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.

O-Acetylation of Peptidoglycan Is Required for Proper Cell Separation and S-layer Anchoring in Bacillus anthracis

“…In the host, B. anthracis remodels its peptidoglycan as suggested by the abundant production of peptidoglycan hydrolases during infection (47). Because purified B. anthracis peptidoglycan stimulates an inflammatory response in vitro (48), controlling the composition of muropeptides shed during multiplication in the host may enable better survival of the pathogen.…”

Characterization of Anti- Salmonella enterica Serotype Typhi Antibody Responses in Bacteremic Bangladeshi Patients by an Immunoaffinity Proteomics-Based Technology

“…A number of high-throughput immunoaffinity screens have been developed to evaluate host-pathogen interactions during human infection, including IVIAT and PELS (11,12,17,38,39). Here we describe a further advance of such approaches, IPT, and used this to identify 57 S. Typhi proteins recognized by antibodies of various isotypes from individuals bacteremic with S. Typhi in Bangladesh.…”

Identification of Toxoplasma gondii in-vivo induced antigens by cDNA library immunoscreening with chronic toxoplasmosis sera

“…Furthermore an important reason for the introduction of IVIAT was to reduce the need to use animals. In addition other researchers have used in-vitro grown organisms in studies which employ IVIAT [24,25].…”