title: Effects of endogenous D-alanine synthesis and autoinhibition of Bacillus anthracis germination on in vitro and in vivo infections.
abstract: Bacillus anthracis transitions from a dormant spore to a vegetative bacillus through a series of structural and biochemical changes collectively referred to as germination. The timing of germination is important during early steps in infection and may determine if B. anthracis survives or succumbs to responsive macrophages. In the current study experiments determined the contribution of endogenous D-alanine production to the efficiency and timing of B. anthracis spore germination under in vitro and in vivo conditions. Racemase-mediated production of endogenous D-alanine by B. anthracis altered the kinetics for initiation of germination over a range of spore densities and exhibited a threshold effect wherein small changes in spore number resulted in major changes in germination efficiency. This threshold effect correlated with D-alanine production, was prevented by an alanine racemase inhibitor, and required L-alanine. Interestingly, endogenous production of inhibitory levels of D-alanine was detected under experimental conditions that did not support germination and in a germination-deficient mutant of B. anthracis. Racemase-dependent production of D-alanine enhanced survival of B. anthracis during interaction with murine macrophages, suggesting a role for inhibition of germination during interaction with these cells. Finally, in vivo experiments revealed an approximately twofold decrease in the 50% lethal dose of B. anthracis spores administered in the presence of D-alanine, indicating that rates of germination may be directly influenced by the levels of this amino acid during early stages of disease.
authors.affiliation: Department of Microbiology and Immunology, The University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
authors.forename: Matthew T
authors.initials: MT
authors.lastname: McKevitt
authors.forename: Katie M
authors.initials: KM
authors.lastname: Bryant
authors.forename: Salika M
authors.initials: SM
authors.lastname: Shakir
authors.forename: Jason L
authors.initials: JL
authors.lastname: Larabee
authors.forename: Steven R
authors.initials: SR
authors.lastname: Blanke
authors.forename: Julie
authors.initials: J
authors.lastname: Lovchik
authors.forename: C Rick
authors.initials: CR
authors.lastname: Lyons
authors.forename: Jimmy D
authors.initials: JD
authors.lastname: Ballard
mesh_terms: D000409:Alanine / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D000411:Alanine Racemase / Q000737:chemistry; D000818:Animals; D000881:Anthrax / Q000378:metabolism* / Q000382:microbiology*; D001408:Bacillus anthracis / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D005260:Female; D008264:Macrophages / Q000276:immunology / Q000382:microbiology; D051379:Mice; D008807:Mice, Inbred BALB C; D008811:Mice, Inbred DBA; D013171:Spores, Bacterial / Q000737:chemistry / Q000254:growth & development / Q000378:metabolism; D013237:Stereoisomerism; D014774:Virulence
chemical_list: D000411:Alanine Racemase; D000409:Alanine