Doubt — and a reportedly royal severed head — haunts a murky corner of forensic science these days, as researchers squabble over an unearthed packet of mummified remains thought to have belonged to King Henry IV of France. 
 
 The mystery has produced a frightful case of regret among two researchers who were part of the first team to investigate the purportedly royal noggin. This week, French pathologist Geoffroy Lorin de la Grandmaison and Leslie Eisenberg, an American forensic anthropologist, wrote to the British Medical Journal and urged the retraction of the 2010 study that first identified the disembodied head as belonging to Henry. 
 
 At the heart of the macabre drama is an embalmed head with several vertebrae still attached. The remains were found in 1919 in the Royal Basilica of St. Denis outside Paris and reportedly secreted away by a civil servant. Reappearing almost a century later, the specimen still had its soft tissue and organs intact, right down to the open mouth and partially closed eyes. 
 
 On the basis of CT imaging and digital facial reconstruction, French medical examiner and forensic osteo-archaeologist Philippe Charlier and a multidisciplinary team — including Eisenberg and Lorin de la Grandmaison — in 2010 identified the head as that of the charming and rakish monarch known variously as "the Green Gallant" and "Good King Henry." 
 
 Even a mushroom-like growth on his nose and evidence of a pierced right ear seemed to point to King Henry IV. Though beloved by most of his people, the Bourbon monarch was assassinated in 1610 after 21 years on the throne. 
 
 That was just the beginning of his misfortune. In 1793, marauding revolutionaries sacked Paris' churches and desecrated the graves of many a purported royal. Legend had it that Henry's remains got the same treatment that befell his descendant Louis XVI: Dead or not, it was off with his head. 
 
 But researchers from the University of Leuven in Belgium were not so sure the head belonged to Good King Henry. 
 
 Obtaining a sample of the mummified tissue, they conducted a genetic analysis and compared it with DNA samples given by three male descendants of the House of Bourbon. Analyzing the Y chromosome of the three descendants, as well as genes that reveal matrilineal descent, they found clear similarities among the three descendants. But none of those matched the DNA of the disembodied head. 
 
 The team further compared the Y chromosome taken from the mysterious head with that taken from a blood-soaked handkerchief said to contain the genetic material of Louis XVI. (That "sample" was collected on Jan. 21, 1793, when a spectator at La Place de la Revolution in Paris thrust out the cloth to capture blood gushing from Louis' headless body.) The two samples did not match. 
 
 Though widely used in genealogical research, paternity testing and forensics, the Y chromosome's pattern of repeated DNA letters is not as definitive a test of genetic relatedness as are the patterns found on other chromosomes. So perhaps this still was Henry's head? 
 
 "Impossible!" declared Jean-Jacques Cassiman, an emeritus professor at the University of Leuven and coauthor of the Belgian team's study, whose results were published in the European Journal of Human Genetics this month. 
 
 The findings came on top of an earlier dismissal by French historian Philippe Delorme. Among other things, Delorme noted that the head bore no sign of craniotomy, as would be typical for a member of the royal family at that time. In such an esteemed person, craniotomy incisions would have been made in life (as a treatment for infection, head trauma and other ills) and in death (as a means to harvest "rondels," diskettes of bony tissue that were subsequently worn around the neck as an amulet). 
 
 Charlier, the lead investigator of the 2010 study, has been stout in defense of his findings. Illegitimacy — not of his methods but among many princelings of the French court — may explain the genetic mismatch, Charlier has argued. Extramarital liaisons among members of the court make it "hopeless," he said, to ascertain lineage using DNA tests over so many generations. 
 
 Such protests have failed to discourage defections from Charlier's team of researchers. Hence this week's letter by two of Charlier's coauthors and two others who have challenged the 2010 findings, saying that "robust scientific arguments ... negate the conclusions." 
 
 "The retraction of the article is now justified, as a rigorous scientific anthropological study should have excluded the hypothesis (and the findings) that the head belonged to" Henry, the four wrote. 
 
 Nailing down the identity of long-lost bones is tricky, and disputes among forensic scientists are common, said Alison Galloway, a forensic anthropologist at UC Santa Cruz who has followed, but not been involved in, the royal kerfuffle. 
 
 "The original analysis looked good," Galloway said. But Charlier's team should have known that its conclusions would be challenged, she said. 
 
 "If you had a group of 100 skulls — all men of European ancestry, let's say — and you had a photograph of a person, about 10 of those skulls could superimpose pretty well on that photograph," she said. Although facial reconstruction makes for great visuals in a TV drama, "it's a technique much better used to exclude somebody" than to identify him definitively. 
 
 With grieving relatives, criminal defendants and judges looking on, forensic scientists typically stick with conclusions that will hold up in a court of law. Only rarely do their disputes become the object of royal speculation and historical ghost stories. 
 
 Charlier's investigation, and the ensuing fracas, may be more of a divertissement than a serious professional debate, Galloway said. "Why are we so worried about whether it's Henry's head?" 
 
 melissa.healy@latimes.com ||||| This photo dated Thursday Dec. 16, 2010 and provided by Galaxie Presse shows what is believed to be the head of former French King Henry IV pictured in the 1930s. A 400-year-old skull that French researchers believe was the head of celebrated King Henri IV was squirreled away for decades in the attic of a now-retired public servant, and now has been handed over to one of his Bourbon dynasty descendants. (AP Photo/Galaxie Presse) NO SALES Three years ago, French researchers declared that a centuries-old mummified head was that of the beloved King Henri IV. But now a new study says, "Non!" 
 
 The original conclusion was based largely on facial reconstruction techniques and signs the skull had injuries similar to those suffered by the monarch. The new study looked at DNA instead. 
 
 It found a genetic mismatch between the head and three living male relatives of the 17th-century French king. The researchers concluded the head didn't come from anybody in the royal lineage. 
 
 Henri IV was one of France's best-loved kings, credited with bringing religious peace to the country and building Parisian landmarks like the Pont Neuf bridge during his reign from 1589 to 1610. He was the first of the Bourbon monarchs and grandfather of the Sun King Louis XIV. 
 
 It's "impossible" that the head belongs to Henri IV, said Jean-Jacques Cassiman, an emeritus professor at the University of Leuven, one of the authors of the new study. The research was published online Wednesday in the European Journal of Human Genetics. 
 
 The same study also found a blood sample previously attributed to King Louis XVI, one of Henri's descendants, lacked any royal ties. Louis XVI died on the guillotine during the French Revolution and many spectators reportedly soaked their handkerchiefs in the king's blood. But Cassiman and colleagues found the lingering traces of powdered blood on the handkerchief they tested did not match the Bourbon family DNA. 
 
 Cassiman said he wasn't sure who the head or the blood sample belonged to. The DNA from both failed to match samples from other royal families on record. 
 
 This is a schematic representation of the paternal line of the three living Bourbons and the French kings Henry IV and Louis XVI. Credit: KU Leuven 
 
 Amid the frenzy of the French revolution, the royal tombs at Saint Denis near Paris were dug up and some accounts claim Henri IV's head got chopped off and stolen. Cassiman and his co-authors dispute that Henri was decapitated and believe the king is still buried in Saint Denis after the tombs were resealed in 1817. 
 
 "That is impossible to verify," said Michael Rowe, a senior lecturer in European history at King's College London, who was not connected to either study. He said there wasn't much evidence one way or the other to suggest what happened to Henri's head. But Rowe said that Henri IV's reputation as one of France's best kings might have saved his body from decapitation. "If the revolutionaries were going to spare any of the kings, it would have been Henri IV." 
 
 This is a schematic representation of the maternal line of the French kings Henry IV and Louis XVII. Credit: KU Leuven 
 
 Philippe Charlier, author of the 2010 study that identified the head as Henri IV, dismissed the new research. He said he and colleagues have found an exact match between a three-dimensional comparison of the ancient head and Henri IV's death mask. He said the work will be published soon in a forensic journal. 
 
 Charlier added there were numerous doubts about the paternity of various kings in the Bourbon line, which might explain why there was no link between the DNA of modern-day descendants and the ancient head. 
 
 "It is hopeless to try to match a family tree and a series of genetic links (over) such a long period," he said in an email. 
 
 Frank Ruehli, of the University of Zurich and the Swiss Mummy project, agreed there could be questions about paternity in the Bourbon line and also raised concerns about the authenticity of the ancient DNA samples, as he said it was very difficult to get good data from a sample several centuries old. 
 
 "The best would be to look at (Henri IV's) heart as well," he said. "If you have multiple tissues from the same body showing the same results, that would be a positive identification," he said. Ruehli said given the lack of complete data, it was impossible to know for sure whom the head belonged to. 
 
 Charlier said the mummified head is currently in a Parisian bank vault awaiting reburial but that there are no exact plans yet. 
 
 Explore further: Dried squash holds headless French king's blood, study finds 
 
 More information: www.nature.com/ejhg ||||| Image copyright University of Leicester 
 
 Analysis of DNA from Richard III has thrown up a surprise: evidence of infidelity in his family tree. 
 
 Scientists who studied genetic material from remains found in a Leicester car park say the finding might have profound historical implications. 
 
 Depending on where in the family tree it occurred, it could cast doubt on the Tudor claim to the English throne or, indeed, on Richard's. 
 
 The study is published in the journal Nature Communications. 
 
 But it remains unknown when the break, or breaks, in the family lineage occurred. 
 
 In 2012, scientists extracted genetic material from the remains discovered on the former site of Greyfriars Abbey, where Richard was interred after his death in the Battle of Bosworth in 1485. 
 
 'Overwhelming evidence' 
 
 Their analysis shows that DNA passed down on the maternal side matches that of living relatives, but genetic information passed down on the male side does not. 
 
 We may have solved one historical puzzle, but in so doing, we opened up a whole new one Prof Kevin Schurer, University of Leicester 
 
 However, there are a wealth of other details linking the body to the Plantagenet king. 
 
 "If you put all the data together, the evidence is overwhelming that these are the remains of Richard III," said Dr Turi King from Leicester University, who led the study. 
 
 Speaking at a news briefing at the Wellcome Trust in London, she said that the lack of a match on the male side was not unexpected; Dr King's previous research has shown there is a 1-2% rate of "false paternity" per generation. 
 
 The instance of female infidelity, or cuckolding, could have occurred anywhere in the numerous generations separating Richard III from the 5th Duke of Beaufort (1744-1803), whose living descendants provided samples of male-line DNA to be compared against that of Richard. 
 
 Image copyright University of Leicester Image caption Wendy Duldig and Michael Ibsen are 14th cousins, descended from Richard's eldest sister Anne of York 
 
 "We may have solved one historical puzzle, but in so doing, we opened up a whole new one," Prof Kevin Schurer, who was the genealogy specialist on the paper, told BBC News. 
 
 Investigation of the male genealogy focused on the Y chromosome, a package of DNA that is passed down from father to son, much like a surname. Most living male heirs of the 5th Duke of Beaufort were found to carry a relatively common Y chromosome type, which is different from the rare lineage found in the car park remains. 
 
 Richard III and his royal rival, Henry Tudor (later Henry VII), were both descendants of King Edward III. The infidelity could, in theory, have occurred either on the branch leading back from Henry to Edward or on the branch leading from Richard to Edward. 
 
 Henry's ancestor John of Gaunt was plagued by rumours of illegitimacy throughout his life, apparently prompted by the absence of Edward III at his birth. He was reportedly enraged by gossip suggesting he was the son of a Flemish butcher. 
 
 "Hypothetically speaking, if John of Gaunt wasn't Edward III's son, it would have meant that (his son) Henry IV had no legitimate claim to the throne, nor Henry V, nor Henry VI," said Prof Schurer. 
 
 Image copyright University of Leicester Image caption Turi King says there is a greater than 99% probability that the body is that of Richard 
 
 Asked whether a break in the branch of the tree leading to the Tudors could have implications for the legitimacy of the present-day royal family, Prof Schurer replied: "Royal succession isn't straightforward inheritance from fathers to sons, and/or daughters. History has taken a series of twists and turns." 
 
 The breakage was statistically more likely to have occurred in the part of the family tree which does not affect Royal succession - the most recent stretch - simply because more links in the chain exist there. 
 
 The lack of any match for the Y-chromosome lineage is quite curious and suggests an intriguing new avenue for dynastic DNA studies Prof Martin Richards, University of Huddersfield 
 
 And Dr Anna Whitelock, a reader in early modern history at Royal Holloway - University of London, told BBC News: "It's important to note that Henry VII claimed the throne "by right of conquest" not blood or marriage - his claim was extremely tenuous. 
 
 "Henry VII was descended from Edward III from the Beaufort line - the Beauforts were legitimised by half-brother Henry IV but not in succession. Royal succession has been based on many things in the past: ability to lead troops, religion, connections - not always seniority by royal blood." 
 
 She added: "The Queen's right to reign in based on the 1701 Act of Settlement that restricted succession to Protestant descendants of Sophia of Hanover. A medieval false paternity does not challenge the current Queen's right to reign." 
 
 Blue-eyed and blond 
 
 Richard's maternal-line - or mitochondrial - DNA was matched to two living relatives of his eldest sister Anne of York. Michael Ibsen and Wendy Duldig are 14th cousins and both carry the same extremely rare genetic lineage as the body in the car park. 
 
 Richard III was defeated in battle by Henry Tudor, marking the end of the Plantagenet dynasty and the beginning of Tudor rule, which lasted until Queen Elizabeth I died childless in 1603. 
 
 Richard's battered body was subsequently buried in Greyfriars. As the Leicester team uncovered the male skeleton, the curvature in its spine became obvious. The condition would have caused one of the man's shoulders to be higher than the other, just as a contemporary of Richard described. 
 
 Genes involved in hair and eye colour were also tested. The results suggest Richard III had blue eyes, matching one of the earliest known paintings of the king. However, the hair colour analysis gave a 77% probability that the individual was blond, which does not match the depiction. 
 
 But the researchers say the test is most closely correlated with childhood hair, and in some blond children, hair darkens during adolescence. 
 
 Image copyright University of Leicester Image caption The curvature in the spine of "Skeleton 1", later confirmed as Richard, was obvious during its excavation 
 
 The researchers took all the information linking the body to Richard III and carried out a statistical test known as Bayesian analysis to determine the probability that the body was indeed his - or not. Despite the absence of a male-line genetic match, the results came back with a 99.999% probability that the body was that of the Plantagenet king. 
 
 Commenting on the study, Prof Martin Richards, a population geneticist at the University of Huddersfield, told BBC News: "The work seems to have been done with great care and looks very convincing to me." 
 
 He said Richard III's maternal DNA type was very rare, and carried an additional genetic variant not previously seen before that "seems to be unique amongst a database that includes several thousand Europeans". 
 
 "So I agree that their assessment of the match probability is very conservative and it's very likely to be him," Prof Richards said. 
 
 He added that, given the apparent certainty of the body's identity, "the lack of any match for the Y-chromosome lineage is quite curious and suggests an intriguing new avenue for dynastic DNA studies". 
 
 Dr Ross Barnett, a specialist in ancient DNA at the University of Copenhagen, agreed that the work was "interesting and thorough". 
 
 Dr Barnett had previously raised questions over a preliminary analysis of the maternal-line DNA. But he told BBC News: "Now the paper is here and available for scrutiny, I have no further complaints. The team are excellent and I would expect the analysis to be robust." 
 
 Follow Paul on Twitter.
– Forensic researchers have a royal dispute on their hands: They can't agree on whether a mummified head belongs to France's Henry IV, explains the Los Angeles Times. The question seemed settled in 2010 when a team of researchers used facial-reconstruction techniques to conclude that it was indeed "Good King Henry," who was assassinated in 1610. But now a second team of scientists says it isn't so because DNA tests don't match Henry's living relatives. They wrote this week to the British Medical Journal urging a retraction of the earlier study, and, in fact, two members of the 2010 team agree that their conclusions were faulty. Not so, says the lead researcher from 2010. He explained to phys.org in an earlier story that it's useless to make conclusions based on DNA results in part because the French royals were such philanderers. "It is hopeless to try to match a family tree and a series of genetic links (over) such a long period," he said. Until things get settled, the mummified head is resting in a bank vault in Paris. (In other mummified news, click to read about a big find in Peru's capital.)