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Title: Detection of Agar, by Analysis of Sugar Markers, Associated with Bacillus Anthracis Spores, After Culture

Abstract

Detection of small quantities of agar associated with spores of Bacillus anthracis could provide key information regarding its source or growth characteristics. Agar, widely used in growth of bacteria on solid surfaces, consists primarily of repeating polysaccharide units of 3,6-anhydro-L-galactose (AGal) and galactose (Gal) with sulfated and O-methylated galactoses present as minor constituents. Two variants of the alditol acetate procedure were evaluated for detection of potential agar markers associated with spores. The first method employed a reductive hydrolysis step, to stabilize labile anhydrogalactose, by converting to anhydrogalactitol. The second eliminated the reductive hydrolysis step simplifying the procedure. Anhydrogalactitol, derived from agar, was detected using both derivatization methods followed by gas chromatography-mass spectrometry (GC-MS) analysis. However, challenges with artefactual background (reductive hydrolysis) or marker destruction (hydrolysis) lead to the search for alternative sugar markers. A minor agar component, 6-O-methyl galactose (6-O-M gal), was readily detected in agar-grown but not broth-grown bacteria. Detection was optimized by the use of gas chromatography-tandem mass spectrometry (GC-MS-MS). With appropriate choice of sugar marker and analytical procedure, detection of sugar markers for agar has considerable potential in microbial forensics.

Authors:
; ; ; ; ; ;
Publication Date:
Research Org.:
Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
949122
Report Number(s):
PNNL-SA-60079
Journal ID: ISSN 0167-7012; JMIMDQ; 400904030; TRN: US200907%%406
DOE Contract Number:  
AC05-76RL01830
Resource Type:
Journal Article
Journal Name:
Journal of Microbiological Methods, 74(2-3):57-63
Additional Journal Information:
Journal Volume: 74; Journal Issue: 2-3; Journal ID: ISSN 0167-7012
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ACETATES; BACILLUS; BACTERIA; DERIVATIZATION; DETECTION; GALACTOSE; HYDROLYSIS; MASS SPECTROSCOPY; POLYSACCHARIDES; SACCHAROSE; SPECTROSCOPY; SPORES

Citation Formats

Wunschel, David S, Colburn, Heather A, Fox, Alvin, Fox, Karen F, Harley, William M, Wahl, Jon H, and Wahl, Karen L. Detection of Agar, by Analysis of Sugar Markers, Associated with Bacillus Anthracis Spores, After Culture. United States: N. p., 2008. Web. doi:10.1016/j.mimet.2008.04.003.
Wunschel, David S, Colburn, Heather A, Fox, Alvin, Fox, Karen F, Harley, William M, Wahl, Jon H, & Wahl, Karen L. Detection of Agar, by Analysis of Sugar Markers, Associated with Bacillus Anthracis Spores, After Culture. United States. https://doi.org/10.1016/j.mimet.2008.04.003
Wunschel, David S, Colburn, Heather A, Fox, Alvin, Fox, Karen F, Harley, William M, Wahl, Jon H, and Wahl, Karen L. 2008. "Detection of Agar, by Analysis of Sugar Markers, Associated with Bacillus Anthracis Spores, After Culture". United States. https://doi.org/10.1016/j.mimet.2008.04.003.
@article{osti_949122,
title = {Detection of Agar, by Analysis of Sugar Markers, Associated with Bacillus Anthracis Spores, After Culture},
author = {Wunschel, David S and Colburn, Heather A and Fox, Alvin and Fox, Karen F and Harley, William M and Wahl, Jon H and Wahl, Karen L},
abstractNote = {Detection of small quantities of agar associated with spores of Bacillus anthracis could provide key information regarding its source or growth characteristics. Agar, widely used in growth of bacteria on solid surfaces, consists primarily of repeating polysaccharide units of 3,6-anhydro-L-galactose (AGal) and galactose (Gal) with sulfated and O-methylated galactoses present as minor constituents. Two variants of the alditol acetate procedure were evaluated for detection of potential agar markers associated with spores. The first method employed a reductive hydrolysis step, to stabilize labile anhydrogalactose, by converting to anhydrogalactitol. The second eliminated the reductive hydrolysis step simplifying the procedure. Anhydrogalactitol, derived from agar, was detected using both derivatization methods followed by gas chromatography-mass spectrometry (GC-MS) analysis. However, challenges with artefactual background (reductive hydrolysis) or marker destruction (hydrolysis) lead to the search for alternative sugar markers. A minor agar component, 6-O-methyl galactose (6-O-M gal), was readily detected in agar-grown but not broth-grown bacteria. Detection was optimized by the use of gas chromatography-tandem mass spectrometry (GC-MS-MS). With appropriate choice of sugar marker and analytical procedure, detection of sugar markers for agar has considerable potential in microbial forensics.},
doi = {10.1016/j.mimet.2008.04.003},
url = {https://www.osti.gov/biblio/949122}, journal = {Journal of Microbiological Methods, 74(2-3):57-63},
issn = {0167-7012},
number = 2-3,
volume = 74,
place = {United States},
year = {Fri Aug 01 00:00:00 EDT 2008},
month = {Fri Aug 01 00:00:00 EDT 2008}
}