Monday, December 15, 2014

A reconsideration of the DNA forensics in the Duke lacrosse case


Introduction
In the spring of 2006 three players on the Duke lacrosse team were indicted for rape and kidnapping.  The accuser claimed to have been raped by three men at least one of whom ejaculated inside of her body.  Some DNA results emerged in the spring of 2006. Eventually additional evidence was uncovered, and the overwhelming perception that this evidence was exculpatory brought about a major turning point in public opinion prior to the declaration of innocence.  After evidence of his misconduct emerged, the district attorney withdrew from the case, which then fell to the office of the Attorney General of North Carolina to investigate.  After the investigation was complete, the Attorney General declared the three players to be innocent in the spring of 2007, and the criminal matter ended.

The Duke lacrosse case is generally perceived as a highly visible case of prosecutorial misconduct and a rush to judgment by the press and the general public.  However with the 2014 publication of William Cohan’s book The Price of Silence, this view has now been challenged, and Mr. Cohan’s view of the DNA evidence is at odds with many who have commented on the case.  Therefore, a reevaluation of the DNA evidence is warranted on the basis of a general understanding of this case, as well as other cases in which the DNA evidence plays a central role, such as the Knox/Sollecito case, a frequent subject of this blog.

The initial results from the rape kit:  Is the absence of evidence evidence of absence?

The initial results from the NC SBI laboratory did not find semen, blood, or saliva.  Subsequently Y-chromosomal (Y-STR) tests were performed at a private laboratory.  Investigators found no DNA from any Duke lacrosse player in the rape kit items but did find DNA from Mangum’s boyfriend.  DNA from plastic fingernails was also extracted.  Some of the results from the private lab were released in a brief report issued on May 10, 2006, but other results were withheld.  Apart from the plastic fingernails (discussed below), how significant was the lack of DNA?

One blogger posed questions along these lines to several highly qualified experts, including Dan Krane, Elizabeth Johnson, and William Thompson.  Dan Krane responded: “It is worth noting that DNA tests are amazingly sensitive (DNA profiles can be generated from as little material as that left behind in a fingerprint) and Y-STR tests have the potential of determining a male’s DNA profile even when a female’s DNA is present in hundreds or thousands of times greater quantities. Scientists are always wary of asserting that the absence of evidence is not proof of absence but it certainly is reasonable to expect to find a rapist’s DNA associated with a victim when the victim presents herself to investigators within hours of an attack and when she has not bathed, the rapist did not use a condom and ejaculation occurred.”
  These experts gave thoughtful, nuanced replies. The consensus view seemed to be that the degree to which the absence of evidence was evidence of absence lies in the particulars of each case.

What then are the particulars of the Duke lacrosse case?  A site at Reed College lists the following guidelines for preserving evidence of a possible sexual assault:

 “Do not shower or douche
Try not to urinate. Urinating may reduce the ability to detect “date rape” drugs
 If there was oral contact, do not smoke, eat, or brush teeth
 Do not change clothes. If you have already changed your clothes, place them in a paper bag (plastic may destroy evidence) If you haven’t changed, keep the original clothes on and bring an extra set to wear home from the hospital
Go to a hospital with the capability of providing a SAFE exam and request the exam. The cost of a SAFE examination is paid for from a state fund.”

The alleged victim in the Duke lacrosse case was taken into custody shortly after the incident and did not have the opportunity to shower or to change clothes.  Therefore, the chances of preserving evidence would have been high, if an actual assault had occurred.  William Anderson wrote, “Furthermore, no one — no one — who inspected the room (and police gave that bathroom a thorough inspection, taking swabs everywhere and testing for DNA) found any evidence of bleach products having been recently used. They found no DNA of Mangum, and none of [Collin] Finnerty and [Reade] Seligmann, and nothing else that demonstrated that either of those two young men had ever been in that bathroom.”  Parenthetically, Taylor and Johnson reported that no fingerprints belonging to Mr. Finnerty were found in the bathroom (Until Proven Innocent, p. 183).

Unfortunately, the question of whether or not the accuser said that the victims used condoms has been not entirely without controversy.  Stuart Taylor and KC Johnson wrote (Until Proven Innocent, p. 327), “In her report from the night of the alleged attack, [SANE nurse Tara] Levicy had written without qualification that Mangum had said repeatedly…the rapists had used no condoms and that she was sure that at least one had ejaculated—in her mouth.  But on January 10 [2007, nine months later], according to [Linwood] Wilson, Levicy stated that Mangum “said ‘no’ but wasn’t really sure” whether her attackers had used condoms.”  Even if one finds Ms. Levicy to be credible about Ms. Mangum’s lack of certainty (which I do not), DNA may be transferred by simple physical contact, as is borne out in studies of simulated strangulation.  In addition, perspiration contains DNA, and sebaceous fluid is a probable source of touch DNA, according to a study by Vecchiotti, Filippini and their coworkers.  Thus even when a condom is used during a sexual assault, there may be transfer of DNA.

Asked whether or not the use of a condom could have been detected, Elizabeth Johnson responded, “Testing for these substances is not typically done, despite what you see on CSI. There has been some research done re spermicides on condoms, but none of this is done as part of a typical test and validated methods for casework aren’t there yet.”  Of course, even if condom use were detected, it would not differentiate between consensual and nonconsensual sexual activity.

The lack of body fluids is absence of evidence in its own right.  It also weakens the DNA evidence, meaning that a given DNA profile found in the absence of a particular fluid might have arisen from any biological tissue or fluid.  Peter Gill describes framework or hierarchy of levels (Misleading DNA Evidence, p. 19) at which DNA evidence can be evaluated:
1.     The sub-source level refers to the strength of evidence of the DNA profile itself.
2.     The source level is an evaluation of the strength of the DNA profile if it can be associated with a particular body fluid, such as semen or blood
3.     The activity level associates the DNA with the crime itself, e.g. sexual assault.
4.     The highest level deals with the ultimate issue of guilt/innocence.

Professor Gill’s book gives examples of cases in which a forensic scientist attempted to claim a higher level for the evidence than was warranted, based upon principles of forensics and probability.  The DNA of the rape kit is sub-source DNA, in this categorization.

The lack of DNA from the accused coupled with the lack of body fluids and the lack of other evidence in the bathroom is strongly exculpatory.  Whether or not it constitutes “proof of innocence” cannot be determined without defining the term.  However, I would weight this lack of evidence more heavily that eyewitness testimony, even if the latter came from a credible witness, owing to the well-known problems with eyewitness testimony.  Ms. Mangum’s credibility is questionable on a number of grounds.

The plastic fingernails in the trashcan
Ms. Mangum’s painted fingernails were placed in the trashcan of the bathroom and the players pointed them out to the police investigators days later.  David Evans could not be excluded as a donor.  The summary of conclusions from the office of the Attorney General of North Carolina stated on p. 12, "First, statistically, the chance of randomly selecting an individual from the population that could be included in this sample would be approximately 1 in 1000." With respect to the autosomal (standard DNA) profile, David Evans (along with about 2% of the population) cannot be excluded as a donor.  Likewise in the Y-chromosomal (YSTR) testing, David Evans cannot be excluded as a donor.  Although the summary report did not explain the calculations, the value of 1 in 1000 probably derives from combining the data from the two types of DNA testing.

The author of a 2014 book on the case (The Price of Silence), William Cohan clearly thought that the DNA was highly incriminating, discussing it on pp. 277-278 and pp. 325-326, among other places.   In the final chapter, Denouement, he returned to the subject on p. 602:  “What remains unresolved is, if in fact it was David Evans’s DNA on Mangum’s red plastic fingernails, how did it get there?”

DNA evidence beneath real fingernails typically is probative, not necessarily conclusive, evidence against a defendant.  Studies have shown that mixtures of DNA under fingernails are seen about 5% of the time (Gill, pp. 43-46).  However, there is less to the DNA from the plastic fingernails than meets the eye.  Taylor and Johnson wrote (Until Proven Innocent, p. 221), “And it would later become clear…that the DNA that might (or might not) have come from Evans had not necessarily been lodged “under” a plastic fingernail.  Rather, the SBI had extracted and mixed together all of the DNA taken from all surfaces of the three used plastic fingernails found in the wastebasket in Evans’ bathroom.”  Moreover, a case in Britain involving a taxi driver, David Butler suggests some additional caveats.  His DNA was apparently found on the fingernails of a murdered woman Anne Marie Foy.  He was convicted partially on the basis of the DNA evidence but later released.  A plausible theory is that he handled money which later carried his DNA to the victim.  There are not many forensic details that have been made public, but a few things can be gleaned from the information that is available.  One is that Mr. Butler's skin was unusually flaky.  Two is Hannah Barnes' report that "The victim was also wearing a glitter nail polish, which proved particularly attractive to dirt - and DNA." 

The defense correctly noted that David Evans was not uniquely identified in either of the tests.  The defense’s position was that if David Evans were a donor of the DNA, then there existed a plausible route for innocent DNA deposition, namely secondary transfer from materials in trashcan, such as dental floss and tissue paper.  Defense attorney Joseph Cheshire, "Any expert and any person in the world will tell you that your [own] DNA is in your bathroom." (Cohan, p. 242).  Saliva is a good source of DNA, for example.  The office of the attorney general of North Carolina accepted the possibility of secondary transfer as a reasonable explanation.  In the summary document pertaining to this case, the authors wrote: “Third, to the extent that Evans’s DNA could not be excluded, the SBI experts confirmed that the DNA could easily have been transferred to the fingernails from other materials in the trash can.”

It is uncontroversial that the presence of DNA does not indicate the time or manner of its deposition.  This cuts both ways: one cannot be certain that the DNA on the plastic fingernails arrived by secondary transfer, but neither can one rule it out.  Secondary transfer in DNA forensics is well established, and tertiary transfer has been observed under some circumstances.  Neither the amount of DNA nor the completeness of the DNA profile is a reliable guide as to whether the DNA arrived by primary transfer or secondary transfer.  Joe Minor and Suzanna Ryan have written several well-researched and readable articles that cover the subjects of DNA found on touched objects and secondary transfer.  Review articles by Van Oorshot and colleagues and by Meakin and Jamieson in academic forensic journals may be consulted for more detailed information.

Shedding and transferring DNA is perhaps more common than the general public appreciates.  Leslie Prey wrote, "We all shed DNA, leaving traces of our identity practically everywhere we go. Forensic scientists use DNA left behind on cigarette butts, phones, handles, keyboards, cups, and numerous other objects, not to mention the genetic content found in drops of bodily fluid, like blood and semen (Van Oorschot & Jones, 1997)."  The issues of shedding and transferring DNA were perhaps not very important when DNA profiling was in its infancy.  However, the introduction of techniques based on the polymerase chain reaction (PCR) meant that much smaller amounts of DNA are needed now than in the first generation of DNA profiling.  This is because the DNA is increased in amount (amplified) many times over prior to the analysis.  The more recent development of low copy number DNA profiling means that still smaller amounts of DNA can be typed, less than a dozen cells.

Although secondary transfer from the waste in the trashcan is highly plausible, primary or secondary transfer via the players themselves is also possible.
  Joe Minor wrote, “On one occasion, I swabbed my own hand after handshakes at a social function to determine the presence of other epithelial cells.  The laboratory was able to obtain a mixture of my DNA as well as two other individuals.”  Suzanna Ryan wrote, “One study performed by Lowe, et al., was designed to highlight a ‘worst case’ scenario and involved two individuals. The first was determined to be a poor shedder and the second a good shedder. These two shook hands for one minute. The poor shedder had washed their hands immediately prior to the experiment whereas the good shedder had not. After shaking hands the poor shedder held a sterile plastic tube for 10 seconds. The tube was then swabbed and tested for the presence of DNA. This experiment was performed on two sets of good shedder/poor shedder pairs. Surprisingly, in one of the pairs, only the good shedder's DNA was obtained from the plastic tube, with no evidence of a mixture including the poor shedder!”

  Results from Vecchiotti’s laboratory published in 2014 are consistent with this result.  Suppose that David Evans shook hands with Ms. Mangum when she arrived. That or a number of other events, such as handling money, might also transfer DNA.

  As a team captain and resident of the house, Mr. Evans might have shaken hands with Ms. Mangum when she entered or touched money that she later handled.

As persuasive as these arguments are however, one even more powerful argument may also be offered in Mr. Evans’ defense using the tables that Professor Giannelli presented in the chapter “DNA Profiling” within the book Race to Injustice.  At least two other males contributed DNA to the sample (not lacrosse players nor other men who attended the party).  In the YSTR profile ten of sixteen loci (locations within the Y chromosome) show the presence of three contributors and two more loci show the presence of at least two contributors.  In the autosomal profile some loci (places within other chromosomes) clearly show that at least three individuals contributed DNA.  This is very problematic for the prosecution.  If one acknowledges that this DNA arrived in a way unrelated to a sexual assault, then how is it possible to exclude the possibility that Mr. Evans’s DNA also arrived innocently?

By itself, the presence of what may be Mr. Evans’ DNA on the fingernails seems inculpatory; in the context of the other profiles and given the strong possibility of secondary transfer by multiple mechanisms, it is close to meaningless as evidence.  Paul Giannelli summed up the DNA on the plastic fingernails: “There was some probative value--but not much.”  Followers of the Knox-Sollecito case will recognize that the presence of additional male DNA donors is basically the same situation that exists with Mr. Sollecito’s Y-chromosomal profile that is associated with the bra clasp.  The Gary Leiterman case is perhaps the starkest example of this conundrum for the prosecution; the additional profile belongs to someone who could not possibly have been present at the crime scene.

Two more issues deserve attention, one of which is the difference in how the SBI versus DNA security interpreted the mixture in the autosomal DNA profile.  Taylor and Johnson wrote (Until Proven Innocent, p, 221), “The SBI had reported to Nifong that there were ‘multiple contributors’ to the DNA mixture but that ‘no conclusion [could] be rendered’ as to whether any of this fingernail DNA was even a possible partial match with any of the players…Based on the same testing process that had led the state lab to find no basis for any conclusion at all, DNA Security offered a singularly weak conclusion: that 98% of the male population could be ‘excluded as a contributor to the mixture’ and that David Evans was in the other 2 percent.”  Section 21C of the motion to compel discovery also quotes the same portion of the SBI report, and footnote 70 emphasized that the SBI had said “that no conclusion ‘could be rendered as to the contribution of DNA profiles from the buccal swabs of the suspects’ in the minor profile BS 4522.” 

The defense was rightly concerned about this apparent discrepancy.  It is possible that DNA Security used a suspect-centered analysis, meaning that they analyzed the mixture with Evans’s reference profile in hand.  However in “Painting the target around the matching profile: the Texas sharpshooter fallacy in forensic DNA interpretation,” William Thompson observed, “In the absence of clear standards for ‘inclusion’ and ‘exclusion’, different experts evaluating the same evidence may reach different conclusions: one may conclude that a particular suspect is ‘included’, while another concludes that the same suspect is ‘excluded’.”  Indeed, Dror and Hampikian have shown that the same mixed DNA profile may be interpreted differently by different groups of analysts.  Unfortunately the interpretation of DNA mixtures is presently still a somewhat subjective process; therefore, the difference between the two labs may have an innocent explanation.  However, another take-home message from this case is that the defense should re-analyze DNA mixtures from items of evidence.

Finally, Dr. Meehan’s own DNA may have contaminated an item of evidence.  Such contamination events are not uncommon.  The ABA’s standard 4.1a states in part:

“(ix) reports of laboratory contamination and other laboratory problems affecting testing procedures or results relevant to the evaluation of the procedures and test results obtained in the case and corrective actions taken in response…”

If there were a contamination event, Dr. Meehan’s laboratory should have reported it.  However, laboratories do not always do so.  In the Adam Scott and Gary Leiterman cases, the laboratories simply ignored evidence of DNA contamination in the negative controls (negative controls are experiments which should have no DNA in them).  There are also instances in which negative controls were faked in some way, as noted by William Thompson in “Tarnish on the gold standard.”  The reasons to fake having performed the negative controls include covering up a contamination event and increasing laboratory throughput.  Given the existence of DNA contamination and of instances of samples being mixed up or mislabeled in some way, it is surprising and disappointing that anyone opposes complete disclosure of the raw data, contamination logs, standard operating procedures, and any other pertinent records.

The DNA evidence released in the fall
On 27 October 2006 Nifong released some 1800 pages of material to the defense, but still did not include a complete report.  Although he lacked training in this area, defense attorney Brad Bannon did an extensive study of the material and was able to glean the essential information that the Y- chromosomal DNA of four unidentified men (men who were not at the party or Mangum’s boyfriend) had been found in the items of evidence from the rape kit.  When this information become public knowledge, it was a major turning point in the case, one that helped to shift public opinion in favor of the three accused students.

These profiles are important for at least three reasons.  One is that the finding of DNA serves as evidence that the tests were working, in other words the results functioned as a positive control.  The technique of amplifying DNA using the polymerase chain reaction (which is central to modern profiling) is susceptible to inhibitors of DNA polymerase, the enzyme that copies the DNA.  The lack of a DNA profile might in principle, arise from the presence of inhibitors, as opposed to the absence of DNA.  Of course, it is likely that the lab ran positive controls, but this sort of information might not come to light unless a DNA expert had reviewed the case for the defense.  Two is that Ms. Mangum’s sexual activity provides an explanation for the edema noted in her examination.  Three is that Ms. Mangum had claimed no sexual activity for a week, and these results raised doubts about her credibility, which would have been a central issue if the case had come to trial.

Using the information that they had obtained a month and a half earlier, the defense teams jointly crafted a motion to compel discovery and filed it on 13 December 2006.  Besides the information gleaned above, the defense teams learned that DNA Security had tested more items than it had disclosed in its report:

There are significant gaps in the item sequence numbers…The worksheet shows that the DNA extraction was performed n July 12, 2006, on that item, as well as items labeled “16074C” and “16075C,” which are nowhere otherwise identified in the DNA Security materials.  But on this worksheet itself, under the “name” column, someone typed “Bobby” and “Owen.”  “Quantification Worksheet Q-181”75 and “PCR Worksheet Number A-464”76 reflect quantification & amplification work on those three extractions the following day, July 14, 2006.  “Analysis Worksheet Number G-592” reflects analysis of those extractions the following day, July 14, 2006.  But nowhere in the materials do the DNA profiles generated from those analyses appear.  The phenomenon repeats itself throughout the underlying materials from DNA security.77

Students of the Knox/Sollecito case are familiar with this kind of withholding of evidence, except on a larger scale; there are many samples for which DNA quantification was performed but for which no electropherogram (loosely speaking, an electropherogram is a picture of the sizes of the DNA fragments) was produced.

DNA Security’s standard operating procedures with respect to its reports were shown to be in marked contrast with its conduct in this case.
52.  Included in the materials provided to the Defendants from DNA Security are the laboratory’s standard operating procedures, which include guidelines for drafting “Reports and Documentation” of analyses performed by the lab in each case:

Reports shall include:
·      Case identifier.
·      Description of evidence examined.
·      Description of methodology.
·      Name of each DNA locus analyzed.
·      Results for each DNA test.
·      Interpretative statement of conclusions…

That the results for each test should be part of the report is very clear.  Yet Dr. Meehan chose to disregard this standard for reasons that are open to speculation.  Perhaps he acted as he did because he wished to establish a business relationship with the office of the district attorney or because of the publicity that working on a high-profile case would bring his firm.  The defense motion also noted, “the troubling effect of its limited scope of reporting is that it allowed DNA security to produce a report in this case that avoided disclosure of exculpatory evidence, either in the form of potential contamination noted in its testing, or as it relates to the discovery of DNA from multiple male sources…”  It is difficult to overstate how different the actual results are from the results as portrayed in the incomplete report from DNA Security produced on 12 May 2006.

DNA laboratory reports and the discovery of DNA evidence
The American Bar Association’s model rules make several important points regarding DNA evidence, one of which also insists that the reports be complete.  Standard 3.3a states, “(a) A summary of all DNA testing and data interpretation should be recorded promptly in a report.”  Standard 4.1a states in part, “(xi) material or information within the prosecutor’s possession or control, including laboratory information or material, that would tend to negate the guilt of the defendant or reduce the punishment of the defendant.”

Robert Mosteller discussed Nifong’s transgressions: “Nifong had an ethical duty under Rule 3.8(d) to provide the exculpatory information in a timely fashion, which is not explained further by the rule, but lacks any suggestion that prolonged unjustified delay is authorized.”  Kenneth Williams concurred, writing, “By withholding exculpatory evidence from the defense, Nifong violated the U.S. Constitution, the laws of North Carolina, and Rule 3.8d of the North Carolina Rules of Professional Conduct.  Specifically, the State Bar found that ‘Nifong did not make timely disclosure to the defense of all evidence or information known to him that tended to negate the guilt of the accused’ and that he ‘failed to make a reasonably diligent effort to comply with a legally proper discovery request.’”

Professor Paul Giannelli commented on the incomplete laboratory reports, “In any event, no attorney should have to search through the haystack for the exculpatory needle.  A laboratory report should be comprehensive and include a section specifying the limitations of the technique used in the analysis.  The report should also be comprehensible to laypersons.” (emphasis mine)  Professor Giannelli noted that Dr. Brian Meehan also failed to meet the standards for reports laid out by the American Society of Crime Directors/Laboratory Accreditation, even though his laboratory relied upon this organization for accreditation.

Conclusions
The circumstances surrounding the absence of evidence coupled with the presence of DNA from other men is persuasive evidence of absence; no one who attended the party sexually assaulted Ms. Mangum.  The DNA that may be from David Evans on the plastic fingernailsis very weak evidence against him; in isolation it would raise the possibility of his participation in a sexual assault.  However, there are plausible alternative explanations for how his DNA arrived; therefore, if this were the only piece of evidence in the case, it would not come anywhere close to the threshold of reasonable doubt.  In the context of the absence of Mr. Evans DNA in the rape kit and the presence of other men’s DNA on the plastic fingernails, it loses almost all significance.

Regrettably, the withholding of evidence is a common theme in miscarriages of justice; however, two factors were not in Mr. Nifong’s favor.  In addition to North Carolina’s open discovery law, the defense would also have been helped by the norms of discovery of DNA evidence as set forth in the ABA standards, which would have allowed expert review of the data in its raw form.  Without any discovery of the exculpatory DNA evidence, the case might have played out much differently, especially if Mr. Nifong had been able to escape censure.  If Mr. Nifong had brought the case to trial, it is difficult to predict how a jury would weigh the fingernail DNA evidence against potential alibi witnesses for Mr. Evans (Mr. Finnerty and Mr. Seligman had electronic alibis).  Juries have occasionally ignored strong alibi evidence in favor of weak or questionable evidence, as in the cases of Jonathan Fleming and Russ Faria.  Fortunately, the state's investigators came to the conclusion that Mr. Nifong should have, and Attorney general Roy Cooper ended the matter.

A partial list of authors and references
 William Anderson “Duke:  Why the DNA mattered.”  William Anderson teaches economics at Frostburg State University in Maryland.  He was one of the first bloggers to call attention to the many problems in the Duke lacrosse case.

William Cohan, The Price of Silence, Scribner, 2014.  Mr. Cohan is a writer on business affairs.

Paul Giannelli, “DNA Profiling” in Race to Injustice (2009), Michael Siegel, ed.
Paul Giannelli is Albert J. Weatherhead III and Richard W. Weatherhead Professor of Law at Case Western University in Ohio.  He was the Reporter for the American Bar Association Criminal Justice Standards on DNA evidence, and he specializes in the area of scientific evidence.  William Thompson explained, "The Reporter for an ABA Standards Group or Study Committee is the person whose job it is to take down and 'report' the findings, conclusions and determinations of the group or committee.  It is the reporter who prepares the first draft of the report and of the annotations and notes that accompany a report.  The Reporter usually works under the direction of a committee chair, but has independent responsibility for getting details right."


Peter Gill, Misleading DNA Evidence, Academic Press, 2014.  Peter Gill is Professor of Forensic Genetics at The University of Oslo, Norway and is a professor at the Norwegian Institute of Public Health.  He is chair of the DNA commission of the International Society of Forensic Genetics, and he has published more than 180 peer-reviewed papers.

Georgina Meakin and Alan Jamieson, “DNA transfer: Review and implications for casework,” Forensic Science International: Genetics 7 (2013) 434–443.  Professor Jamieson is the head of the Forensic Institute in Glasgow, Scotland.

Robert P. Mosteller, Exculpatory Evidence, Ethics, and the Road to the Disbarment of Mike Nifong: The Critical Importance of Full Open-File Discovery, 15 George Mason Law Review 257-318 (2008).  Robert Mosteller is J. Dickson Phillips Distinguished Professor of Law at The University of North Carolina, Chapel Hill.  He teaches Evidence, Criminal Procedure Investigation, Constitutional Criminal Procedure and co-directs the Trial Advocacy Program.

Joe Minor, “Touch DNA: From the Crime Scene to the Crime Laboratory,” Forensic Magazine 4/12/13.  Joe Minor teaches forensic science and is a forensic DNA consultant.

Suzanna Ryan “Touch DNA Analysis: Using The Literature To Help Answer Some Common Questions.”  Forensic Magazine.  Suzanna Ryan is a contributor to law enforcement and forensic magazines, and she is a consultant and expert witness.

Stuart Taylor and KC Johnson, Until Proven Innocent, St. Martin’s Press, 2007.
KC Johnson hosted Durham in Wonderland, a blog that was a major source of information related to the case.  Stuart Taylor is a freelance journalist and a contributing editor for the National Journal, specializing in law.  He is a nonresident fellow of the Brookings Institution.

William Thompson, Painting the target around the matching profile: the Texas sharpshooter fallacy in forensic DNA interpretation.” Law, Probability and Risk (2009) 8, 257-276.

William Thompson, “Tarnish on the Gold Standard.”  William Thompson is a professor in the Department of Criminology, Law and Society at the University of California, Irvine.  His areas of expertise include forensics and human judgment and decision-making.

Van Oorshot R.A., Ballantyne, K.N., and Mitchell, R.J.  “Forensic trace DNA:  a review.  Investigative Genetics 1 (2010) 1:14. doi: 10.1186/2041-2223-1-14.  Roland Van Oorshot is a member of the Forensic Services Department of the Victoria Police, Victoria, Australia.

Kenneth Williams “An Examination of the District Attorney’s Alleged Unethical Conduct,” in Race to Injustice (2009), Michael Siegel, ed.  Kenneth Williams is a member of the faculty of the South Texas College of Law in Houston, TX.

Tuesday, December 2, 2014

Gary Leiterman and DNA contamination


Jane Mixer was initially thought to be the victim of a serial killer, John Norman Collins.  Jane Mixer was murdered in1969 near Ann Arbor, MI, but the DNA testing was done until 2002, a gap of approximately 33 years.  One item of evidence from the Mixer case showed DNA from John Ruelas, and several locations (on some panty hose and a towel) showed DNA from Gary Leiterman.  Despite the fact that the state had no other evidence against Leiterman, he was convicted of her murder in 2005.  John Ruelas was never charged.

A drop of blood from Mixer’s hand was preserved, and DNA from John Ruelas was found.  The prosecutor believed that the blood was from John Ruelas, but he did not offer a satisfactory explanation of how his blood came to be there.  Because Ruelas was four years old at the time of the murder and lived about forty miles away, the possibility that his DNA arrived instead via contamination must be considered.  As defense expert witness Theodore Kessis wrote, “The unexpected and never accounted for finding of John Ruelas’ profile on evidence in the Mixer case clearly demonstrates the proposition that contamination can and does occur between samples from different cases.”

The presence of Ruelas’ DNA is most easily explained by the fact that samples from John Ruelas were also processed in the crime laboratory of the State Police of Michigan contemporaneously with the samples from the Mixer murder.  But can the absence of Mixer’s DNA in the blood drop also be explained?  In 2004 Peel and Gill (“Attribution of DNA profiles to body fluid stains,” International Congress Series 1261, pp. 53-55) performed a series of experiments, in which a good DNA shedder handled the substrate (cotton or glass) for a blood stain, either before or after the blood from a different individual was placed on the substrate.  The blood was either diluted or the stain had been left to sit for some months, allowing for possible DNA degradation over time.  They used leucomalachite green as a presumptive test for blood.  Peel and Gill wrote, “…the more dilute or degraded the stain, the higher the contribution of the substrate handler’s DNA to the resulting profile.  A positive presumptive test could be obtained from samples when a profile originating from the body fluid was no longer detectable.”  Although it is tempting to associate the DNA that one finds in a stain to that stain, such an association is occasionally in error.  In some cases substrate controls can be helpful in determining whether or not DNA is associated with a particular stain.

It is essentially 100% certain that Ruelas’s DNA arrived on the items from the Mixer case via contamination, although the exact route is unclear.   Gary Leiterman’s DNA was also in the laboratory at that time.  No body fluid could be associated with Leiterman’s DNA associated with evidence from the Mixer case.  Sub-source DNA such as this is weaker in probative value than DNA associated with a particular tissue or body fluid.  Taking these facts and ideas into consideration, Leiterman’s DNA probably also arrived via contamination. 

A second line of evidence also implies that contamination is the most likely explanation.  A lab worker performed a negative control experiment during the time that a sample from the panty hose was being tested.  The negative controls will only show the presence of DNA in an electropherogram if DNA has been introduced unexpectedly into the experiment.  Theodore Kessis wrote, “Review of the electropherograms associated with this negative control sample (NEG 041902) reveals that it was contaminated, a fact that cannot be disputed since Dr. Milligan himself labeled it with a note indicated as much (Appendix 8 – Electropherogram sample NEG 041902).  Remarkably, Dr. Milligan stated in his 7/15/02 testimony that no contamination events had occurred during the course of his testing and that if any had, he would have documented them in his reporters (p. 141-21 and 142-4).  Equally difficult to rectify here is the fact that when asked if he had ever committed an error, Dr. Milligan’s replied that he could never recall making one.”  CBS News reported that, “Lab supervisor Jeffrey Nye says he retraced every step and he does not believe there is any issue of contamination. ‘No issue whatsoever,’ he says.”  This is an astonishing statement.

This lack of disclosure of a contamination event and a similar occurrence in the Adam Scott case (Peter Gill, Misleading DNA Evidence, p. 22) demonstrates that one cannot implicitly rely upon a laboratory to report accurately the results of negative control reactions.  Yet this is valuable information; a jury might choose to discount testimony from a lab if it knew contamination had happened.  It is also worth recalling what William Thompson noted in “Tarnish on the Gold Standard,” which is that the some laboratory workers tamper with negative controls in various ways.  For these reasons full disclosure of the negative controls in the form of raw (meaning unprocessed) data is the best course of action for a judicial system.  Students of the Knox/Sollecito case will not be surprised to learn that the position of the Michigan State Police crime laboratory was foursquare against disclosure of the raw data.  Professor Thompson wrote, “The Deputy Director of the Michigan State Police issued a statement on May 12, 2005 opposing ‘the allowance of releasing raw electronic data for subsequent manipulations using software and parameters not validated by the Michigan State Police Forensic Laboratory’ and declaring that ‘it is the position of the Michigan State Police Forensic Science Division that any release of this (sic) data for processing with non-validated parameters is tantamount to evidence tampering.’”26  This is a self-evidently nonsensical position, and it also forces one to ask why the processing parameters chosen by a forensic laboratory are necessarily the optimal ones.

Conclusions
The DNA evidence against Gary Leiterman is compromised so completely by the presence of the DNA from John Ruelas that it scarcely should be called evidence at all.  Exactly how their DNA came to be on items of evidence from the murder of Jane Mixer is not known.  Dr. Theodore Kessis commented on the Benjamin LaGuer case:  “It is highly improbable that any given forensic DNA laboratory will take it upon itself to contact its accrediting bodies or the press and state for the record how often they make mistakes… To best understand the weaknesses associated with DNA testing we must rely upon the empirical, the occasions in which such deficiencies are revealed either by the press or internal review of a lab’s documentation of such problems by a defense expert.  A close look at either reveals that indeed many instances of DNA testing errors have lead to the false conviction of individuals.”  Regrettably, even the sworn testimony of laboratory personnel may be seriously in error, as in the Leiterman case.  Nor can a jury be counted upon to accurately weigh the odds of contamination.    Quite the contrary, juries sometimes discount alibi evidence, such as the Farah Jama case, or the fact that the defendant lived in one city and claimed never having been to the city where the crime occurred, as happened in the Adam Scott case.  Both cases are now generally believed to be cases of DNA contamination.

Sunday, July 20, 2014

Retesting the knife for DNA: a dubious rationale, but an exculpatory result


Part 38 in the Knox/Sollecito case

Background on the DNA profiling of Sample 36-I
In overturning the Hellmann-Zanetti acquittal of Knox and Sollecito, the Supreme Court of Cassation (SCC) published a motivations document in 2013 (the previous blog entry critiqued section 12 of the motivations report on genetic investigations).  The present entry discusses section 11, another DNA-related portion but one that focused entirely on the retest of Raffaele’s kitchen knife, the subject of two previous entries here.  One of the court-appointed independent experts, Dr. Carla Vecchiotti, had found the sample in question (36-I) by swabbing the knife in 2011.  Because this amount was far below the recommended amount of DNA for the reagent kit, she decided that the amount was too small (its concentration was estimated to be 5 pg/┬ÁL).  Pro-guilt commenters sometimes imply that because the defense had the opportunity to object to certain aspects of DNA testing in 2007, that their subsequent requests for the raw forensic data should be turned aside (this argument is problematic on multiple grounds as discussed in the previous blog entry).  Yet if we accept that an objection to some aspect of testing should be lodged immediately, then either the prosecution's observers should have objected to not amplifying 36-I in the spring of 2011, or the prosecution should have admitted that it lost the opportunity to do so when it did not.

Why did the Supreme Court of Cassation demand an amplification of the new sample?
In section 11 of their motivations report the CSC wrote,
“This trace was not subjected to genetic analysis – through a decision made by one of the experts, Professor Vecchiotti, alone, without documented prior authorization to that effect by the Court, who had also given a mandate to attribute the DNA on the present findings on the knife and on the bra hook- because the amount was not sufficient to provide a reliable result, amounting to Law (sic) Copy Number. This choice, however, met the subsequent sharing of the group, on the assumption that such a small sample would not have allowed two amplifications necessary for a reliable result (p. 84 of sentence).

“So that when the Procurator General and the civil plaintiffs demanded to complete the examination, strongly as a result of the scientific contribution of Professor Novelli, geneticist of undisputable fame recognized by the same Court (p. 79 sent.) on the availability of equipment able of operate with safety also quantities of less than ten picograms, in the areas of diagnostic character (even on embryos) in which the claim to certainty is certainly no less pressing than that which animates the legal field, the Court rejected that proposal, assuming that the methods to which Prof. Novelli had referred were "in the experimental phase” (p. 84 sent.), thus freely interpreting and misrepresenting the assumption of bias, which in fact was to remember the use of these diagnostic methods in areas in which you can be certain of the result.

“…Especially since the renewal of genetic investigations were requested in 2011, after four years from the initial time and over which the evolution of instrumentation and methods of investigation had marked significant milestones, as was emphasised by the advisor to the Procurator General, Professor Novelli. Just on receipt of the information from the consultant mentioned who - under the constraint of the obligation to truth, spoke of cutting-edge techniques -, the Court fell into a new gross misrepresentation of argument concerning the reliability of the results of investigations carried out assuming no new findings of such remedies, even through developments emerging at a later time, concerning reasonableness of the grounds (Section I, 25.6 .2007, n. 24667).

“…[the survey must] bring to analysis also the newly sampled trace, according to the most accurate and modern “experimental” analytical techniques, under pain of violation of the law for not making a decisive test and the fallout in terms of manifest illogical reasoning (again for obvious incompleteness of the inferential platform, to have overlooked data that is not only important, but crucial), as was correctly pointed out by the public plaintiff.”

Leila Schneps and Coralie Colmez summarized Novelli’s testimony and offered their opinion in the chapter, “The test that wasn’t done,” in their 2013 book Math on Trial (pp. 83-86):  "In court on September 5 [2011] and supported by expert prosecution witness Giuseppe Novelli, Stefanoni explained that newer generations of DNA analysis kits existed in 2011 that had not been available in 2007, and these new kits could give results on as small as a couple of cells.  She wanted a new analysis performed to confirm that her previous work was correct.  The prosecution agreed and asked the judge to order the new tests...Judge Hellmann missed a major opportunity to get at the truth.”  According to the CSC, techniques that are appropriate for diagnosis in embryology are also appropriate for forensic investigations.  There have been advances in technology, and the new tests should take advantage of them.  The results will be "decisive."

Not every observer sounded convinced of the probability-based rationale Schneps and Colmez put forward.  Professor Daniel H Kaye wrote, “If these experts’ concern — that the original DNA test was simply detecting traces of Kercher's and Sollecito’s DNA that investigators inadvertently transferred to the knife and bra clasp, respectively — then repeating the tests could well continue to detect that DNA — and prove nothing more than the original tests did.” In other words one weakness in the rationale provided by the SCC is that a newer generation of kits cannot circumvent the problem that one is dealing with DNA in the low template range (see below). 

Can one equate pre-implantation genetic diagnosis with DNA forensics?
It may be that Professor Novelli left the court with the impression that single-cell PCR was new technology, but single-cell PCR in diagnostics is at least twenty years old.  Yet the problem with using diagnostic practices to justify further low template testing on the knife is that it ignores that one is still dealing with low template quantities and all the problems that such analysis has in a forensic setting.

In the article “LCN DNA Analysis: Limitations Prevent 'General Acceptance'” Dr. Angela von Daal wrote, "The use of PCR for analysis of very low levels of DNA in the field of pre-implantation genetic diagnosis (PGD) has been used to justify general acceptance of LCN analysis in the wider scientific community(12). This argument is flawed. PGD analysis is not analogous to LCN analysis for several reasons. PGD uses pristine cellular DNA from a single source, whereas forensic LCN samples are mostly mixtures and are likely in a state of apoptosis. The complexities of profile interpretation issues seen with STR analysis (e.g., stutter) are not an issue for PGD testing(13). However the most significant difference is that the samples derived from the mother, father and embryo are single-source and the parental samples are of known genotype."  In other words the connection between pre-implantation genetic diagnosis and low template DNA forensics is tenuous and trying to extrapolate from the former to the latter is dubious at best.  Why Professor Novelli chose to ignore the problems in his analogy is something only he can explain.

Did the SCC understand the unique problems of low template DNA profiling?
And the problems of innocent transfer of DNA get more severe as the sample size gets smaller.  Sara Gino testified for the defense in the trial of the first instance, and some of what she had to say is pertinent to this issue. From the Massei report (p. 258, English translation): “She reaffirmed that [the risk of] contamination exists, and emphasised that in minimal quantities of DNA there is not necessarily a greater risk of contamination but it was easier to notice the effects of the contamination and be misled (‘...It's not that the risk of contamination is greater; but it is easier to see the contamination...’ page 92).” In response to a question on this subject, Professor Dan Krane expounded on Sarah Gino’s testimony, “There is absolutely no question but that contamination is a much greater problem in LCN cases than conventional DNA testing. The reasons that it is a greater problem are both because it is easier to detect contaminants ([Sarah] Gino's point) and because it is easier to transfer (and to transfer without knowing) smaller amounts of DNA than larger amounts of DNA.”

Proper low template profiling demands that one rework the entire process of DNA forensics, from collection of the evidence all the way through the final step of analysis of the electropherogram.  True low copy number LCN profiling requires ultra-clean, dedicated facilities in order to perform it properly, as clearly discussed in nontechnical language in an article in the New Zealand Herald. Without those extra precautions, the odds of contamination are necessarily greater (or else the precautions are a waste of time and money).  In a review article in Nanomedicine Professor Novelli and coauthors wrote, “Very few laboratories perform low template DNA typing properly, because it requires dedicated facilities and great experience, although there are several published methods for the interpretation of such profiles [80-82].”  This blog has previously discussed the need for proper air handling facilities.

Even so, within the forensic DNA community there are a number of critics of LCN profiling.  Dr. Budowle and colleagues, for example, argued that low template analysis should be restricted to identification of bodies and the generation of leads.  In other words using the practices in embryology to justify further testing on the knife was predicated on a very weak analogy and one that has been seriously questioned by molecular biologists.  Professor Novelli should have known this and should have acknowledged this limitation to the court.

What other forensic issues did the Supreme Court of Cassation ignore?
In forensics it is impossible to interrogate the DNA as to the time or manner of its deposition.  With respect to the knife, the DNA may have arrived from secondary or tertiary transfer prior to its collection or during its handling.  Regrettably, the knife was removed from its package at the police station, a breach of good practice.  The person who repackaged it had previously been to the women’s apartment earlier that day, a second error.  In addition, the negative controls have never been produced in the form of raw data or probably any other form.

What did the proponents of the retest expect to learn?
As this blog has repeatedly argued, the DNA from Sample 36B in 2007 was probably from laboratory contamination.  Pro-guilt commenters have resisted this interpretation.  In a story from 27 April 2013 the BBC’s Ruth Alexander asked Coralie Colmez this question.  “’So what this means in the case of the knife in the murder is that if it were tested again, and once again the DNA was Meredith's profile we could be a lot more certain that the DNA on the knife is indeed Meredith's,’ Colmez says.”  On the other hand suppose that the test came out differently.  “And if the knife were tested again and the DNA did not match Meredith Kercher's profile? That would be good news for Knox and Sollecito, she says.
‘This would mean that this major piece of evidence against them would be discredited.’”  In other words either way the results turned out, there would be useful, arguably decisive, new information.

What did the new test actually tell us about the knife?
Andrea Vogt wrote, "The RIS Wednesday deposited their forensic report on trace 36i, a spot of DNA identified (but not earlier tested) on the kitchen knife alleged to be the murder weapon. “Cento Percento” (100 percent) said Major Berti, discussing compatibility. The RIS found that the DNA was compatible with Amanda Knox, and excluded that it was that of Sollecito, Guede or Kercher. The RIS expert was asked only a few questions from attorneys and the judge. The judge asked why the RIS had done two amplications of the DNA and not 3 or 4. Major Berti described that two is considered the minimum number of amplifications necessary, according to today’s forensic standards, doing less (or more) might have diminished the reliability of the results. The judge also asked about the age of the equipment used. Berti responded that the forensic kit used this time has been commercialized since 2010 and available for use since 2011. At one point the judge stopped a line of questioning by Knox’s Rome attorney Carlo Dalla Vedova, who was asking why the RIS described Knox’s DNA as “fluids” when a prior expert had said the trace did not come from blood. Nencini said: That question was not put to the RIS by this court, it was not their job to determine that. The other experts’ reports are in the case files for everyone to read, he noted, adding: “We cannot put words in the mouth of this expert that were said by another expert.”

The presence of Ms. Knox’s DNA on the handle knife has never been in dispute, and the finding of additional DNA on the blade is not inculpatory.  A recent review article on trace DNA noted that DNA transfers can occur after an item of evidence is packaged; therefore, the DNA on the blade may have arisen from transfer after the knife was packaged, or from direct transfer: chef’s knives are often gripped both at the handle and the base of the blade.  If anything, the result is modest additional exculpatory evidence.  If the knife were the murder weapon, it would have Meredith’s blood.  Somehow all of Meredith’s blood would have to be removed, yet DNA from Meredith and Amanda would remain, along with starch.  A letter to the court from Professor Bruce Budowle indicates that it would be difficult to clean a knife of blood.  There was additional DNA in sample 36-I belonging to an unknown person, a finding which further calls into question the hypothesis that it was the murder weapon.

Are the new results exculpatory?
In early 2014 Leila Schneps wrote, “By the third trial, when a new attempt was made to collect DNA from the knife (which had been swabbed again during the appeal trial, though no tests were then conducted) there was no match to Meredith – a result welcomed by Knox's defence team, though it did not in fact impact on the findings of the first trial.”  This passage presents quite a contrast with the earlier quote from Coralie Colmez (above).

There are two important issues.  The first point is that Meredith Kercher’s DNA was not found, and this was the whole reason for performing the test.  The people who argued for the test did so on the basis that the retest might improve the reliability of the 2007 result.  They should at the very minimum concede that the original result on the knife has now been “discredited,” as Ms. Colmez wrote.  The second point is that the 2013 result did indeed impact on the finding of the first trial.  The Carabinieri noted that at least two amplifications of the DNA must be done, and Stefanoni apparently did only one.  Therefore, the lack of retesting with respect to the original result is indeed makes it unreliable by the expert testimony of the Carabinieri, which support the testimony of Conti and Vecchiotti.  In other words the result and the testimony are a one-two punch that should have knocked the kitchen knife right out of the trial.

Is the Nencini court being impartial and objective with respect to the forensics of this case?
An early report from Andrea Vogt on Judge Nencini’s motivations document from the Florence court indicates that Nencini harshly criticized the rationale for Conti and Vecchiotti’s not completing the test in 2011.  For the reasons given above, this argument has no merit.  A cynical observer might conclude that it is a canard to distract future courts from the real issue, which is that Meredith Kercher’s DNA was not found, and this was supposed to be a decisive test.  Pro-guilt commenters might now argue that the test would have been decisive if Meredith’s DNA had been observed in 36-I but any other result is inconclusive.  However, such an argument prompts a question:  Why should the Nencini court order a test that might only be beneficial to the prosecution and not also grant requests that are favorable to the defense?  This blog has long argued that the failure of the prosecution to release the raw DNA data constitutes a serious breach of discovery, rendering the trial unfair.

Another serious logical problem with the SCC and the Nencini court is the selective indignation about the lack of amplification of 36-I in 2011.  Mrs. Stefanoni has failed to produce many electropherograms which showed positive quantification of DNA, some of which may contain highly probative evidence.  The defense has every right to see these data under any reasonable standard of discovery.  One profoundly wishes that the Nencini court had demanded the production of the electropherograms that originated from a putative semen fraction, to take just one example.  The fact that it did not do so is only one reason to call its objectivity into question.

Update (13 August 2014)
In the Lindy Chamberlain case a forensic worker, Joy Kuhl, invented a new test for fetal hemoglobin.  The defense witnesses were sharply critical of her methods.  “On the matter of specificity and testing solution, [Ian] Barker wanted to know whether [Professor Richard] Nairn believed that over two hundred tests, referring to Kuhl's tests, which showed a negative reaction to adult haemoglobin would be sufficient to conclude that the anti-sera was not specific to adult haemoglobin. Nairn answered that it would depend on the test, a bad test is a bad test no matter how many times it is repeated.”  Another author quoted Richard Nairn: "Two hundred bad tests are poorer than one good test."