title: Optimization of a sample processing protocol for recovery of Bacillus anthracis spores from soil.
abstract: Following a release of Bacillus anthracis spores into the environment, there is a potential for lasting environmental contamination in soils. There is a need for detection protocols for B. anthracis in environmental matrices. However, identification of B. anthracis within a soil is a difficult task. Processing soil samples helps to remove debris, chemical components, and biological impurities that can interfere with microbiological detection. This study aimed to optimize a previously used indirect processing protocol, which included a series of washing and centrifugation steps. Optimization of the protocol included: identifying an ideal extraction diluent, variation in the number of wash steps, variation in the initial centrifugation speed, sonication and shaking mechanisms. The optimized protocol was demonstrated at two laboratories in order to evaluate the recovery of spores from loamy and sandy soils. The new protocol demonstrated an improved limit of detection for loamy and sandy soils over the non-optimized protocol with an approximate matrix limit of detection at 14spores/g of soil. There were no significant differences overall between the two laboratories for either soil type, suggesting that the processing protocol will be robust enough to use at multiple laboratories while achieving comparable recoveries.
authors.affiliation: U.S. Environmental Protection Agency, National Homeland Security Research Center, 26 W. Martin Luther King Drive, MS NG16, Cincinnati, OH 45268, United States. Electronic address: Silvestri.Erin@epa.gov.
authors.forename: Erin E
authors.initials: EE
authors.lastname: Silvestri
authors.affiliation: Pegasus Technical Services, Inc., 46 East Hollister St., Cincinnati, OH 45219, United States. Electronic address: Feldhake.David@epa.gov.
authors.forename: David
authors.initials: D
authors.lastname: Feldhake
authors.affiliation: U. S. Geological Survey, Coastal and Marine Science Center, 600 4th Street South, St. Petersburg, FL 33701, United States. Electronic address: dgriffin@usgs.gov.
authors.forename: Dale
authors.initials: D
authors.lastname: Griffin
authors.affiliation: U. S. Geological Survey, Coastal and Marine Science Center, 600 4th Street South, St. Petersburg, FL 33701, United States. Electronic address: jlisle@usgs.gov.
authors.forename: John
authors.initials: J
authors.lastname: Lisle
authors.affiliation: U.S. Environmental Protection Agency, National Homeland Security Research Center, 1200 Pennsylvania Avenue, N.W., MS 8801R, Washington D.C. 20460, United States. Electronic address: Nichols.Tonya@epa.gov.
authors.forename: Tonya L
authors.initials: TL
authors.lastname: Nichols
authors.affiliation: U.S. Environmental Protection Agency, National Homeland Security Research Center, 1200 Pennsylvania Avenue, N.W., MS 8801R, Washington D.C. 20460, United States. Electronic address: Shah.Sanjiv@epa.gov.
authors.forename: Sanjiv R
authors.initials: SR
authors.lastname: Shah
authors.affiliation: Pegasus Technical Services, Inc., 46 East Hollister St., Cincinnati, OH 45219, United States. Electronic address: Pemberton.Adin@epa.gov.
authors.forename: Adin
authors.initials: A
authors.lastname: Pemberton
authors.affiliation: U.S. Environmental Protection Agency, National Homeland Security Research Center, 26 W. Martin Luther King Drive, MS NG16, Cincinnati, OH 45268, United States. Electronic address: fschaefer@zoomtown.com.
authors.forename: Frank W
authors.initials: FW
authors.lastname: Schaefer
mesh_terms: D001408:Bacillus anthracis / Q000254:growth & development / Q000302:isolation & purification; D001431:Bacteriological Techniques / Q000379:methods*; D002498:Centrifugation; D003470:Culture Media; D004783:Environmental Microbiology; D007753:Laboratories; D012987:Soil / Q000737:chemistry / Q000145:classification; D012988:Soil Microbiology; D013010:Sonication; D013171:Spores, Bacterial / Q000254:growth & development / Q000302:isolation & purification*
chemical_list: D003470:Culture Media; D012987:Soil
keywords: Bacillus anthracis; Indirect processing; Loam; Sand; Soil; Spores