Unraveling anthrax

Los Alamos National Laboratory's Bioscience Division researchers have developed technologies that can uniquely identify the origins of biological organisms based on information in the DNA.

The technique, known as Amplified Fragment Length Polymorphism (AFLP), is used to create a library of genetic profiles for hundreds of different Bacillus anthracis strains, the organisms that cause anthrax in livestock and humans. Specific DNA fragments from the AFLP profile are then used to design a new set of fragments, known as polymerase-chain-reaction, or PCR, primers, that can specifically detect these fragments in complex samples.

Based on the analysis of tissue samples, Los Alamos researchers proved that the victims of the 1979 anthrax outbreak in the former Soviet Union were infected with at least four different strains of B. anthracis . This provided definitive evidence that the deaths were not caused by a natural infection. It was later revealed that the deaths had been caused by the accidental release of B. anthracis spores from a Soviet military biological research facility suspected by western intelligence experts of producing large quantities of spores. More recently, Los Alamos DNA analysis of samples from Iraq in the aftermath of the Gulf War was directly linked to Iraq's disclosure of an offensive biological warfare program that included the use of B. anthracis.

A team of Los Alamos researchers, including Paul Jackson, Karen Hill and Larry Ticknor, uses AFLP markers as genetic characters to determine the relationships among bacterial isolates. AFLP DNA fragment libraries have been developed for a large number of Bacillus species and a smaller number of pathogens. Researchers use the libraries to analyze medical, veterinary, forensic and environmental samples to determine their microbial content. The goal of the project is to generate an AFLP profile from a sample containing unknown microbes, compare it electronically to all the archived profiles, and thereby determine its phylogeny, and possibly, its exact identity and geographic origin.

The technology recently was used to properly classify a misidentified pathogen that severely infected the wound of a French peace-keeping soldier wounded in Bosnia. The microbe isolated from the infection was misidentified as a B. thuringiensis strain commonly used as a biopesti- cide. This caused considerable public concern about the use of this species for insect control. Los Alamos analyses showed that the pathogen actually was related closely to B. anthracis, explaining why the soldier suffered a serious infection, and was only remotely elated to the B. thuringiensis strains used as biopesticides.

B. anthracis, discovered in the 1870s, was the first organism shown to cause a particular disease. It causes anthrax in animals, mostly cattle, horses, goats, sheep and in humans. Cutaneous anthrax in humans occurs most frequently on the hands and forearms of persons working with infected livestock or products from these animals. It results in sores that develop coal-black scabs. The term anthrax comes from the Greek word for coal.

Cheryl Kuske, a technical staff member in the Lab's Bioscience Division, is interested in understanding the relationships between the pathogenic B. anthracis bacterium and non-pathogenic close relatives of B. anthracis that are naturally present and widespread in the environment. She has been comparing sets of genes from the pathogen with DNA from several different nonpathogenic Bacillus species to identify genes that are unique to the pathogen. These genes have potential for use in rapid, DNA-based detection strategies for B. anthracis in environmental samples for use in forensic analysis.

Another Bioscience Division project addresses the host-pathogen interface. Michael Altherr and Tom Brettin's work will create novel databases and integrated computational tools to investigate the interaction between pathogens and human cells, including B. anthracis. Several key technologies have emerged through this project that allow the analysis of the entire transcriptional and translational components of a cell. The knowledge gained from these studies may someday help lessen the consequences of exposure to the pathogens and aid in the development of vaccines.