No abstract

No abstract

There is little published data on the performance of biological indicator tests and immunoassays that could be used by first responders to determine if a suspicious powder contains a potential biothreat agent. We evaluated a range of biological indicator tests, including 3 protein tests, 2 ATP tests, 1 DNA test, and 1 FTIR spectroscopy instrument for their ability to screen suspicious powders for Bacillus anthracis (B. anthracis) spores and ricin. We also evaluated 12 immunoassays (mostly lateral flow immunoassays) for their ability to screen for B. anthracis and ricin. We used a cost-effective, statistically based test plan that allows instruments to be evaluated at performance levels ranging from 0.85 to 0.95 lower confidence bound of the probability of detection at confidence levels of 80% to 95%. We also assessed interference with 22 common suspicious powders encountered in the field. The detection reproducibility for the biological indicators was evaluated at 108 B. anthracis spores and 62.5 μg ricin, and the immunoassay detection reproducibility was evaluated at 107 spores/mL (B. anthracis) and 0.1 μg/mL (ricin). Seven out of 12 immunoassays met our most stringent criteria for B. anthracis detection, while 9 out of 12 met our most stringent test criteria for ricin detection. Most of the immunoassays also detected ricin in 3 different crude castor seed preparations. Our testing results varied across products and sample preparations, indicating the importance of reviewing performance data for specific instruments and sample types of interest for the application in order to make informed decisions regarding the selection of biodetection equipment for field use.

There is little published data on the performance of hand-portable polymerase chain reaction (PCR) systems that can be used by first responders to determine if a suspicious powder contains a potential biothreat agent. We evaluated 5 commercially available hand-portable PCR instruments for detection of Bacillus anthracis. We used a cost-effective, statistically based test plan to evaluate systems at performance levels ranging from 0.85-0.95 lower confidence bound (LCB) of the probability of detection (POD) at confidence levels of 80% to 95%. We assessed specificity using purified genomic DNA from 13 B. anthracis strains and 18 Bacillus near neighbors, potential interference with 22 suspicious powders that are commonly encountered in the field by first responders during suspected biothreat incidents, and the potential for PCR inhibition when B. anthracis spores were spiked into these powders. Our results indicate that 3 of the 5 systems achieved 0.95 LCB of the probability of detection with 95% confidence levels at test concentrations of 2,000 genome equivalents/mL (GE/mL), which is comparable to 2,000 spores/mL. This is more than sufficient sensitivity for screening visible suspicious powders. These systems exhibited no false-positive results or PCR inhibition with common suspicious powders and reliably detected B. anthracis spores spiked into these powders, though some issues with assay controls were observed. Our testing approach enables efficient performance testing using a statistically rigorous and cost-effective test plan to generate performance data that allow users to make informed decisions regarding the purchase and use of field biodetection equipment.

Robust and highly specific methods for the detection of the protein toxin ricin are of interest to the law enforcement community. In previous studies, methods based on liquid chromatography−tandem mass spectrometry shotgun proteomics have been proposed. The successful implementation of this approach relies on specific data evaluation criteria addressing (1) the quality of the mass spectrometric data, (2) the confidence of peptide identifications (peptide-spectrum matches), and (3) the number and sequence specificity of peptides detected. We present such data evaluation criteria and use a novel approach to establish the limit of detection for this ricin assay. Specifically, we use logistic regression to determine the probability of detection for individual ricin peptides at different concentrations. We then apply basic rules from probability theory, combining these individual peptide probabilities into an overall assay limit of detection. This procedure yields an assay limit of detection for ricin at 42.5 ng on column or 21.25 ng/μL for a 2-μL injection. We also show that, despite the conventional wisdom that detergents are deleterious to mass spectrometric analyses, the presence of Tween-20 did not prevent detection of ricin peptides, and indeed assays performed in buffers that included Tween-20 gave better results than assays performed using other buffer formulations with or without detergent removal.

No abstract

Background The BioFire Bone and Joint Infection (BJI) Panel is a sample-to-answer test for the qualitative detection of nearly 40 different bacteria, yeast, and antimicrobial resistance (AMR) genes in synovial fluid (SF). The panel aims to improve on current culture-based diagnostics, particularly for detection of anaerobes (e.g. Finegoldia magna, Kingella kingae, Cutibacterium, Anaerococcus and Peptoniphilus species, and others) in about an hour. Analytical performance of the panel (Limit of Detection (LoD), analytical reactivity and specificity, interference, reproducibility), and specimen storage conditions are described. Methods LoD for each analyte was estimated from serial dilutions and confirmed at the lowest titer with ≥95% detection. A collection of >350 isolates representing genetic and geographic diversity of analytes was tested near LoD to assess analytical reactivity, and more than 420 near-neighbor, commensal, pathogenic, or environmental off-panel species were evaluated for assay specificity. Reproducibility was evaluated in a multi-laboratory multi-variable study, and the impact of storage and potentially interfering substances on the accuracy of test results was also assessed. Testing was performed with Investigational Use Only kits. Results The confirmed LoD for bacteria and yeast ranged from 100 - 10,000 CFU/mL. Sequence analysis and testing demonstrated clinically appropriate specificity and reactivity with a variety of isolates and different AMR gene types. Accurate and reproducible organism and AMR gene detection was observed with repeated testing of samples over several days (99.9% agreement with the expected results), and detection was not affected by potentially interfering substances nor by refrigerated sample storage. Conclusion The BioFire BJI Panel is a robust, accurate, and easy-to-use multiplex PCR test capable of detecting many aerobic and anaerobic bacteria, yeast, and AMR genes in synovial fluid specimens. Rapid and reliable molecular detection of possible BJI pathogens may advance the diagnosis and effective management of bone and joint infections. Note This panel has not been evaluated by the FDA or other regulatory agencies for diagnostic use. Disclosures Nicholas Francis, n/a, BioFire Diagnostics (Employee) Laurence Barbier, n/a, Biomerieux (Employee) Caroline Dubost, n/a, Biomerieux (Employee) Elodie Billet, n/a, Biomerieux (Employee) Joel Manwaring, n/a, BioFire Diagnostics (Employee) Josh Southwick, n/a, BioFire Diagnostics (Employee) Tyson Dawson, n/a, BioFire Diagnostics (Employee) Jess Gann, n/a, BioFire Diagnostics (Employee) Kevin Ekins, n/a, BioFire Diagnostics (Employee) Jennifer Arce, MS, BioFire Diagnostics/BioMerieux (Employee) Briana Flaherty, n/a, BioFire Diagnostics (Employee) Harmonie Durand, n/a, Biomerieux (Employee) Chris Cantrell, n/a, Biomerieux (Employee) Elizabeth Amiott, n/a, BioFire Diagnostics (Employee)

No abstract