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ARS Home » Research » Publications at this Location » Publication #233384

Title: Detection and bioavailability of two biothreat toxins

Author
item Brandon, David
item Cheng, Luisa Wai Wai
item He, Xiaohua
item Rasooly, Reuven
item Scotcher, Miles
item Stanker, Larry
item Carter, John

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 11/13/2008
Publication Date: 11/13/2008
Citation: Brandon, D.L., Cheng, L.W., He, X., Rasooly, R., Scotcher, M.C., Stanker, L.H., Carter, J.M. 2008. Detection and bioavailability of two biothreat toxins. Meeting Abstract. Poster No. 2.

Interpretive Summary:

Technical Abstract: Potential bioterror incidents involving intentional adulteration of food seem likely to involve cheap, easily prepared crude toxins. We hypothesized that the impurities in such preparations may influence the stability and oral bioavailability of the toxins in food matrices. We have recently focused on ricin and botulinum neurotoxin (BoNT), comparing the bioavailability of crude versus pure forms and developing sensitive test methods that accurately measure toxins and marker compounds. Using the mouse bioassay, the “Gold Standard,” to test for BoNT bioavailability in food, we found that bioavailability was affected by both toxin purity and the matrix. The results of bioassay also showed that crude BoNT is more stable to heating than pure toxin, an important consideration in threat and risk assessment. Although well validated and reliable, animal models have several drawbacks, and we are therefore developing and evaluating alternative tests, especially immunochemical and activity-based formats. We have developed new monoclonal antibody reagents (mAbs) that bind BoNT and ricin with high affinity, affording assay sensitivities exceeding those of rodent bioassays. These antibody reagents have also proven useful for sample preparation and production of portable tests for field use. MAbs have been used for sample preparation in the form of immunomagnetic beads. After scavenging a large volume of liquid food, the beads are sequestered with a magnet, and the highly specific peptidase activity of BoNT is detected by fluorescence resonance energy transfer (FRET), using a suitably labeled synthetic peptide substrate. While BoNT exhibits a characteristic peptidase activity, ricin exerts toxicity through its glycosidase activity, removing adenine from 28S rRNA with exquisite specificity, resulting in ribosomal inactivation. The inhibition of translation of luciferase mRNA by ricin provides a cell-free luminescence assay to measure active toxin (Hale, 2001). The stability of ricin in various food matrices was studied using this approach. Crude ricin, such as the lethal material found recently in a Las Vegas hotel room, contains traces of multiple components of the castor bean, including DNA. This genetic material can be detected with excellent sensitivity and specificity by a quantitative PCR. Although we found that some food matrices interfere with the PCR and the cell-free translation assay, both methods are sufficiently sensitive so that dilution of liquid foods or food extracts provides adequate sample preparation for reliable detection of sublethal amounts of toxin.