title: Detection and tracking of a novel genetically tagged biological simulant in the environment.
abstract: A variant of Bacillus thuringiensis subsp. kurstaki containing a single, stable copy of a uniquely amplifiable DNA oligomer integrated into the genome for tracking the fate of biological agents in the environment was developed. The use of genetically tagged spores overcomes the ambiguity of discerning the test material from pre-existing environmental microflora or from previously released background material. In this study, we demonstrate the utility of the genetically "barcoded" simulant in a controlled indoor setting and in an outdoor release. In an ambient breeze tunnel test, spores deposited on tiles were reaerosolized and detected by real-time PCR at distances of 30 m from the point of deposition. Real-time PCR signals were inversely correlated with distance from the seeded tiles. An outdoor release of powdered spore simulant at Aberdeen Proving Ground, Edgewood, MD, was monitored from a distance by a light detection and ranging (LIDAR) laser. Over a 2-week period, an array of air sampling units collected samples were analyzed for the presence of viable spores and using barcode-specific real-time PCR assays. Barcoded B. thuringiensis subsp. kurstaki spores were unambiguously identified on the day of the release, and viable material was recovered in a pattern consistent with the cloud track predicted by prevailing winds and by data tracks provided by the LIDAR system. Finally, the real-time PCR assays successfully differentiated barcoded B. thuringiensis subsp. kurstaki spores from wild-type spores under field conditions.
authors.affiliation: Edgewood Chemical Biological Center, Aberdeen Proving Ground, Maryland, USA.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Emanuel
authors.forename: Patricia E
authors.initials: PE
authors.lastname: Buckley
authors.forename: Tiffany A
authors.initials: TA
authors.lastname: Sutton
authors.forename: Jason M
authors.initials: JM
authors.lastname: Edmonds
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Bailey
authors.forename: Bryan A
authors.initials: BA
authors.lastname: Rivers
authors.forename: Michael H
authors.initials: MH
authors.lastname: Kim
authors.forename: William J
authors.initials: WJ
authors.lastname: Ginley
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Keiser
authors.forename: Robert W
authors.initials: RW
authors.lastname: Doherty
authors.forename: F Joseph
authors.initials: FJ
authors.lastname: Kragl
authors.forename: Fiona E
authors.initials: FE
authors.lastname: Narayanan
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Katoski
authors.forename: Sari
authors.initials: S
authors.lastname: Paikoff
authors.forename: Samuel P
authors.initials: SP
authors.lastname: Leppert
authors.forename: John B
authors.initials: JB
authors.lastname: Strawbridge
authors.forename: Daniel R
authors.initials: DR
authors.lastname: VanReenen
authors.forename: Sally S
authors.initials: SS
authors.lastname: Biberos
authors.forename: Douglas
authors.initials: D
authors.lastname: Moore
authors.forename: Douglas W
authors.initials: DW
authors.lastname: Phillips
authors.forename: Lisa R
authors.initials: LR
authors.lastname: Mingioni
authors.forename: Ogba
authors.initials: O
authors.lastname: Melles
authors.forename: Daniel G
authors.initials: DG
authors.lastname: Ondercin
authors.forename: Beth
authors.initials: B
authors.lastname: Hirsh
authors.forename: Kendall M
authors.initials: KM
authors.lastname: Bieschke
authors.forename: Crystal L
authors.initials: CL
authors.lastname: Harris
authors.forename: Kristin M
authors.initials: KM
authors.lastname: Omberg
authors.forename: Vipin K
authors.initials: VK
authors.lastname: Rastogi
authors.forename: Sheila
authors.initials: S
authors.lastname: Van Cuyk
authors.forename: Henry S
authors.initials: HS
authors.lastname: Gibbons
mesh_terms: D000391:Air Microbiology; D001408:Bacillus anthracis / Q000302:isolation & purification; D001413:Bacillus thuringiensis / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D001431:Bacteriological Techniques / Q000379:methods*; D058893:DNA Barcoding, Taxonomic / Q000379:methods*; D008954:Models, Biological; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D013171:Spores, Bacterial / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D013194:Staining and Labeling / Q000379:methods; D013997:Time Factors