomeA.00380-13

Whole-Genome Sequences of 94 Environmental Isolates of Bacillus cereus Sensu Lato

Géraldine A Van der Auwera a,b,a,b,, Michael Feldgarden b, Roberto Kolter c, Jacques Mahillon a
PMCID: PMC3790080  PMID: 24092776

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 (16).

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, 1522).

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
a

Reference strain for the pXO2-like plasmid pAW63.

b

Strains predicted to carry other plasmids of interest.

c

Strains predicted to carry plasmid fragments on the chromosome.

d

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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