A decade ago this month, a microbiologist at Northern Arizona University, in Flagstaff, took a special delivery from the US government. Federal investigators wanted the scientist, Paul Keim, to identify the anthrax that appeared in letters mailed to news organizations and US lawmakers. Overnight, he used PCR to determine that the anthrax sent was the Ames strain, commonly used in research—but that was just the beginning of a scientific investigation that would catapult the still wet-behind-the-ears science of microbial forensics to the forefront of the criminal inquiry.

Ten years on, Keim's PCR-based technique seems downright quaint in comparison with modern, speedy DNA sequencing. “In a lot of ways we've matured,” says Bruce Budowle of the University of North Texas Health Science Center in Fort Worth. But there are challenges ahead, adds Budowle, who retired in 2009 from the US Federal Bureau of Investigation (FBI), where he was involved in the anthrax studies as a senior scientist in the laboratory division: “In a lot of ways, we've got a long way to go... We haven't grown in the interpretation of the results and what they might mean.”

Overall, the country has improved in many aspects of preparedness. The US government spent $60 billion on biodefense over the last decade, including the 2004 founding of Project BioShield. The $5.6 billion initiative, managed by the government's Biomedical Advanced Research and Development Authority (BARDA) since 2006, is charged with stockpiling medicines and funding research on new therapies that could be used in instances of bioterrorism. And the spending continues: last month, BARDA awarded a five-year $68 million contract to the New Jersey company Elusys Therapeutics to develop a prophylactic treatment against anthrax.

At the same time, investments continue in the area of microbial forensics, which encompasses chemical analysis, carbon dating and microscopy in addition to DNA sequencing. Shortly after the 2001 anthrax scare, the FBI and Department of Homeland Security created the National Bioforensic Analysis Center, housed at the US Army Medical Research Institute for Infectious Diseases (USAMRIID) in Fort Detrick, Maryland. (Ironically, USAMRIID employed Bruce Ivins, who committed suicide in 2008 around the time he was fingered by the FBI as the culprit behind the anthrax letters.) Researchers there have converted medical tests—intended to identify pathogens in blood samples—to work for other kinds of evidence such as soil samples, carpet fibers and clothing.

Although next-generation DNA sequencing has radically accelerated the decoding of pathogen genomes since 2001, analysis techniques have not kept pace. “The biggest challenge is databases and statistical power,” says Jacques Ravel, a microbiologist at the University of Maryland School of Medicine in Baltimore. Ravel helped sequence the 2001 anthrax strain while at The Institute for Genomic Research in Rockville, Maryland. If the next biological strike involves, say, bubonic plague, sequencing the attack strain will be no good unless investigators have a vast array of plague bacteria for comparison.

Virtual reality

The FBI is starting with a “virtual collection,” says Jason Bannan, senior biological programs advisor for the Bureau's laboratory in Quantico, Virginia. That is, an FBI working group is figuring out what strains are available in collections across the country, so it can quickly access them when needed. Once this work is complete, the Bureau will proceed to calling in samples for a physical strain library.

Over the past decade, government leaders have also focused on how to better integrate the biodefense efforts of disparate agency cultures at, for example, the FBI and the US Centers for Disease Control and Prevention. In 2009, the White House issued a microbial forensics strategy, noting that current capabilities only “scratch the surface” of what's needed. Key recommendations included better technology and interagency coordination. To address these issues, last year the government established the Interagency Microbial Forensics Advisory Board to coordinate research and development among different departments.

The various agencies “are generally singing off the same page now,” says Thomas Inglesby, director of the Center for Biosecurity at the University of Pittsburgh Medical Center in Pennsylvania.

But some experts worry that the collaboration has not yet trickled down to local health authorities, too. At an Institute of Medicine workshop on pathogen surveillance held in mid-September in Washington, DC, Joe Gibson, director of epidemiology for the Marion County Public Health Department in Indianapolis, Indiana, noted that patient privacy laws and a general reluctance of schools and health departments to share epidemiological data hinder his department's access to surveillance data and suggested that building trust and clarifying how data will be used would help foster cooperation.

Even as scientists hammer out the details of bioterrorism countermeasures, they are already using advances in the microbial forensics field to deduce the origins of naturally occurring threats to public health. Biologists have used these techniques to ferret out the sources of severe acute respiratory syndrome (SARS), recent E. coli infections in Germany and the cholera outbreak in Haiti following the 2010 earthquake.

“On the whole, there's a better appreciation of what microbial forensics can and can't deliver,” Inglesby says. The science can only provide a clue, not a conclusion. Good old-fashioned detective work, he adds, is still necessary to catch the bad guy.