strategies for the detection of unknown biological agents dr. peter j. stopa us army edgewood...
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STRATEGIES FOR THE DETECTION OF UNKNOWN
BIOLOGICAL AGENTS
Dr. Peter J. Stopa
US Army Edgewood Chemical Biological Center
Aberdeen Proving Ground, MD
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OBJECTIVES FOR UNKNOWNS DETECTION
Differentiate between natural and artificial events.Implement and maintain protective postures.Confirm whether forces have been attacked.Initiate treatment in the absence of a specific biological agent hazard.
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Potential Classification Parameters
ClassifyPathogen/non-Pathogen
Aerosol Additives
Naturalvs.
Man-made Bacteria
Encapsulants
Virus
Protein Peptide Toxin
Non-Protein Toxin
Bio/non-Bio
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Natural vs. Man-made
Parameter Test Rationale Status
Particles Particle-size Analysis
Most man-made aerosols are > 3 um
Commercial
Particle-shape Analysis
May see uniform distribution of similar particle shapes.
Commercial
Elemental Analysis
Spectrometric analysis
May see “unnatural” elements or ratios of elements appear.
Commercial
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Aerosol Additives
Parameter Test Rationale Status Silica Particles Silica Analysis Silica particles can
be used to improve aerosol dissemination.
Commercial – Various commercial manufacturers have silica tests available.
Protein Content Standard Protein Determinations
Protein content in air is low. Could be used as an additive or agent itself.
Available (Field) – various protein detection kits are routinely used.
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Bio/non-Bio Determination
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Pathogen/non-pathogen Determination
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Bacterial Specific Detection
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Viral Specific Detection
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Protein /Peptide Specific Detection
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Non-Protein ToxinSpecific Detection
Parameter Test Rationale Status Virulence Determination
Binding to receptors. Sentinel Cell Assay
A variety of toxins bind to gangliosides. Measure binding to a sentinel cell.
Available (Lab) –.
Molecular Weight Determination
Mass Spectrometry Scope of potential physiological active materials is great. Only MS can probably address.
Demonstrated in the laboratory.
Enzymatic Activity
Substrate Analyses A variety of toxins inhibit specific enzymatic processes.
Demonstrated in the laboratory.
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Determination of Appropriate Equipment
How do you determine what you need?
Need to identify: The end user. How they operate. Constraints. Performance Criteria. How important are each of these factors in
determining the best equipment?) (Weighting Factors)
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A Computer-based Approach
Can identify each of the parameters and subsequent weighting factors and plug them into a statistical model.Identify appropriate candidate technologies and score them according to your criteria.Obtain the final results.See how they fared and why.Make purchasing decisions.
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Biological DetectionMore difficult than chemical. No vapor pressure – sampling for detection is more
difficult. Present in low concentrations. May be hard to discriminate an intentional release
against background levels of similar “natural” biological materials.
Technologies not as mature. Clinical personnel sometimes do not understand “the
field”. Most confirmatory technologies are slow (i.e., culture). Microbiologists are reluctant to adopt instrumented
methods (need to see it or smell it to believe it).
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Biological Detection Process
Collect samples
Chemical Property Characterization
Generic detection Biological agent
characteristics
Specific identification Identification kits Reference laboratory
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Chemical Properties
From July/August 2003 Homeland First Response
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Chemical Properties
From July/August 2003 Homeland First Response
Premise:
• Biological materials can be viable only within a pH range of 5-9.
• Turbidity and density in water may give an indication of particle size and solubility.
• Presence of Protein may indicate a biological agent.
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Chemical Tests
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WHY GENERIC DETECTION?
Rapidly provide the Incident Commander enough information to justify the response to a suspected biological terrorist threat.Minimize response resources to false threats.Evaluate if the sample can be further analyzed with immunoassay tickets on site or sent to a laboratory for confirmation.Should not rely on immunoassay tickets alone. Limited library - some materials that a terrorist may
use are not on the threat list. For ticket results to be valid, the sample must be
within certain concentration, pH, and ionic strength guidelines.
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Biological Properties
Generic detectors that detect biological properties are relatively rapid, inexpensive, and easy to use.
More information than the chemical property test alone –i.e., is it LIVE?
Confirmatory identification must be performed in the laboratory.
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Generic Detection Strategy
Use various properties to detect the presence of a biological material in a suspect sample. Luminescence – ATP –
viability (cell battery). DNA – present in all
biological materials. (blueprint).
Protein – also present in all biological materials. (building block).
Particle size/concentration.
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Biological Detection Instruments
Particle analyzer
Fluorometer (DNA)
Luminometer (ATP)
Colorimeter (protein)Particle Analyzer
Colorimeter
Fluorometer
Luminometers
Integrated System
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Expected Results for Generic Detection
SAMPLE ATP DNA PROTEIN
Bacteria + + ? (1)
Toxin - ? (2) +
Virus - + ? (1)
(1) Possibly from culture media.
(2) If protein concentration is high enough (> 10 ug/ml), will probably see contaminating DNA from protein source.
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Identification
Immunoassay TicketsAnd Readers Biological Laboratory
Field Instruments(PCR)
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Comments
Experience from Fall 2001 showed that no single technology was 100% accurate.
Both IA platforms and PCR platforms missed samples for various reasons.
Culture was able to detect in all of the samples – but TOO slow to be of value in the field.
Need redundant capabilities.
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CONCLUSIONS
Tests and equipment exist for use by first responders for field screening of biological samples.
Additional discussion is needed among the policy makers, field responders, and reference laboratories to develop or accept a workable system.