An optimized MLST scheme for the Bacillus cereus group

Our research group, led by Prof. Anne-Brit Kolstø at the Laboratory for Microbial Dynamics (LaMDa), Department of Pharmaceutical Biosciences, School of Pharmacy, University of Oslo, has designed and applied the first MLST scheme for phylogenetic analysis of B. cereus group bacteria (Helgason et al. 2004, Appl. Environ. Microbiol. 70:191-201), leading to the identification of several clonal lineages comprising strains isolated from clinical sources. Alternative schemes have been subsequently developed by several other groups (Ko et al. 2004, Infect. Immun. 72:5253-61; Priest et al. 2004, J. Bacteriol. 186:7959-70; Candelon et al. 2004, Microbiology 150:601-611; Sorokin et al. 2006, Appl. Environ. Microbiol. 72:1569-1578).

In order to provide the community with the most robust and practical scheme, we have developed, in collaboration with Dr. Alexei Sorokin, INRA, France, a new combined and optimized MLST scheme for the B. cereus group adapted from three of the four available schemes. This new scheme is described in Tourasse, Helgason et al. 2006, J. Appl. Microbiol. 101:579-593. The seven genes chosen are listed below. The total length of the fragments used for MLST is 2,658 bp.

gene locus in B.cereus ATCC 14579 encoded protein genomic position in B.cereus ATCC 14579 total gene length fragment length used for MLST fragment position in the gene
adkBC0152adenylate kinase137017−137667651411163−573
glpTBC0656glycerol−3−phosphate permease / transporter653160−6518111,350330958−1287
glpFBC1034glycerol uptake facilitator protein1014999−1015835822372151−522
panCBC1541pantoate−β−alanine ligase1489715−1490563849350307−656
pycABC3947pyruvate carboxylase3930297−39268513,4473632443−2805
ccpABC4672catabolite control protein A4612709−4611711999418567−150
ptaBC5387phosphate acetyltransferase5304503−5303532972414181−594

The adk gene fragment has been shortened from 450 bp (as originally published in Helgason et al. 2004 ) to 411 bp, by trimming the first 16 bp and the last 23 bp. The glpF fragment has also been shortened from 381 (as originally published in Priest et al. 2004) to 372 bp by trimming the first 9 bp. These changes provide better sequencing reads, as the trimmed regions are close to the primers used for PCR amplification and sequencing.

The positional distribution of the seven genes on the 5,411,809 bp chromosome of the type strain of B. cereus, ATCC 14579, is shown below (oriC and dif indicate the origin of replication and the replication termination region, respectively). The relative positions are similar in the chromosomes of other completely sequenced B. cereus, B. thuringiensis, and B. anthracis strains.

Gene_locations_picture

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