Probing the DNA of Death

NEW YORK —  The human remains of the day shelter in the shade of a white tent near the East River’s edge.

Here, set off by wilted bouquets, are the unanswered questions of the World Trade Center dead, stored in the 16 refrigerated trailer trucks parked on this shaded sliver of vacant pavement.

Each question is an anonymous flake of human bone or fleck of flesh picked from the rubble and preserved in a scientific limbo. There are 14,994 of them.

This is death divorced from identity--body parts without names to claim them; names without remains that can be mourned.

These human fragments are the dark heart of an unconventional murder mystery that will take years to solve. There is no question who committed the Sept. 11 attack or why terrorists may have acted as they did; little question now of the 2,797 names of those who died. The only answer authorities still seek--at a cost of $58 million since last fall--is how to settle those names on these unidentified remains.

Only half the dead have been identified so far.

The search has brought investigators to the edge of what science can discern of death.

By necessity, forensic specialists led by Dr. Robert Shaler, head of forensic biology at the New York chief medical examiner’s office, are virtually reinventing the science of identification.

Their pursuit of identity has turned the busiest morgue in the U.S. into a laboratory at the forefront of human gene research. They are forging the new tools of 21st century forensic medicine.

They are creating better ways of handling DNA, perfecting new genetic testing techniques and developing computer programs to analyze genetic variations. The new techniques one day may help diagnose inherited traits across the entire human population.

“We are moving into new territory,” Shaler said. “I feel as nervous as I did the first day of the attack.”

*

In its scale and scientific demands, the federally financed World Trade Center investigation is unique in the annals of crime and forensic medicine, experts say.

In an effort that rivals the Human Genome Project, Shaler has marshaled a national network that includes the New York State Police, the FBI, six biotechnology companies, a score of DNA consultants, computer software developers and an advisory committee of 30 forensic experts that has met every eight weeks to thrash out technical issues.

So crushed, burned, waterlogged and corroded are these remains that they defy conventional identification techniques, forensic specialists quickly discovered.

Consequently, almost half of the identifications made so far have been solely on the basis of genetic testing--682 of the 1,411 named--and DNA analysis helped in the identification of 343.

Of the rest, there is not enough undamaged DNA to build a normal genetic profile.

Even now, no one can even tell how many people these refrigerated tissues encompass. No one knows how many people vaporized in the fiery crashes and collapsing towers.

The fires took three months to extinguish. Crews sifted 1.6 million tons of debris for nine months to exhume the remains.

Death crushed some people so fiercely that only genetic analysis has been able to tell their intermingled cells apart.

Death made a jigsaw of others. More than 180 pieces of one victim have been identified so far. The slayers and the slain mingle, flesh and bone, in an autopsy test tube.

The forensic experts so far have matched 4,930 body pieces to 1,411 of the victims and given over many of them for burial or cremation.

“If this had occurred in 1980, or even 1990, the forensic work would have to stop now, with only half the people identified after heroic, intensive work,” said Clyde Snow, a forensic anthropologist who helped pioneer modern identification techniques.

By pushing the state of the forensic art, Shaler is hoping to identify remains of at least 600 more victims.

In recent weeks, the forensic experts have invented more refined testing techniques to extract usable DNA. They are retesting every one of the thousands of unclaimed pieces of bone, flesh, hair and clotted blood samples.

Conventional genetic testing--the mainstay of the effort so far--is reaching its limits. Shaler is resorting to experimental genetic techniques never before used on such a scale.

Since June, DNA experts at Celera Genomics in Rockville, Md., have processed 19,000 DNA specimens from victims and relatives, examining the genetic material contained in the thousands of mitochondria in every cell.

The DNA in these cellular power plants is inherited directly from each person’s mother. It is much smaller than DNA found in the nucleus of a cell but is much tougher and more likely to survive intact in damaged tissue.

This month, forensic experts at Orchid Cellmark Inc. in Germantown, Md., expect to begin tests that look for single variations in the 3 billion characters of the human genome. These differences--called single nucleotide polymorphisms--are part of the reason every individual is unique. So far, scientists have mapped 1.4 million of these molecular variations in humankind.

Forensic experts are contemplating more exotic methods that might allow them to rebuild shattered fragments of DNA the way a cosmetic surgeon can rebuild a face.

Even for those professionally hardened to death, the work is emotionally exhausting.

It is not uncommon to find forensic technicians weeping at their computer terminals. Several have asked for transfers. One lab expert quit under the strain earlier this year. A second broke down emotionally and was hospitalized.

There is no room for error. Perfection is the acceptable legal standard.

Even so, they must anticipate failure.

Morgue technicians already are drying these tissues, like petals of rare flowers, to preserve them for a day when more advanced technology may allow the identification of the most damaged remains

“If somebody wants to look at this 20 years from now, they will be able to scrutinize it,” Shaler said. “Somebody, someday will challenge what we have done.”

*

When the first hijacked airliner hit the World Trade Center, Shaler was holding his regular Tuesday morning staff meeting to plan the workload for the 90 forensic experts under his direction.

Shaler, 59, has unblinking blue eyes shielded by frameless glasses. A closely trimmed white beard softens his expression. He is light on his feet as he sidesteps the cardboard boxes of unfinished cases that crowd his office cubicle.

In more ways than one, Shaler is married to his work. His wife, whom he met at a medical examiners conference, is a forensic scientist for the New Jersey State Police.

After almost 14 years in the medical examiner’s office, he is no stranger to the confusion that fogs the circumstances of violent death.

The city’s Office of the Chief Medical Examiner, led by Dr. Charles Hirsch, is the country’s busiest, handling 25,000 deaths a year, including 3,300 homicides and sexual assaults. Hirsch recruited Shaler to found the forensic biology laboratory.

As conflicting reports filtered back to the medical examiner’s office on Sept. 11, a seven-member emergency morgue team mobilized and headed downtown, in accordance with the city’s long-standing plans for any air disaster.

It was no surprise when they left without asking anyone from the forensic DNA team to go with them. They were never included in disaster drills.

Within hours, the sobering scale of the World Trade Center disaster emerged. Early reports placed the dead at 6,700 or more. Morgue experts were braced to handle as many as a million body parts. The numbers of the missing and murdered continued to fluctuate for months.

In all, they would find 19,924 body parts.

For nine months, human remains recovered from the Trade Center site or culled from rubble trucked to the Fresh Kills landfill on Staten Island were delivered to the open-air bay by the side entrance of the medical examiner’s office. Even now, tissue still turns up. The most recent fragment was delivered from the World Trade Center area Monday.

There, under a canopy on a cul-de-sac blockaded by six metal barricades and under police guard, Shaler and his staff took possession of the dead.

As the only full-time forensic anthropologist on the medical examiner’s staff, Amy Zelson Mundorff was the first to examine remains when they came through the door, the last one to sign them out when they finally were identified.

Pathologists, dental experts and fingerprint analysts searched each new set of remains for identifying marks or features that might help distinguish it from the other dead--a distinctive tattoo, a pattern of freckles, a healed bone fracture, a birthmark, a wedding ring.

Each piece was cataloged, tagged with a bar code and measured.

“We would go through the bags and sort out the cases,” Mundorff said. “Anything that was not attached to something else got its own case number. We would take the time, if we could, to put the pieces back together, if we could make it into one case.”

She kept at it 12 hours a day, six days a week.

For many emergency workers, the difference between the quick and the dead at the World Trade Center was uncomfortably close.

On that Tuesday morning, Mundorff had rushed to the World Trade Center shortly after the first aircraft crashed into the building, as part of the disaster team dispatched to assess the scene.

Pressing forward with other emergency workers, she was caught in the tumbling outwash of debris from the fall of the south tower. Slammed against a wall, she suffered a concussion and broken ribs. Rubble and broken glass slashed her legs. One of her co-workers fractured his skull.

For the first month, she looked at sets of remains through two blackened eyes.

“We were too close,” she said. “When I look at the cases on the table, I feel lucky.”

*

As Mundorff and her colleagues sorted through bag after bag, it became clear that the mainstays of forensic identification--detailed skeletal analysis, dental X-rays and comparison with existing medical records--were all but useless.

There rarely was enough bone to make a recognizable skeleton; no faces that could be reconstructed; few teeth to be matched to dental records; even fewer whole fingers to print.

“So many of our traditional anthropological techniques are obsolete in a mass disaster like this,” Mundorff said. “It is frustrating. We have to take what we know and apply it in a different way.”

Of the 2,797 dead, only 287 whole bodies were found. Only 185 people could be identified solely from their teeth; 70 from their fingerprints alone.

By contrast, remains of the 44 people who died in the crash of United Airlines Flight 93 near Shanksville, Pa., were all quickly identified, aided by accurate passenger manifest information.

All but five of the 189 people who died at the Pentagon have been identified. Forensic experts were able to take advantage of military identification tags and extensive government medical files.

Remains of nine hijackers from those two crash sites also have been identified.

But the victims of the World Trade Center attack included office workers and visitors from 60 countries, shredded now beyond all recognition.

To identify the fragments, the medical examiner’s technicians would have to look deep into the heart of the human cell.

Properly treated, the nucleus of a cell can yield the long, twisted thread of DNA that forms the unique molecular signature for every individual. Stretched to its full length, that strand of DNA is almost 6 feet long. It weighs barely a billionth of a gram.

Only one-tenth of a single percent of DNA differs from one person to the next.

Scientists can use these variable regions to generate a DNA profile of an individual, using samples from blood, bone, hair and other body tissues. In conventional genetic testing, a series of chemical probes will bind to the DNA sample in a distinctive pattern for an individual.

That, in turn, can be compared to DNA from the victim’s personal effects or with that of relatives.

Robotic gene sequencers and supercomputer bioinformatics systems would have to do what family and loved ones could not.

“We take snippets of tissue--less than a cubic centimeter,” Shaler said. “It is not much, but there are tons of cells in there. You just need to find 100 good cells.”

It is technology whose time had come.

By last year, 120 DNA crime laboratories were involved in more than 16,000 cases and were analyzing more than 265,000 samples from convicted offenders. In New York alone, genetic testing is being used to reevaluate evidence in 17,000 unsolved rapes.

As a new tool of mass identification, genetic testing helped investigators identify all 230 of the victims of TWA Flight 800, which crashed in the ocean off Long Island in 1996. It helped resurrect the identities of the murdered from the mass graves of Bosnia and Guatemala. It reunited families with remains of long-missing soldiers killed in Korea and Vietnam.

In the process, genetic testing also altered public expectations about the anonymity of death. With so much scientific knowledge to be had from the code of life, a nameless death seemed an affront.

Even America’s Unknown Soldier was no longer unknown.

In 1998, the remains of an unidentified American serviceman from the Vietnam War, buried beneath the Tomb of the Unknowns in Arlington National Cemetery, was removed from his public tomb, identified through genetic testing and returned to his family.

The crypt has since remained vacant.

Just as genetic testing altered the forensic science of identification, however, the demands of the World Trade Center dead would in turn change the technology of genetic testing.

*

As a laboratory technique, modern genetics depends on highly automated machinery that can perform the mindless, repetitive chemical work of gene sequencing at high speed, running as many as 800,000 genotypes a day.

Scientists around the world dump more than 100 million bases each week into a public data repository. It takes arrays of supercomputers to make sense of the raw genetic sequences, by identifying the patterns of DNA code that indicate genes or the presence of identifying markers.

Although New York City has the largest forensic DNA lab in the country, the demands of the World Trade Center investigation instantly swamped its resources.

The medical examiner quickly drafted the New York State Police forensic DNA laboratory in Albany and two of the world’s largest genetic sequencing companies: Myriad Genetic Laboratories Inc. in Salt Lake City and Celera Genomics, which was the first private company to sequence the human genome. A third company, Applied Biosystems Inc. in Foster City, Calif., provided equipment and expertise.

Two of the most experienced private forensic DNA identity laboratories--Orchid Cellmark and the Bode Technology Group in Springfield, Va.--also joined the investigation.

Laboratory Corp. of America, which has 900 DNA centers around the country, and the New York State Police collected almost 7,000 razor blades, combs, toothbrushes, lipsticks and other items that had belonged to victims, which could be tested for DNA and compared to remains. From relatives, they also collected more than 6,800 DNA swab samples to be compared with victims’ genetic profiles.

Soon, couriers were crisscrossing the country carrying sealed pouches full of bar-coded vials of DNA.

But no one had ever tried to use these high-speed laboratory DNA extractors and gene sequencers to handle so many tissue samples so badly damaged.

Nor were research laboratories accustomed to the unforgiving demands of forensic identification or the legal niceties of handling evidence.

From the outset, it was more than the robots and the prepackaged biochemical kits could handle.

Shaler’s technicians processed the first 3,000 sets of remains by hand, extracting the DNA themselves.

The chemical extraction solutions had to be customized by trial and error, the timing adjusted and procedures revamped. As the weeks went on, every one of the processing methods was customized to handle these unusual human cells, said Brian Ward, president of operations at Myriad.

Scientists were retrained. Laboratories were rebuilt and secure evidence rooms constructed to hold the DNA. The reliability of new procedures was tested again and again, then certified by the New York Department of Health.

At Celera the new labs were ready by November, but it took until June to develop computer programs that could analyze all the data the high-speed sequencers spit out.

“There has never been a laboratory using high-speed robotics to process such a large number of mitochondrial samples--not the FBI, not the Defense Department, not anywhere,” said Rhonda Roby, forensic manager for Applied Biosystems.

Almost 20,000 DNA samples were sent to Myriad for analysis on its 12 high-speed sequencing robots.

More than 13,000 bone samples were shipped out for testing and analysis to the Bode laboratory. So were DNA extracts from 4,000 remains and 3,000 DNA samples gathered from family members or personal effects.

“The bones are the hardest to do,” said Tom Bode, general manager of Bode Technology Group. “Most of them have been subjected to 2,000 degrees Fahrenheit for weeks on end.”

The pressure of so many dead pushed the company’s workers and technology to new extremes.

When Alaska Airlines Flight 261 crashed two years ago off the Ventura County coast, it took the Bode laboratory three months to process DNA from 860 bones recovered from the ocean, then considered a record pace.

By comparison, the lab processed 2,000 bone fragments from the World Trade Center dead in one week last October. Working around the clock, they continued to process 1,000 new samples a week for the rest of the fall.

As the weeks turned into months, however, only half the remains yielded useful DNA. Almost one-fourth showed no DNA at all.

Shaler soon realized that technicians would have to test all the unclaimed remains again, in the hope that they had learned enough from the year to coax a name from even these damaged cells.

*

In the DNA identification room, Shaler squinted at the matrix of the dead scrolling across the computer monitor.

Each numbered cluster of items detailed a set of body pieces, personal effects and DNA from families of victims that all have a genetic sequence in common.

Shaler clicked on a case number with the computer mouse.

The microprocessors brooded briefly, then flagged one collection of remains with a yellow tag.

“This looks like a potential mix-up,” Shaler said.

Retesting revealed that two people--a man and a woman--had died in the same spot, their remains intermingled in the crush of tons of falling Trade Center rubble. The tissues were recovered by rescue workers and collected in a single 50-milliliter test tube. Muscle clung to bone.

“People died together,” Shaler said. “We have a lot of that.”

Down the hall, three industrial freezers were stocked with vials of World Trade Center DNA. Nearby, a walk-in chiller held a roomful of bones gleaned from the rubble. Red plastic containers holding personal effects of disaster victims were stacked six deep to the ceiling.

On Shaler’s computer display, it was all reduced to symbols: life code and computer code.

In this molecular search-and- rescue effort, they are aided by a sophisticated computer matching program developed during the course of the investigation by an Ann Arbor, Mich., company called Gene Codes Forensics.

The program can cross-match thousands of gene sequences at a keystroke.

“DNA is the sexy part, but the real underpinning of all this is computer software,” Shaler said. “Without this, it would be impossible.”

Shaler moved the computer cursor down the ranks of the dead.

He clicked on a case number.

The screen blossomed into a matrix of genetic sequences from 63 body parts, matched against the DNA extracted from the bristles of a toothbrush brought in by a victim’s family.

“They all match this one person,” Shaler said. “And we have the identification.”

Even so, the computer’s conclusion must be independently confirmed by a second set of genetic tests, then laboriously cross-checked against all the family records and personal effects on file, to make sure that no one puts the right name to the wrong body.

“Even if you were 99% perfect, you would have still misidentified 28 people,” said Mike Hennessey, project leader for the World Trade Center identification system.

The heart of the system is the program developed by Gene Code Forensics. With 96,000 lines of code, the software links every item in the investigation, information once scattered through 22 different databases and countless file cabinets at laboratories in five states. With it, Shaler and his forensic team can simultaneously sort in seconds information from three different types of DNA tests on almost 20,000 partial human remains and compare it to more than 3,000 DNA samples gleaned from cheek swabs of victims’ kin and genetic material taken from almost 8,000 personal effects.

Other computer programs allow morgue analysts to cross-check family relationships and calculate the statistical probabilities of a match between genetic samples and family groups.

In some cases, Shaler and his team have been able to use the computer programs to meld partial fragments of DNA from different but related samples, to create virtual genetic profiles for comparison purposes.

The company delivered the first version of the program, called the Mass Fatality Identification System, in December.

It made 55 conclusive identifications its first day. Now the team revises it every week to handle the changing demands of the investigation.

“This requires a level of quality assurance beyond anything we have ever done, because of the consequences of a mistake and the emotional pressure,” said Howard Cash, the president of Gene Code Forensics. “Nothing can ever be wrong.”

Every Friday morning, Cash flies from Michigan to New York, hand-carrying the latest update of the software.

When he walks through the lobby of the medical examiner’s office, he passes a child’s note taped to the wall, written in red crayon on white construction paper: “I hope you find the people.”

*

The dead transform the landscape they occupy, even as a temporary measure.

In this way, a cornfield becomes a battlefield monument. A stretch of invasion beach becomes a place in the heart.

So too this sloping asphalt slab where the homeless dead are parked has--without planning or public ceremony--become a memorial.

There is no plaque.

There is the smell of the dead, the steady hum of cooling fans and the roar of the rush-hour traffic on FDR Drive beyond the barbed-wire fence.

By the morgue tent, three unpainted plywood walls have been erected to give visiting families an easel to write their thoughts of love and remembrance. An architect is at work on drawings for a temporary mausoleum.

Along the walls, mourners taped up a photograph of the wife, the husband, the father, the mother, the son, the daughter, the friend whose remains have yet to be identified and released.

Every Friday afternoon, Charles Flood, an Episcopal chaplain from Philadelphia, lights a single candle by the morgue tent and holds a prayer service. He reads aloud the 23rd Psalm, should any among the living be there to listen.

“This DNA process is all wrapped up in feelings of wanting to conquer death,” Flood said. “It is very American. It is a way of saying we will prevail: This dust shall have a name again.”

Some families cannot bring themselves to come. A few cannot stay away.

There is a woman who comes every day and leaves one fresh flower. A couple from New Jersey paid their first visit not so many evenings ago.

Their son is here. They know the trailer where his remains are stored. More of him may yet be identified. They agonized over how much longer to wait before holding his funeral.

They left his picture by the trailer.