Discovery may improve anthrax detection

Laboratory scientists with collaborators at Northern Arizona University and Los Alamos National Laboratory, have discovered new DNA regions unique to the bacterium that causes anthrax, potentially providing a way to improve the disease’s detection.

The scientists’ research, conducted during the past year, was presented Wednesday during a session at the general meeting of the American Society for Microbiology at the Salt Palace Convention Center.

In her presentation, Livermore biomedical scientist Lyndsay Radnedge discussed how the researchers have found 20 DNA regions or "signatures" unique to Bacillus anthracis, the bacterium that causes anthrax.

Currently, most DNA-based tests for B. anthracis are based on plasmid sequences, which can be genetically unstable, occasionally yielding false positives or false negatives. A plasmid is a small piece of DNA separate from the chromosome that is transferable between microorganisms.

The new DNA signatures represent an increased repertoire of chromosomal markers that can be used for anthrax detection.

The team’s DNA signatures, which range from about 100 bases of DNA to 800 bases of DNA, were derived from the Ames strain of the B. anthracis bacterium. A robust set of DNA signatures should produce no false negatives for all strains of B. anthracis, and no false positives for closely related bacteria.

"Significantly, the DNA signatures we’ve discovered are found in all of the diverse strains of B. anthracis in the culture collection at NAU," Radnedge said.

The signatures are being checked against — and are so far different from — other strains of Bacillus and related microbes. Of equal importance, they are not represented in the collection of pathogenic non-anthrax Bacillus strains so far examined, decreasing the possibility of false positive results.

During the next two months or so, the DNA signatures will undergo an extremely rigorous screening process to select the optimal signatures. Once screened, the signatures are expected to be submitted to the Atlanta-based Centers for Disease Control for further validation.

Once primers are developed from the B. anthracis signatures, they can be used for rapid, specific, DNA-based pathogen detection on many platforms, including the Livermore-developed portable instrument known as the Handheld Advanced Nucleic Acid Analyzer, (HANAA). They can also be used in a detection system developed by Livermore and Los Alamos scientists that was used at the Salt Lake City Olympic Games. These polymerase chain reaction-based systems can be used to detect and identify pathogens based on their DNA sequence within an hour.

In addition to Radnedge and project leader Gary Andersen, other Livermore biomedical scientists who have worked on the project are Cheryl Strout, Silvia Gamez-Chin, Anne Marie Erler, Julie Avila and Paula McCready.

The DNA signatures were evaluated using the B. anthracis collection of Northern Arizona University microbiology professor Paul Keim, and species closely related to B. anthracis from the collection of Los Alamos National Laboratory.

Additional related research is under way at Livermore by Andersen’s group to characterize other microbes in the environment that might interfere with these assays. In further research, Livermore and Los Alamos scientists are sequencing different pathogens for future signature development to broaden the suite of pathogens that can be rapidly detected.

The signatures for B. anthracis were developed under funding primarily from the NNSA’s Chemical and Biological National Security Program as well as the DOE Office of Biological and Environmental Research and other federal sponsors.