Before this happened, however, Rostropovich told Ivanov that he was about to set out on a visit to Japan. Ivanov, still in England, remembered that in 1892 Nicholas II as tsarevich had visited Japan. In Otsu, the heir to the Russian throne suddenly had been attacked by a sword-wielding Japanese. The blow, aimed at his head, glanced off his forehead, bringing a gush of blood but failing to bite deep. The wound was bound with a handkerchief. For one hundred years, a museum in Otsu had kept the blood-soaked handkerchief in a small box. For DNA comparison purposes, nothing could provide more accurate positive identity than achieving a match between bone material of unknown origin and blood from a known person. Ivanov was eager to go to Japan, but, as always, “there was no money. The English said, Why should we pay for this? The Russians said, We have no money.” Eventually, Rostropovich arranged for Ivanov’s trip. “It was the money we were going to use to dig up George,” Ivanov said. “So, instead of George, we did Japan.”
The Japanese were not anxious to give up or even to disturb the handkerchief, but Rostropovich spoke to his friend the emperor of Japan, and the emperor spoke to the relevant authorities. When Ivanov arrived he was permitted to remove and take with him a strip of the handkerchief three inches long and one eighth of an inch wide. Unfortunately, back in Gill’s laboratory in England, Ivanov ran into difficulties. “The handkerchief had been handled by too many people,” he said. “Cells slough off from fingers. There was a lot of blood on the handkerchief, but who knows how much of it was Nicholas’s? And there was a lot of dust and dirt. It would be impossible to say that any result you got from that handkerchief was reliable. There were too many other possible contaminants.”
Having failed with both George and Japan, Ivanov came up with a third possible source of DNA for comparison to the piece of the presumed tsar’s femur at Aldermaston. In 1916, Nicholas II’s younger sister Grand Duchess Olga married Colonel Nicholas Kulikovsky, a commoner. With Kulikovsky, Olga had two sons, Tikhon, born in 1917, and Guri, born in 1919. In 1948, Olga and her family moved to Canada, where Kulikovsky bought a farm and raised cattle and pigs. Guri Kulikovsky died, but in 1992, when Gill and Ivanov began their work together, Tikhon, at seventy-five, was living in retirement in Toronto. He was, by that time, Tsar Nicholas II’s only living nephew and, as such, the best available source for comparative mitochondrial DNA. If the femur from Body No. 4 had belonged to Nicholas II, it should match perfectly with DNA from Tikhon Kulikovsky.
Mr. Kulikovsky, however, refused to cooperate. When Ivanov wrote to him, explaining the purposes of the investigation and asking for a blood sample, he received no reply. Ivanov tried again through Bishop Basil Rodzianko of the Orthodox Church in America, and, finally, through Metropolitan Vitaly, the head of the Russian Orthodox Church Abroad. Ultimately, Kulikovsky replied to Ivanov. “He told me he believed this whole bones business was a hoax,” Ivanov recalled. “He said, ‘How can you, a Russian man, be working in England, which was so cruel to the tsar and to the Russian monarchy?’ He said, ‘For political reasons, I will never give you a sample of my blood or hair or anything.’ ” Ivanov was disappointed, but he did not give up. “At that time, it was critical,” Ivanov said. “He was the closest relative. I spent a lot of my own money talking with him and his wife by telephone, assuring them that I was not a KGB agent. And they replied, ‘Then probably the only reason for your investigation is to prove that Tikhon Nicholaevich is not of royal blood.’ ” Ivanov gave up. “Okay, so we forgot about this Tikhon,” he said. “And after we published our work, some people wrote that our analysis was not accurate because we didn’t use the blood of Tikhon Kulikovsky. The fact is that his blood is no longer necessary. We found two other relatives. They gave us their blood, and we had everything we needed for our research.”
To locate the other two relatives, the Aldermaston genealogists looked again at the family tree. Because the chain of similar mitochondrial DNA is repeated indefinitely down through generations of females, they focused on the women closest by blood to Tsar Nicholas II. Beginning with his mother, Dowager Empress Marie, they found an unbroken line of five generations of mothers and daughters leading to a contemporary descendant willing to help. The tsar’s sister Grand Duchess Xenia had one daughter, Princess Irina. This Irina married Prince Felix Yussoupov, famous for having murdered Rasputin. Irina and Felix produced one child, a daughter, also named Irina. This second Irina married Count Nicholas Sheremetyev, with whom she had one child, a daughter, Xenia. Upon her marriage, young Countess Xenia Sheremetyeva became Xenia Sfiris. Now in her early fifties, Mrs. Sfiris lives in Athens and Paris, and it was in Athens that she received the FSS’s appeal for help. An exuberant, warmhearted woman, she agreed immediately. Following instructions, she pricked her finger, let some blood run into a paper handkerchief, where it dried, put the handkerchief in an envelope, and took it to the British Embassy. From there, it went, via diplomatic pouch, to Aldermaston.
The other donor of DNA material given to identify Nicholas II was found on what must seem an infinitely remote branch of the massive European royal family tree. Nevertheless, although the line stretched back over six generations, the connection was as reliable and productive as it was in the case of Mrs. Sfiris. James George Alexander Bannerman Carnegie, third Duke of Fife, Earl Macduff, and Lord Carnegie, is a sixty-six-year-old Scottish nobleman and farmer who descends from a common female ancestor of Tsar Nicholas II. She was Louise of Hesse-Cassel, a German princess who married King Christian IX of Denmark. One of her daughters became Empress Marie Feodorovna of Russia, the mother of Nicholas II. Another, older daughter, Alexandra, married the Prince of Wales, later King Edward VII. Queen Alexandra’s daughter Louise married the first Duke, of Fife. In 1929, Louise’s daughter Maud produced James, who, in 1959, succeeded to the title. The duke was willing to donate blood but, not wishing to incur publicity, made it a condition that he remain anonymous. Inevitably, in the course of time and with an investigation of this significance, knowledge leaked out.
As Gill and Ivanov expected, Xenia Sfiris’s mitochondrial DNA matched perfectly with that of the Duke of Fife. But when the matching 782 base pair letters lengths of the Greek woman and the Scottish peer were compared to the same section in the mitochondrial DNA extracted from the presumed tsar, there was a mismatch. A single letter was different. At a position numbered 16169, Xenia Sfiris and the Duke of Fife had a T; in this position Nicholas had a C. The other 781 pairs were in identical sequence. To check their data, Gill and Ivanov did a second mitochondrial DNA extraction from the bone believed to be the tsar’s. They cloned the DNA in this region after amplifying it with PCR and then transformed the product into E. coli bacteria. When fresh sequencing of these new clones was performed, seven of the clones were found to have a T at position 16169, thus matching Mrs. Sfiris and the Duke of Fife. But twenty-eight clones still presented the single spelling mistake, a mismatching C. The Aldermaston scientists concluded that Tsar Nicholas II had possessed two forms of mitochondrial DNA, one of which matched his relatives exactly; the other, at a single point, did not. This rare condition is known as heteroplasmy.
This single mismatched base pair letter caused great anxiety at Aldermaston. In their paper, the two scientists presented their interpretation of what they had found: “We consider … that the mitochondrial DNA extracted from the tsar was genetically heteroplasmic. This complicates the interpretation because the strength of the evidence depends upon whether we accept a priori that a mutation has occurred in the tsar. The probability of a single mutation was calculated to be approximately one in three hundred per generation, but this estimate does not take account of the incidence of heteroplasmy (much of which may be undetected).”
Gill understood that this one-letter mismatch raised questions about the validity of his findings. He believed that a mutation did occur, although he admitted that the odds against a mutation in any given generation were long. “A mutation is thought to occur [in a family] about once in three hundred generations,” he
said. But he insisted that he is talking primarily about a heteroplasmy, which he found, not a mutation, which he could not prove but which is the probable cause of that heteroplasmy. “Heteroplasmy is different from a mutation in nuclear DNA; it means that there are two types of mitochondrial DNA in the same person. What we did was demonstrate that there are two types of mitochondrial DNA in the tsar. One of these types differed by just one base; the other was identical to the relatives.’ That’s pretty good evidence that there has been a real mutation. Bear in mind that we’re working right at the frontiers of knowledge. The actual incidence of this type of phenomenon is not really known, and we suspect that it is much more common than originally had been anticipated.”
In July 1993, after ten months of work, Gill and Ivanov were ready to announce their results to the world. The Forensic Science Service convened a press conference and, on July 10, a large hall at the bleakly modern Home Office building in Queen Anne’s Gate was filled with reporters, photographers, and television cameramen. Dr. Janet Thompson, the director general of the FSS, presided. Aware that questions might be asked as to who had borne the cost of this research, she began by expressing hope that “the FSS will soon be able to put the techniques used, once validated, into practice in criminal casework to the benefit of the criminal justice system as a whole.”
Gill explained what he and his colleagues had done. He described how the sex of the specimens had been determined, how the family relationship between five skeletons had been established, how Prince Philip’s blood had made certain the identity of Alexandra Feodorovna and her daughters, and how the heteroplasmy found in the tsar’s DNA had complicated the effort to make an absolute statement about Nicholas. Nevertheless, the Aldermaston team announced that, given the DNA evidence and adding it to the anthropological and historical evidence provided by others, they were 98.5 percent certain that these were the Romanovs. This percentage, Gill said, was based on the most conservative interpretation of the DNA evidence. A more generous interpretation would increase the probability to 99 percent. Pavel Ivanov took a broader view of what had been done. “We are very close to the last part of this mystery, to one of the great mysteries of the twentieth century, one of the great mysteries of my country, of Russia,” he said.
The press conference produced headlines: RIDDLE OF ROMANOV REMAINS IS SOLVED (Financial Times), DNA TESTS IDENTIFY TSAR’S SKELETON (The Times), TSAR NICHOLAS’S BONES IDENTIFIED (The Washington Post). Tass told Russian readers that “British scientists” were “almost without any doubt” that the remains found in Siberia were those of Tsar Nicholas II and his family. Seven months later, in February 1994, Peter Gill, Pavel Ivanov, and others put their findings in print in their own words, publishing a description of their work in Nature Genetics, the authoritative journal of their profession. Their findings and article have never been challenged or even mildly criticized, in print or orally, by another DNA scientist.
* McCrery, a florid, enthusiastic man who was a policeman before he went to Cambridge to study Russian history, takes considerable credit for engineering the British-Russian effort. Hearing about the discovery of the bones, he says, he rang up Avdonin in Ekaterinburg. Avdonin put him on to Pavel Ivanov. Ivanov told him that the best place in the world for DNA testing was Aldermaston and gave him the name of Peter Gill. McCrery telephoned Gill, who, he says, was “quite excited, but not sure the Home Office would approve. Well, Kenneth Clark, [then] the home secretary, lives around the corner from me and I’ve known him for years. He is my M.P. So I contacted him and pointed out how prestigious it would be for the Forensic Science Service to be involved. ‘Will you give permission?’ And Clark said, ‘It’s a wonderful idea.’ So then I rang Ivanov back and Ivanov said, ‘How do I get there? I don’t have any money.’ So I said, ‘I’ll pay for it.’ Actually, somebody in Russia paid for Ivanov’s trip, but I got in touch with Applied Biosystems, which makes gene scanners and other machines they use in DNA work, and asked them whether they would pay Ivanov’s expenses in England. They said yes, and they came up with three to five thousand pounds for him to live on for ten months. So he came and brought the bones.”
* In fact, the security men of both monarchs were so concerned about the possibility of terrorism that King Edward never actually set foot on the soil of the Russian Empire. All meetings were held on the two yachts.
CHAPTER 9
DR. MAPLES VERSUS DR. GILL
William Maples, after his examination of the bones and his presentation of his findings at the Ekaterinburg conference in July 1992, was unwilling to let go of the Romanov inquiry. In his talk at the conference, he recommended that there be further archaeological exploration of the burial site and more extensive photo documentation and DNA testing of the remains. Apparently, he intended to do—or at least to supervise—most of this himself. In April 1993, Maples, Dr. William Hamilton, and Mrs. Maples returned to Siberia, assisted by two airline tickets provided by the television program Unsolved Mysteries. In Ekaterinburg, Maples rephotographed the skeletal remains more carefully than he had been able to do on his previous trip. He also removed one tooth from each of the skulls, except that of Dr. Botkin, which had few teeth to spare, and that of Kharitonov, where only the top of the skull was available. From Botkin and Kharitonov, he took leg bone fragments. The teeth, he believed, would be far more suitable for accurately identifying the Imperial family by DNA testing than the pieces of femurs taken to Britain by Pavel Ivanov. Along with these teeth, Maples carried away from Ekaterinburg a decree from the Sverdlovsk regional prosecutor authorizing him to export the bones, oversee DNA tests, and report the results back to the Sverdlovsk authorities. Curiously, no one bothered to tell Dr. Vladislav Plaksin, the chief medical examiner of the Russian government, or Pavel Ivanov, then in his seventh month working with Peter Gill at Aldermaston.
Returning to Florida, Maples held the Russian teeth in his laboratory for six weeks, then “transferred custody” to Lowell Levine, who carried them to California and, in June 1993, gave them to Dr. Mary-Claire King, who held two professorships at the University of California, Berkeley, one in epidemiology in the School of Public Health, the other in genetics in the Department of Molecular and Cell Biology. According to Maples, Dr. King “is the foremost forensic genetics scientist in the United States and one of the most highly regarded scientists in this field anywhere in the world.” She drafted the report prepared for the National Academy of Sciences on the use of DNA for forensic identification purposes. She has worked with a United Nations team in Argentina to identify kidnapped children and reunite them with their families. She has assisted the United Nations in El Salvador to try to identify the remains of victims of a mass murder in the village of El Mozote. Doctors Maples, Levine, and Baden knew her because she had worked with them on the remains of American servicemen brought back from Vietnam. By 1993, Maples said, she had more experience with mitochondrial DNA than the British Forensic Science Service. And she had a far larger database. According to Maples, Dr. King’s database held mitochondrial DNA information on one thousand people; Peter Gill and Aldermaston had only about three hundred. “In this field,” said Dr. Maples, “there simply isn’t anyone to compare with Dr. King.” Michael Baden and Lowell Levine agreed.
Maples and his colleagues had a secondary regard for Peter Gill, and, until they met him at the Ekaterinburg conference in 1992, they had never heard of Pavel Ivanov. Unable to understand Russian, they were not sure what Ivanov had told the meeting about his arrangements to test the bones in England. Nevertheless, Ivanov was friendly and tried to be helpful. Their return to Moscow that summer was disagreeable for the American team: a dog ran up and down the aisle of the Aeroflot plane; at the domestic airport in Moscow, people shoved and shouted at them. Dr. Ivanov, whose English is fluent, appeared and confidently steered the Americans to safety. The following day, wearing his FBI Academy T-shirt, he showed them through Red Square. He explained what he was doing, that he had been in touch with Gill and was making ar
rangements to do DNA tests in England. The Americans attempted to change his plans. “We offered him the chance to come and work in an American laboratory,” said Baden, “but he went to England because he could get there quicker and they would pay his way.” “The best thing for Ivanov,” Levine said, “was that he personally was going to take the bones to England and he was going to get to stay there.”
William Maples met Peter Gill for the first time and Pavel Ivanov again in July 1993, soon after Gill’s press conference announcing identification of the Romanov bones. Maples was in England, returning to America from his third visit to Ekaterinburg, where he had been filmed by Nova examining and describing the remains. From London, Maples and his wife drove to Aldermaston, where they took Peter Gill and Pavel Ivanov to lunch. The luncheon conversation was polite, this ambience being achieved by both sides ignoring feelings of mutual grievance. Maples was annoyed that Gill had just announced that he was 98.5 percent sure that the bones he had tested belonged to the Romanovs; this event had occurred just as Dr. Maples was arriving in Russia to be filmed by Nova. Ivanov was indignant that Maples, with the permission of the Sverdlovsk authorities, had initiated a second round of DNA tests at Dr. King’s laboratory in California without his being informed and while his own tests with Gill were still under way. There was no discussion at lunch about the heteroplasmy which Gill had discovered in Tsar Nicholas II, or the possibility that it had been caused by a mutation. The scientists did talk briefly about the Aldermaston finding that the three young females had the same mitochondrial DNA as one of the older females and therefore undoubtedly were mother and daughters.