title: Genetic basis for sulfonamide resistance in Bacillus anthracis.
abstract: Natural resistance of field strains of Bacillus anthracis to drugs from the sulfonamide class of antimicrobials that act by inhibiting dihydropteroate synthase (DHPS) has been reported. Though the structure of B. anthracis DHPS has been determined, its connection to the apparent intrinsic sulfonamide resistance of the bacterium has not been established. The aim of this study was to determine if a connection exists between DHPS and the observed sulfonamide resistance of B. anthracis. Microdilution broth assays verified that B. anthracis Sterne is highly resistant to a variety of sulfonamides with minimum inhibitory concentrations (MICs) exceeding 1250 microg/ml. A putative gene encoding DHPS (folP) was amplified from B. anthracis Sterne chromosomal DNA by polymerase chain reaction (PCR) and cloned. Sequence comparisons showed 100% identity with DHPSs from published genome sequences for various strains of B. anthracis. Additionally, expression of folP in B. anthracis Sterne was confirmed. Functionality of the B. anthracis DHPS was confirmed by complementation of an Escherichia coli folP deletion mutant as well as a standard enzyme assay. Concomitant transfer of high level sulfonamide resistance to this mutant along with increased sulfonamide IC(50)values for purified B. anthracis DHPS links DHPS to sulfonamide resistance in B. anthracis. These findings lay the groundwork that will aid future development of antimicrobics that target DHPS to treat anthrax infections.
authors.affiliation: Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA.
authors.forename: Michelle Wright
authors.initials: MW
authors.lastname: Valderas
authors.forename: Philip C
authors.initials: PC
authors.lastname: Bourne
authors.forename: William W
authors.initials: WW
authors.lastname: Barrow
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001408:Bacillus anthracis / Q000187:drug effects* / Q000201:enzymology / Q000254:growth & development; D001431:Bacteriological Techniques; D004094:Dihydropteroate Synthase / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics*; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013449:Sulfonamides / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D013449:Sulfonamides; D004094:Dihydropteroate Synthase