Objectives: To examine susceptibilities of Bacillus anthracis and related species to 24 antimicrobials using and concurrently comparing two methods. cereus, three were resistant to clindamycin and one was resistant to clarithromycin and clindamycin. One B. mycoides was intermediately resistant to quinupristin/dalfopristin and meropenem and one was clindamycin-resistant. All B. pseudomycoides were clindamycin-resistant with one quinupristin/ dalfopristin-resistant. Two B. mycoides/pseudomycoides were intermediately resistant to quinupristin/ dalfopristin and clindamycin and a third was intermediately resistant to clindamycin alone. All isolates were susceptible to chloramphenicol, ciprofloxacin, gatifloxacin, gentamicin, levofloxacin, linezolid, moxifloxacin, rifampicin, streptomycin, tetracycline, tigecycline and vancomycin. MethodsConclusions: This paper expands the list of therapeutic or prophylactic antimicrobials potentially effective against B. cereus group isolates using two testing methods that produced comparable results.

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.

In order to cause the disease anthrax, Bacillus anthracis requires two plasmids, pX01 and pX02, which carry toxin and capsule genes, respectively, that are used as genetic targets in the laboratory detection of the bacterium. Clinical, forensic, and environmental samples that test positive by PCR protocols established by the Centers for Disease Control and Prevention for B. anthracis are considered to be potentially B. anthracis until confirmed by culture and a secondary battery of tests. We report the presence of 10 genes (acpA, capA, capB, capC, capR, capD, IS1627, ORF 48, ORF 61, and repA) and the sequence for the capsule promoter normally found on pX02 in Bacillus circulans and a Bacillus species closely related to Bacillus luciferensis. Tests revealed these sequences to be present on a large plasmid in each isolate. The 11 sequences consistently matched to B. anthracis plasmid pX02, GenBank accession numbers AF188935.1, AE011191.1, and AE017335.3. The percent nucleotide identities for capD and the capsule promoter were 99.9% and 99.7%, respectively, and for the remaining nine genes, the nucleotide identity was 100% for both isolates. The presence of these genes, which are usually associated with the pX02 plasmid, in two soil Bacillus species unrelated to B. anthracis alerts us to the necessity of identifying additional sequences that will signal the presence of B. anthracis in clinical, forensic, and environmental samples.Long considered a biowarfare agent, Bacillus anthracis was used in 2001 in an act of bioterrorism. After bacterial endospores were placed into envelopes and delivered by the United States postal system to unsuspecting victims, 22 people were infected of whom 11 developed inhalational anthrax and 5 died (14, 15). As a result, first responders delivered hundreds of thousands of environmental specimens to laboratories across the nation that are part of the National Laboratory Response Network (LRN), a coordinated system of sentinel, reference, and national laboratories established by the Centers for Disease Control and Prevention (CDC) (6, 7). Of the three Florida Department of Health (FDOH) laboratories with biosafety level 3 facilities designated to receive these specimens, the FDOH Tampa Laboratory received and analyzed 1,046 environmental samples from first responders across west-central Florida over the last 3 months of 2001. Among these specimens, 19 loose powders or swab samples of powders that were brought in by local law enforcement officers and considered plausible threats initially tested positive for potentially carrying a B. anthracis isolate.Because of the initial positive results, the 19 powders and swabs were cultured and all isolates were tested further using the LRN tests, consisting of nonspecific phenotypic physical/ biochemical tests and specific molecular methods for B. anthracis such as gamma phage susceptibility testing, cell wall and capsule detection by direct fluorescence antibody tests (DFA), and amplification of targeted DNA sequences by PCR (31). After more tests...

Bacillus anthracis spores have been used as a biological weapon in the United States. We wanted to develop a safe, rapid method of sample preparation that provided safe DNA for the detection of spores in environmental and clinical specimens. Our method reproducibly detects B. anthracis in samples containing <10 spores.Bacillus anthracis spores, a recent threat, can remain dormant for years while retaining full virulence (5,10,13,20,21,22; S. Endicott, E. Hagerman, and M. Furmanski, Letter, JAMA 284:561-562, 2000). Powders and environmental samples, the most common nonclinical specimens, and nasopharyngeal swabs are submitted to designated laboratories in the national Laboratory Response Network (LRN), which has facilities to work safely on the specimens (6,19,23). High numbers of specimen can overwhelm a laboratory's capability and capacity to perform the tests in a timely fashion. From October to December 2001, the Florida Department of Health's three LRN laboratories each received hundreds of samples per day, which revealed the need for a method to render the samples harmless so the DNA could be safely extracted under biological safety level 2 conditions, to alleviate the bottleneck, and to decrease the turnaround time to the final result. Specimens may contain Ͻ10 spores, and the isolation and identification of B. anthracis may take days (5, 15). The purpose of this work was to develop a method of sample preparation that would provide safe DNA for the detection of Յ10 B. anthracis spores.Previous studies used sonication probes in open tubes and cartridges to hold samples, a method that necessitated the use of a biological safety level 3 environment and sterilization between each sample preparation (2, 3, 7). Dang et al. demonstrated that DNA from autoclaved spores was usable for PCR assays (9). No one explored the sensitivity of these methods or if combinations of methods could be used.(A preliminary report of this work has been presented previously [V. A. Luna, M.