Trail of Odd Anthrax Cells Led FBI to Army Scientist

By Joby Warrick
Washington Post Staff Writer
Monday, October 27, 2008

In late October 2001, lab technician Terry Abshire placed a tray of anthrax cells under a microscope and spotted something so peculiar she had to look twice. It was two weeks after the country's worst bioterrorism attack, and Abshire, like others at the Army's Fort Detrick biodefense lab, was caught up in a frenzied search for clues that could help lead to the culprit. Down the hall, Bruce E. Ivins, the respected vaccine specialist, was looking, too.

Abshire focused her lens on a moldlike clump. Anthrax bacteria were growing here, but some of the cells were odd: strange shapes, strange textures, strange colors. These were mutants, or "morphs," genetic deviants scattered among the ordinary anthrax cells like chocolate chips in a cookie batter.

Unknowingly, Abshire had discovered a key to solving the anthrax case. But it would take nearly six years to develop the technology to allow FBI investigators to use it.

Ultimately the evolving science led investigators to Ivins and a strikingly original collection of anthrax spores that became the focus of the FBI's probe. In a series of interviews over the past month, FBI agents and scientists described, in ways that the bureau has not previously revealed, how the pieces of the forensic puzzle came together -- often in Ivins's very shadow -- and how they eventually concluded that the eccentric vaccine specialist was the culprit.

Ivins, the FBI discovered, had spent more than a year perfecting what agents called his "ultimate creation" -- his signature blend of highly lethal anthrax spores -- and guarded it so carefully that his lab assistants did not know where he kept it. When the FBI later asked Ivins for anthrax spores from his lab, he deliberately bypassed his prize spore collection, agents said, and gave them a false sample.

Ivins's talents also helped give him away, they said. Exceptionally pure concentrations of anthrax spores were Ivins's trademark and placed him in an exclusive class. In the end, the FBI concluded, he was the only one with access to the deadly spores who also possessed the skills and equipment needed to create the extraordinarily powerful bioweapon that was mailed to U.S. Senate offices and news organizations in the fall of 2001.

"He wasn't an expert. He was the expert," said a senior FBI investigator, who answered questions about the still-open case on the condition of anonymity.

Yet the forensic search that started in the glare of Abshire's microscope turned out to be far more arduous and costly than anyone could have predicted. Conducted almost entirely out of the public eye, it was a journey that required use of techniques that had never been tried in a criminal investigation. Some of the technology needed to solve the case had not been invented. And the FBI's top science advisers were warning that the effort would fail.

"We were looking for a needle in a haystack," said Scott Decker, a geneticist who became the FBI's science team leader, "and no one knew if there was even a needle there."

Many outside experts and some lawmakers dismiss the government's case against Ivins as circumstantial, while Ivins's former colleagues and friends argue that he was incapable, technically and constitutionally, of committing an act of mass murder. "Bruce Ivins was a victim of a vicious plot," said Ayaad Assaad, a toxicologist who once worked with Ivins at Fort Detrick, in Maryland.

The questions have prompted an independent review of the FBI's forensic case by a panel of the National Academy of Sciences. In an Oct. 16 letter to the academy, Rep. Rush D. Holt (D-N.J.), a member of the House intelligence committee, asked the panel to investigate whether the bureau's scientific discoveries were "inconsistent with the FBI's conclusions."

The FBI defends its case against Ivins as well as the seven years it took to solve the crime -- an unavoidable delay, officials say, given that the bureau had to invent an entirely new investigative field, microbial forensics, to accomplish it. Investigators say more evidence will be revealed in the coming weeks, some of it in peer-reviewed scientific journals and the rest in documents that will shed new light on Ivins himself. "A lot of [the investigators] probably know Dr. Bruce Ivins better than his own family," the senior investigator said.

But while bureau officials view the evidence against Ivins as overwhelming, any chance at full certainty was lost when Ivins took his own life in July, said Ed Montooth, the FBI special agent in charge of the investigation the bureau dubbed "Amerithrax."

"We were truly disgusted after we knew he had killed himself," Montooth said, "because we knew the only way we'd have justice was to be in court."

Ivins's 'Ultimate Creation'

It was intended for garden-variety animal experiments, but the collection of anthrax spores known as RMR-1029 was anything but ordinary. Ivins, its creator, had devoted a year to perfecting it, mixing 34 different batches of bacteria-laden broth and distilling them into a single liter of pure lethality.

The finished product, a muddy, off-white liquid in a glass flask the size of a small coffee pot, was the greatest single concentration of deadly anthrax bacteria in the country, FBI investigators said.

Ivins began work on it in 1996 with the goal of creating a large repository of highly virulent Bacillus anthracis spores that could be used by his fellow scientists at the U.S. Army Medical Research Institute for Infectious Diseases, or USAMRIID, for years to come. To measure the effectiveness of new anthrax vaccines, the drugs have to be tested against a potent form of bacteria that remained the same from one experiment to the next.

The art of "spore preparation" is a tedious job often relegated to novices and technicians. But Ivins, a veteran researcher with decades of experience, was naturally good at it, according to FBI officials and USAMRIID scientists. With RMR-1029, Ivins established an anthrax gold standard.

"It was his ultimate creation," said Jason D. Bannan, an FBI microbiologist assigned to the Amerithrax case. "This was the culmination of a lot of hard work."

But Ivins could not have known that RMR-1029 contained genetic mutants, in relatively high numbers. A batch of spores like RMR-1029 might be expected to contain, at most, one mutated variant. But Ivins's flask, because of its unusual pedigree, contained five.

Without knowing it, Ivins had provided the FBI a rare clue that in time would lead them to his lab.

The Hunt for Mutants

Within days after letters laden with anthrax microbes reached New York and Washington, Fort Detrick had become the scientific epicenter of the FBI investigation. It had a resident corps of anthrax specialists as well as numerous biocontainment laboratories where deadly microbes could be handled. Dozens of the lab's white coats -- including Ivins -- joined FBI scientists in the search for the culprit.

The experts quickly established that the attacker had used the so-called Ames strain, a virulent form of anthrax bacteria that was the strain of choice within the Army's biodefense complex. They also concluded, by November 2001, that the attack strain had not been altered: The spores were not drug-resistant and contained no foreign additives to make them more lethal. It was a detail the FBI would not disclose publicly for six years.

Top FBI officials hoped that science could provide a link to the bioterrorist, but they soon grasped the difficulty of the task. They searched for traces of human DNA in the anthrax powder, and in the envelopes, but found none.

But what if there were something unique about the spores themselves? The anthrax experts who served as consultants weren't encouraging. Ames-strain bacteria was essentially identical wherever it was found, the advisers said.

"There is Ames and Ames and Ames," said Decker, the science team leader. If the investigators could find even one clear marker, "we'd be pretty lucky," he said.

The breakthrough the FBI sought came not from a big-name scientist but from a technician who had spent years studying anthrax bacteria under a microscope.

Terry Abshire had been tasked with growing colonies of anthrax bacteria from spores recovered from one of the mailings. When the 56-year-old Frederick resident studied the cells, she noticed that a few colonies were different in subtle ways, so she allowed the bacteria to grow for a longer period so as to check again.

"They looked different -- different colors, different textures," said Richard Langham, an FBI scientist who was assigned to work at the Fort Detrick lab. He said it was Abshire's 20 years of experience that allowed her to spot the subtleties.

"A new postdoc working with anthrax probably would not have noticed," he said.

The FBI was fortunate: Not only were there multiple mutations among the attack strain, but they also were the kinds that led to easily detectable physical changes.

Once the mutants were found, FBI scientists could begin pinpointing the subtle alterations in the spores' DNA code that caused them to morph. It took scientists until early 2004 to find all the altered genes and to develop special tests to help find the mutations in other samples of anthrax bacteria.

Even a single genetic mutation is unique, and the FBI had discovered five in the spores used in the anthrax attacks. Now investigators just needed to find the same genetic fingerprint in anthrax spores in the possession of a presumed bioterrorist, somewhere in the world.

The Search Narrows

While some FBI scientists were analyzing genetic mutations, others were scouring the planet for repositories of Ames-strain bacteria. To their surprise, Ames turned out to be quite rare, with only 15 U.S. institutions and three foreign ones possessing live, virulent Ames.

Samples of Ames were collected and added to a repository the FBI had established at Fort Detrick. In a process that ended only in late 2006, bureau scientists picked up 1,072 samples of anthrax bacteria and tested each for mutations identical to the ones in the bioterrorist's letters.

By early 2007, the FBI had a few direct hits, yet the results were perplexing. Each of the matching samples could be traced to Ivins's lab, but only indirectly. Ivins had shared anthrax bacteria with other researchers, and some of them had turned in samples containing the mutations. But Ivins's lab tested clean. He had given the FBI a vial of anthrax bacteria with no mutations.

Bureau officials decided to look again. Ivins, they found, had turned in two samples from his lab. The first was rejected because Ivins had not followed the FBI's detailed instructions and had used the wrong type of test tube. Ivins was asked for a second sample, and this time, investigators deduced, he tried to deceive them. He bypassed his exclusive reserve of spores -- the one he used in his experiments -- and turned in something different. Later tests to determine the origin of the substitute sample came up blank. "Our experts could not tell us where it came from," a senior investigator said.

FBI agents searched Ivins's lab and found the flask labeled RMR-1029. Tests showed it was pure, highly concentrated Ames anthrax bacteria, with genetic mutations identical to those in the attack strain.

Back at the bureau's Washington field office, agents were reconstructing the history of RMR-1029. A giant flow chart, covering most of a wall, recorded each discovery about the origins of the spores and what Ivins did with them. But the agents wondered: Could others, besides Ivins, have gotten access to the flask of spores?

The question drives much of the skepticism about the FBI's case. At a news conference in August, bureau officials estimated that as many as 100 people potentially had access to the biocontainment lab where Ivins kept his collections. Investigators have maintained that other possible suspects were ruled out, but they have never explained how. It is one of the gaps that independent experts and lawmakers have raised since Ivins's death.

In interviews, FBI officials said the list of 100 names included USAMRIID scientists as well as anyone with even a tenuous connection to Ivins's lab, such as visitors or janitors. Each person was investigated, though most could not have gotten to the spores under any reasonable scenario the investigators could construct.

For one thing, no one besides Ivins seems to have known where they were kept. The plain, triangle-shaped storage flask was one of many kept in plastic tubs inside a refrigerated storage room in Ivins's restricted lab. It had only a handwritten label -- RMR-1029, shorthand for "reference material received, No. 1029." When spores were needed for experiments, Ivins alone would retrieve them. "His own people who worked with him on a daily basis didn't know which flask it was," Langham said.

Initially, agents thought Ivins divided his spores into two flasks and kept one in a different building, which would have increased the number of people with potential access. That belief was based on a lab notebook entry that turned out to be erroneous, agents said.

Still, dozens of people were cleared at various times to enter USAMRIID's Building 1425, where Ivins worked and kept his spore collection. Each had to be investigated, even those who lacked the basic knowledge to handle highly lethal bacteria. "An animal handler might have had access," said Bannan, "but he would not have had the capability. And he probably would have expired by now."

Simply obtaining the microbes would have been only the first hurdle. The FBI is convinced that the bioterrorist did not merely steal spores from RMR-1029, but also regrew them and converted them into a highly concentrated powder. And then he repeated the process.

Differences between the two grades of anthrax powders used in the attacks -- the earlier batch sent to New York news outlets was coarser and darker than the powder mailed to the Senate -- confirm that there were at least two production runs. Bureau officials knew they were looking for someone who had repeated access to Ivins's flask as well as talent for sophisticated spore preparations.

The list of suspects narrowed, officials said, until only one was left: Ivins. Ivins alone created and controlled the distinctive collection of anthrax cells that provided the seeds for the attacks. And he was the undisputed master at manipulating the bacteria into dense concentrations of deadly spores. While graduate school microbiologists could have performed most of the tasks, Ivins had the experience and the "good set of hands" required to achieve a spore preparation of such quality, a government scientist said.

"When you go to the true experts and ask them how many people can develop [anthrax spores] into something with this purity and this concentration, they shake their heads," said Montooth, the lead Amerithrax investigator. "Some will say there are perhaps six. Others will say maybe a dozen."

Ivins normally worked with liquid anthrax spore solutions, not dry powders, investigators acknowledge. Ivins's colleagues insist that he had no experience with "dry aerosols" of anthrax spores and would not have known how to make them.

But drying the spores turned out to be no obstacle at all, FBI scientists said. It required only one more step, using a common laboratory machine known as a lyophilizer. Ivins had one in his lab.

"Because he grew spores on a daily basis, he was in a position to make [the powder], and no one would be the wiser," Montooth said.

Would the FBI's evidence have stood the challenge of a court trial? Paul Kemp, a lawyer who represented Ivins, dismissed the government's case as an "orchestrated dance of carefully worded statements, heaps of innuendo and a staggering lack of real evidence."

Bureau officials said they feel cheated at being deprived of the opportunity to prove otherwise. The resentment spilled over in the early hours of July 27, when investigators first learned of Ivins's drug overdose, said Montooth, who recalled getting 253 text messages from fellow agents within minutes after the news broke. For the next week, the members of his team barely slept, Montooth said, because they knew Ivins's suicide meant they "would never get to do what we wanted to do, which was to go to court."

The only solace, he said, came on the day the Amerithrax team sat down with family members of the victims of the attacks. In an FBI conference room, Montooth laid out the still-secret details of the seven-year investigation.

"They thanked us," Montooth said, recalling the families' reaction. "They said, 'We believe you got the right guy.' "

Staff researcher Julie Tate contributed to this report.