title: Application of in vivo induced antigen technology (IVIAT) to Bacillus anthracis.
abstract: In vivo induced antigen technology (IVIAT) is an immuno-screening technique that identifies bacterial antigens expressed during infection and not during standard in vitro culturing conditions. We applied IVIAT to Bacillus anthracis and identified PagA, seven members of a N-acetylmuramoyl-L-alanine amidase autolysin family, three P60 family lipoproteins, two transporters, spore cortex lytic protein SleB, a penicillin binding protein, a putative prophage holin, respiratory nitrate reductase NarG, and three proteins of unknown function. Using quantitative real-time PCR comparing RNA isolated from in vitro cultured B. anthracis to RNA isolated from BALB/c mice infected with virulent Ames strain B. anthracis, we confirmed induced expression in vivo for a subset of B. anthracis genes identified by IVIAT, including L-alanine amidases BA3767, BA4073, and amiA (pXO2-42); the bacteriophage holin gene BA4074; and pagA (pXO1-110). The exogenous addition of two purified putative autolysins identified by IVIAT, N-acetylmuramoyl-L-alanine amidases BA0485 and BA2446, to vegetative B. anthracis cell suspensions induced a species-specific change in bacterial morphology and reduction in viable bacterial cells. Many of the proteins identified in our screen are predicted to affect peptidoglycan re-modeling, and our results support significant cell wall structural remodeling activity during B. anthracis infection. Identification of L-alanine amidases with B. anthracis specificity may suggest new potential therapeutic targets.
authors: Massachusetts General Hospital, Boston, Massachusetts, United States of America. srollins1@partners.org
authors: Sean M
authors: SM
authors: Rollins
authors: Amanda
authors: A
authors: Peppercorn
authors: John S
authors: JS
authors: Young
authors: Melissa
authors: M
authors: Drysdale
authors: Andrea
authors: A
authors: Baresch
authors: Margaret V
authors: MV
authors: Bikowski
authors: David A
authors: DA
authors: Ashford
authors: Conrad P
authors: CP
authors: Quinn
authors: Martin
authors: M
authors: Handfield
authors: Jeffrey D
authors: JD
authors: Hillman
authors: C Rick
authors: CR
authors: Lyons
authors: Theresa M
authors: TM
authors: Koehler
authors: Stephen B
authors: SB
authors: Calderwood
authors: Edward T
authors: ET
authors: Ryan
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000276:immunology*; D001408:Bacillus anthracis / Q000235:genetics / Q000276:immunology*; D020869:Gene Expression Profiling; D008253:Macaca mulatta; D051379:Mice; D008807:Mice, Inbred BALB C; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000942:Antigens, Bacterial; D012333:RNA, Messenger