Each ces gene displays 90 ~ 95% identity

between B. cereus and B. weihenstephanensis, and 95 ~ 100% identity within B. weihenstephanensis learn more isolates. Similar but slightly lower identity levels were observed for the corresponding proteins. Thus, based on the concatenated ces genes and protein sequences, two main clusters, namely “”cereus”" and “”weihenstephanensis”", could be distinguished, and within “”weihenstephanensis”" cluster, two subsequent clades were identified (Figure  1B). Genomic location of the ces gene clusters IS075 harbors a larger plasmid pool than AH187. The cereulide gene cluster of IS075 was observed to be located on a large plasmid with a size similar to that of pCER270 (270 kb) in AH187 (Figure  2A). Like pCER270, IS075 was PCR-positive to the pXO1 backbone genes pXO1-11, pXO1-14, pXO1-45, pXO1-50 and pXO1-55, which all encode hypothetical proteins (data not shown). It was also observed that the IS075 contig containing the ces gene cluster is ca. 180.7 kb with 146 predicted CDSs, of which 85.6% matched to those of pCER270, with a good synteny (Figure  2B). This indicated that the emetic plasmid in IS075 is pXO1-like with high similarity to pCER270. The deduced proteins from 21 predicted CDSs not matching those of pCER270 were blasted with

databases (Nr and Swissprot). The result showed that two matched putative transposases, one was related to putative DNA topoisomerases I, one to putative transcriptional repressors, see more and the others to

hypothetical proteins, all with homologs in other B. cereus group plasmids. Figure 2 Genomic location of the cereulide gene cluster. (A) Genomic location of the cereulide gene cluster of emetic B. cereus group isolates determined by plasmid profiling (L) and hybridization (R). Lane 1: IS075, lane 2: MC118, lane 3: MC67, lane 4: CER057, lane 5: BtB2-4, lane 6: non cereulide-producing B. cereus isolate CER071, lane 7: AH187. The probe used was cesB internal fragment amplified with EmF and EmR primers from the reference strain AH187. pMC118 and pMC67, displaying a larger size than pCER270, are indicated by a dark triangle. (B) Linear arrangement of the contig containing the ces gene cluster of (L) CER057 with the chromosome of KBAB4 and (R) Sclareol IS075 with the plasmid pCER270. Aligned segments are represented as dots (20 ~ 65 bp) and lines (>65 bp), with red and blue colors refer to forward and reverse matching substrings, respectively. For BtB2-4 and CER057, although large plasmid with smaller size to pCER270 was observed in the profile, no hybridization signal was detected (Figure  2A). It was observed that the contig containing the ces gene cluster in CER057 is about 245.4 kb with 215 predicted CDSs, of which 80% and 85% matched those of the chromosomes of AH187 and KBAB4, respectively.