The Anthrax Letters: The Attacks That Shocked America (29 page)

With a small container in hand, Mann returned to the level-3 lab, where she injected the blood with anthrax spores. Connell was present for the first of the injection sessions. “I knew what to expect, but still the next morning I was shocked,” Connell said. During the night the bacteria had split open the blood cells. “It was black. The cells had all been lysed. The devastation was so fast.”

Eventually, anthrax and plague genes, as well those of the other agents in the study, would be tested against RNA in human blood. (RNA, ribonucleic acid, is a close structural cousin to DNA.) Only then will it become clear whether the investigation has succeeded, that is, whether the genes that become active are distinct to each organism. The central importance of RNA to the experiment arises from an understanding of its function in gene expression. And this all became possible because of one of the great discoveries of the 20th century.

For more than 100 years, the fundamental characteristics of living things—the shape of a maple leaf, the color of an eye, the infectivity of an anthrax bacterium—have been understood as expressions of microscopic units called genes. Composed of deoxyribose nucleic acid, or DNA, the structure of the gene was identifed in 1953 by James Watson and Francis Crick at Cambridge University. Perhaps the most astonishing aspect of their discovery was the simplicity of the structure. The myriad characteristics of caterpillars, elephants, yeast, ivy, tulips, and amoebas are essentially described by an alphabet of the same four letters. A, T, C, and G comprise the fundamental structures of all genes. They represent four DNA bases—adenine, thymine, cytosine, and guanine.

The bases are joined in matching pairs; “A” is always paired with “T,” and “C” is always with “G.” The base pairs occur in irregular sequences along a lengthy backbone of sugar and phosphate groups. (The combination of sugar, phosphate, and a base is called a nucleotide.) The structure of DNA resembles a ladder whose sides are made of sugar and phosphates and whose rungs are composed of base pairs.

Splitting the ladder lengthwise results in two separate lines studded with half rungs. Each half rung represents a single base. Thus, if a segment were chopped out from one of the sides, it might contain a sequence of bases that reads TGACCAT. The complementary half rungs on the other side would read ACTGGTA. But a sequence of seven base pairs, as in this example, would make up only a small segment of a gene. Human genes may run as long as several thousand base pairs. It is these lengthy, though uncomplicated, sequences that produce the magic of life.

Watson and Crick immediately understood the profound implications of their discovery despite the restrained language of their published description. Appearing in
Nature
, their article was simply titled “The Molecular Structure of Nucleic Acids” and concluded with a coy observation: “It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.” Only later in his book,
The Double Helix
, did Watson reveal the fullness of their excitement. After the moment of discovery, Watson said that Crick rushed over to a local pub “to tell everyone within hearing distance that we had found the secret of life.”

In the years since their discovery, that secret has led to avenues of research that had previously been unimaginable. It has also produced debates about the propriety of the research. In the 1970s the discovery of techniques to insert DNA from one species into another prompted biologist Erwin Chargaff to wonder whether investigators had “the right to counteract, irreversibly, the evolutionary wisdom of millions of years.” Such concerns have diminished, however, and gene splicing has become a common research and therapeutic tool. For example, insulin is now produced by bacteria that have been genetically programmed to do so. The human gene responsible for the manufacture of insulin is pasted into the DNA of
Escherichia coli
, a common bacterium that inhabits the human gut. The rapidly dividing bacteria are thus transformed into insulinproducing factories.

Still, the potential for harm from the technology does exist, including in the military sphere. Genes could be spliced into microorganisms that would turn them into super killers that are impervious to any known defense, which is exactly what inspired Soviet activities in the 1970s and 1980s. The sad paradox is that in 1972, the year that the Soviet Union, the United States, and other countries established the treaty to ban germ weapons, the Russians expanded their development program. According to Ken Alibek, a former director of the Soviet program, he and his co-workers ignored the Biological Weapons Convention entirely and assumed the United States was also cheating.

With gene-splicing technology at hand, the Soviets embarked on a program to genetically engineer anthrax, plague, tularemia, smallpox, and other pathogens to make them resistant to antibiotics and vaccines. Only in the mid-1990s, with information provided by Alibek and other defectors, did Americans learn how extensive the Soviet program had been. Until the demise of the regime in 1991, some 60,000 people had been engaged in Soviet germ warfare work.

Meanwhile, in other laboratories throughout the world, advances in legitimate gene research kept apace. The genetic underpinnings of many diseases were identified. Some diseases, such as cystic fibrosis, which can provoke suffocating secretions of mucus in the respiratory tract, are attributable to a single gene. Others, like diabetes and cancer, appear to arise from mutations in several genes.

This knowledge became possible only with an understanding of the structure of the gene. Similarly, the development of research techniques, such as the polymerase chain reaction, depended explicitly on the ability to split strands of the genetic material. Unzipping the DNA ladder is the key to PCR’s power to copy a single DNA sequence many million-fold. The new technology also facilitated the cloning of individual genes and the ability to gauge their activity.

In the 1980s a method was developed to anchor a gene segment to a solid surface and stain it with a radioactive or fluorescent dye. The activity of a gene could thus be determined by looking for an interaction between the dyed segment and the target gene from another specimen. Fluorescence changes would signal that the gene was turned on or off.

The actual expression of a gene first involves an unzipped strand of DNA that attracts chemical building blocks to form a complementary strand of RNA. The structure of the RNA is only slightly different from that of DNA, but it has the capacity to direct the formation of a protein. The protein is the end result of gene expression and is what comprises an organism’s distinctive characteristics. Thus, if there is active RNA in a blood sample, a gene has been turned on. It is expressing itself.

By the end of the 20th century, efforts to map the entire human genome were nearly complete. The sequencing of every human gene revealed that a person carries perhaps 30,000 genes made up of 3 billion base pairs of DNA. At the same time, a vastly more powerful research capability had been developed, the microarray. Through miniaturization techniques like those used to manufacture computer chips, thousands of DNA probes could be affixed to an area on a glass slide no larger than a postage stamp. Using fluorescentbased detection, the ability to identify gene activity had suddenly been multiplied 20,000-fold.

“I like working with the technology,” Amol Amin told me. Amin, who holds a Ph.D. from his native India, has, since mid-2001, worked in Nancy Connell’s lab, where he applies his expertise on microarrays. “We are using the oligonucleotide technique to make the chips,” Amin explained, using a term that refers to a short chain of nucleotides, or DNA bases. The technique is one of several now being used to make microarrays.

Amin works in the fourth-floor laboratory of UMDNJ’s new research building on Norfolk Street in Newark, a half mile from the medical school. The red brick structure, called the International Center for Public Health, was completed in the spring of 2002. Much of Connell’s research has been moved there, though not the work dealing directly with biowarfare agents. That is done exclusively at the BSL-3 lab in the basement of the medical school. At the ICPH, Amin handles no pathogens, only the RNA from blood samples that have been infected by the bacteria.

In a small room across the corridor from his laboratory, Amol Amin stood before a 6-foot-wide machine. Labeled “Gene Machine Robot,” the apparatus was invented in the late 1990s. The left section of the machine is comprised of several 2-foot-tall metal columns, each containing 10 small circular shelves the size of a computer disk. Sitting on the shelves were plastic plates indented with 384 tiny wells. “Each well contains segments of a human gene immersed in liquid,” Amin explained. “And the gene segments are all 70 base pairs long.” The gene segments are single stranded DNA to be imprinted on glass slides. The RNA from the blood samples that Mann and Trzop prepared in the BSL-3 lab will later be bathed over the slides to find their complementary DNA imprints.

To the right of the shelves was a flat deck that can accommodate a hundred 1 × 3 inch microscope slides. Dressed in a blue sweater, faded jeans, and sneakers, Amin’s latex gloves were the only sign of attire suggestive of a laboratory researcher. He gingerly placed the last of the hundred blank slides on the deck. He turned to the computer behind him and pressed three keys. After a few seconds, the robot began to act. A 10-inch-long red block glided forward from the rear. Its bottom was lined with pins that were spaced precisely to match the tiny wells. The block of pins stopped and hovered over the wells. Like a family of ducks that bob in unison, the pins descended. They came up, paused, and moved laterally, to the right over the glass slides. Again, they descended, leaving imprints on the slides, rose, and retreated to the wells.

Printing the DNA to the slides was but one step in the experiment that Nancy Connell was overseeing. Months of work lay ahead. But 4 months after the project began, Amin and the others on the staff felt a sense of accomplishment. This was the first transfer of genes to slides. These slides will provide the baseline for possible reactions of genes from blood infected with anthrax, plague, and the other organisms. Amin, whose broad smile pierced through a black beard and mustache, intoned, “I have waited for this day.” He was about to start working with the first batch of RNA recovered from the blood of volunteers.

The templates that Amin was creating were the core of the experiment. Each of the 20,000 dots on his microarrays was a composite of genes from two samples. “Right now, I am making slides with RNA from blood that is uninfected and from blood that has been infected with Vollum anthrax,” Amin said. The slides were washed with fluorescent dye that soaks into the gene segments. If a gene in one sample is active, it will glow red. If active in the other sample, it will glow green. Amin smiled. “But the samples are mixed with each other, and if comparable genes in both samples are active, the gene will glow yellow.” He walked over to a computer to demonstrate. “Here are the genes magnified,” he said. The computer screen was filled with rows of tightly packed traffic lights—circles of red, green, and yellow. “I will be recording the color of every dot,” Amin said. “This will take me quite a long time.”

On Wednesday morning, October 16, 2002, Connell was up, as usual, at 5:30, replying to e-mails, reading journal articles, and reviewing notes for the day’s lectures. At 7 she went into Eloise’s room. “Good morning, sweety. Time to wake up,” Connell said, pecking her 9-year-old daughter’s cheek. Then it was dressing, frying eggs for breakfast, packing lunch for Eloise. Her husband, Mitchell Gayer, an environmental scientist, was off to work at 7:30. Before 8, Connell walked Eloise a mile to school and then jogged back home. No early meetings that morning, so she had a few minutes to practice her cello. At 8:30 she backed her car out of the driveway. Five minutes later she was on Route 78 west, and in another 20 minutes she pulled into the lot behind the medical school.