Corinne Byrdsong Spring 2014 Seminar Relevance Regulation Effects - - PowerPoint PPT Presentation

Corinne Byrdsong Spring 2014 Seminar

Relevance Regulation Effects of exposure Method of action Detection methods PCR analysis

1978 - Bulgarian dissident Georgi Markov assassinated in London April 2013 – Letters addressed to the US President March 2014 – Pennsylvania teen March 2014 – ricin in Georgetown dorm Both a criminal and a public health issue

Ricin is considered a biological and a chemical weapon Classified as Category B biological threat by the CDC

Prohibited by international treaties Biological Weapons Anti-Terrorism Act of 1989 Antiterrorism and Effective Death Penalty Act

• f 1996

USA PATRIOT Act of 2001 Public Health Security and Bioterrorism Preparedness Act of 2002

• Ricinus communis

– Castor beans

• A major protein in

castor beans

• Comes from the waste
• f castor oil production

Image from: http://www.printmag.com/design-inspiration/botany- blueprint-the-castor-bean/ Image from: http://dsonggonplant.blogspot.com/2012/02/ricinus- communis-usually-name-called.html

• Many uses for the oil:

– Cosmetics – Paints, varnishes – Organic fertilizer – Nylon – Synthetic flavors – Potential biofuel

Image from: http://beautyeditor.ca/2010/11/15/my-brief-traumatic-foray-into-the-oil-cleansing-method-or-how-overreaching-can-sometimes- backfire-on-your-skin/

• R. communis grows in most tropical and

temperate regions throughout the world The Anarchist’s Cookbook The Poisoner’s Handbook The Internet Easily and inexpensively obtained

Image from: http://en.wikipedia.org/wiki/The_Anarchist_Cookbook

A plant toxin A lectin Ribosome inactivating protein (RIP)

Image from: http://www.wiley.com/college/boyer/0470003790/cutting_edge/molecular_recognition/ricin.gif

Inhibits protein synthesis

This leads to the death of eukaryotic cells

Image from: http://ibbio.pbworks.com/w/page/28624940/Nucleic%20Acids%20and%20Proteins

Lethal dose depends on route of exposure

Generally, 1 – 20 mg of ricin/kg body weight

Death takes place 36 – 72 hours after exposure, depending on the amount and route

• f exposure

Accidental exposure unlikely

Depends on route of exposure Non-specific but severe

Will ultimately see respiratory, organ failure

Ricin poisoning not contagious

No antidote Treat symptoms Try to get ricin out of body

Recent ingestion – flush stomach with activated charcoal

Know if a person has been exposed Decontamination measures Show ricin to be biologically active Presumptive screening

Rapid results

Confirmatory analysis

Environmental

Sample

Immunological

assays

Mass spectrometry Detection of ricin protein Biological Assays Detection of residual DNA Real-time PCR

Environmental

Sample

Immunological

assays

Mass spectrometry Detection of ricin protein Biological Assays Detection of residual DNA Real-time PCR

Rapid screening Specific High sensitivity Quantitative

Image from: http://classconnection.s3.amazonaws.com/826/flashcards/512826/png/picture131323331655437.png

Amplification of specific sequence Ricin-specific primers Ricin-specific probes ‘Real-time’ data collection

Point when amplification first detected during cycle

Important for:

Risk assessment Medical treatment

Results in 4-5 hours

Detection of R. communis DNA Detect presence of DNA encoding the gene for ricin A-chain Specific nucleic acid sequences

• R. communis DNA as residual contaminant from ricin

preparation Small amounts detected

High detection from various samples

Castor bean suspension Spiked powders

Amount of R. communis DNA

Get idea of how much toxin present

Determined using exponential phase

Doubling of products

Image from: http://www.biotechniques.com/BiotechniquesJournal/specialissues/2008/April/Twenty-five-years-of-quantitative-PCR-for-gene-expression- analysis/biotechniques-45217.html?pageNum=2

Direct investigators to next step

Confirmatory assays

Agricultural plant-breeding programs

Low levels of ricin

Ricin

Biological and chemical agent Easily processed

Detection Methods

Presumptive

PCR Analysis

Confirmatory

Future studies

• Dr. Staton
• Dr. Fenger

Darcie Winkler Kel Daniel AJ Montañez KariAnna Baber

Bundowle, B., Schutzer, S. E., Breeze, R. G., Keim, P. S., & Morse, S. A. (Eds.). (2010). Microbial forensics. (2nd ed.). Burlington, MA: Elsevier/Academic Press. Hutchins, A. S., Astwood, M. J., Saah, J. R., Michel, P. A., Newton, B. R., & Dauphin, L.

• A. (2014). Evaluation of automated and manual dna purification methods for

detecting ricinus communis dna during ricin investigations. Forensic Science International, 236, 10-15. doi: 10.1016/j.forsciint.2013.12.011. Melchior, W.B., Jr., & Tolleson, W.H. (2010). A functional quantitative polymerase chain reaction assay for ricin, Shiga toxin, and related ribosome-inactivating

• proteins. Analytical Biochemistry, 396, 204-211. doi: 10.1016/j.ab.2009.09.024

Puri, P., & Kumar, O. (2011). Integrating immunobased detection and identification methods for ricin analysis: An overview. Journal of Bioterrorism & Biodefense, S2:003. doi: Integrating Immunobased Detection and Identification Methods for Ricin Analysis: An Overview Schieltz, D.M., McGrath, S.C., McWilliams, L.G., Rees, J., Bowen, M.D., Kools, J.J., Dauphin, L.A., Gomez-Saladin, E., Newton, B.N., Stang, H.L., Vick, M.J., Thomas, J. Pirkle, J.L., & Barr, J.R. (2011). Analysis of active ricin and castor bean proteins in a ricin preparation castor bean extract, and surface swabs from a public health

• investigation. Forensic Science International, 209, 70-79. doi:

10.1016/j.forsciint.2010.12.013 Shea, D. A., & Gottron, F. (2010, December). Ricin: technical background and potential role in terrorism. Library of Congress Washington D.C. Congressional Research Service.

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