Performance Assessment of Three Commercial Assays for Direct Detection of Bacillus anthracis Spores

Abstract

Bacillus anthracis, the cause of anthrax, has been used as a bioterrorism agent. Because the isolation and identification of B. anthracis by culture can take days, first response units (hazardous materials [HAZMAT], firemen, police, and hospital personnel) desire a quick and easy test that can be done in the field to detect possible B. anthracis contamination (1, 4). To our knowledge, there are no peer-reviewed published data on commercially available kits that could guide first responders in their search for such rapid detection methods. We tested three lateral flow immunoassay kits that are designed to test for B. anthracis at 10 4 to 10 5 spores per sample: (i) Anthrax BioThreat Alert (BTA) test strips (Tetracore, Gaithersburg, Md.), (ii) BioWarfare Agent Detection Devices (BADD) (Osborne Scientific, Lakeside, Ariz.), and (iii) Anthrax (spore) SMART II (New Horizons Diagnostics, Columbia, Md.). These tests require little technician time and training, and results are available within 15 min.This study was conducted at the Florida Department of Health Laboratory in Tampa, Fla., and employed B. anthracis Pasteur (CDC BC 3132) and Bacillus cereus and Bacillus thuringiensis from our culture collection. Spores were added to the buffer provided by the manufacturer to achieve 10 2 to 10 6 (B. anthracis) or 10 6 (B. cereus and B. thuringiensis) spores per sample. The range of 10 2 to 10 5 for B. anthracis spores was chosen in order to include the manufacturer's claims of sensitivity. We also tested 10 6 spores in order to achieve a clear, easy-to-read positive result. Because one of the kits did not consistently detect spores at the upper limit (10 5 ), we tested 10 6 spores more than once to see if detection was consistent. All tests were performed according to the manufacturer's instructions and were allowed to proceed for 15 min, although the positive results were recognized within 5 min. Spore concentrations were verified by viable plate counts on Trypticase soy agar (Remel, Lenexa, Kans.) in duplicate.

Development of Antibodies against Anthrose Tetrasaccharide for Specific Detection of Bacillus anthracis Spores

“…Such assays have not been well described before. Besides PCR-based detection, only a few immunological detection systems, such as lateral-flow immunoassays, are available, but they offer detection limits that range from only 10 5 to 10 6 spores/ml (12,19). Unfortunately, no information on the target antigens of the antibodies used in those assays is available.…”

Anthrax Spore Detection by a Luminex Assay Based on Monoclonal Antibodies That Recognize Anthrose-Containing Oligosaccharides

“…The sensitivity of spore detection by the Luminex assay was substantially increased compared to the corresponding capture ELISA, with the detection of 10 3 to 10 4 spores per ml. Currently available immunological detection systems offer higher detection limits (11,12,14). The platform can be also used in multiplex assays, and we are currently evaluating the simultaneous detection of different biothreat bacteria in mixed samples.…”

Diagnostic Probes for Bacillus anthracis Spores Selected from a Landscape Phage Library

“…The recombinant phage carry the gene necessary for tetracycline resistance, allowing only those cells infected by phage to form colonies on the plate (26 ). The biological titer of these recombinant phage, expressed as colony-forming units (CFU) 4 is typically 20-fold lower than the physical titer (virions/mL).…”

“…(2,3 )], but to date none has been adapted for real-time continuous monitoring of the environment (4 ). Immunoassay-and biosensorbased (5 ) detection systems are the best prospects for continuous monitoring systems, but they require specific, selective, and stable diagnostic probes with which the pathogen can be detected.…”