title: Bacillus cereus strains fall into two clusters (one closely and one more distantly related) to Bacillus anthracis according to amino acid substitutions in small acid-soluble proteins as determined by tandem mass spectrometry.
abstract: Small acid-soluble proteins (SASPs) are located in the core region of Bacillus spores and have been previously demonstrated as reliable biomarkers for differentiating Bacillus anthracis and Bacillus cereus. Using MS and MS-MS analysis of SASPs further phylogenetic correlations among B. anthracis and B. cereus strains are described here. ESI was demonstrated to be a more comprehensive method, allowing for the analysis of intact proteins in both MS and MS-MS mode, thus providing molecular weight (MW) and sequence information in a single analysis, and requiring almost no sample preparation. MALDI MS was used for determination of MW of intact proteins; however, MS-MS analysis can only be achieved after enzymatic digestion of these proteins. It was demonstrated that the combination of the two different approaches provides confirmatory and complementary information, allowing for unambiguous protein characterization and sequencing. This study established that B. cereus strains fall into two clusters (one closely and one more distantly related) to B. anthracis as exhibited by amino acid substitutions. The closely related cluster was characterized by a beta-SASP with a single amino acid substitution, localized either close to the C terminus (phenylalanine-->tyrosine, 16 masses change) or close to the N terminus (serine-->alanine serine, also 16 masses change). The more distantly related cluster displayed both amino acid substitutions (32 masses change). One strain of B. cereus isolated from a patient with severe pneumonia (an anthrax-like disease) fell into the more distantly related cluster implying that pathogenicity and phylogenicity are not necessarily correlated features. Unlike PCR and DNA sequencing, protein sequence variation assessed by ESI MS-MS, essentially occurs in real-time, and involves simply extracting the protein and injecting into the instrument for analysis.
authors.affiliation: Department of Pathology, Microbiology and Immunology, University of South Carolina School of Medicine, Columbia, SC 29208, USA. castanha@gw.med.sc.edu
authors.forename: Elisangela R
authors.initials: ER
authors.lastname: Castanha
authors.forename: Marvin
authors.initials: M
authors.lastname: Vestal
authors.forename: Steve
authors.initials: S
authors.lastname: Hattan
authors.forename: Alvin
authors.initials: A
authors.lastname: Fox
authors.forename: Karen F
authors.initials: KF
authors.lastname: Fox
authors.forename: Danielle
authors.initials: D
authors.lastname: Dickinson
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000502:physiology*; D001408:Bacillus anthracis / Q000235:genetics*; D001409:Bacillus cereus / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D019143:Evolution, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D017422:Sequence Analysis, DNA / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D001426:Bacterial Proteins; C030069:spore-specific proteins, Bacillus