The likelihood of DNA contamination

Part 31 in the Knox/Sollecito case

The subject of DNA contamination has been a frequent topic of this blog. DNA contamination is again near the forefront of the Kercher murder and the second trial of Amanda Knox and Sollecito. The court-appointed, independent experts, Drs. Conti and Vecchiotti, have issued a report that raises the strong possibility of contamination. The odds that a forensic DNA sample are contaminated are hard to determine yet very important. Let us first examine a specific statement made with respect to environmental contamination and this case, then look at the question more generally. A subsequent entry will examine the collection practices of the Rome lab and Conti’s and Vecchiotti’s evaluation of their work in this case.

Professor Giuseppe Novelli, a researcher into medical genetics and forensic DNA profiling, said, «Il contaminante va dimostrato, dove nasce e dove è. Il gancetto contaminato dalla polvere? Più probabile che cada un meteorite e butti giù questo tribunale» "The contaminant needs to be demonstrated, where it comes from and where it is. The clasp contaminated by dust? It's more likely that a meteorite comes down and knocks down this courthouse." (translation by komponisto)

Let us assume that Dr. Novelli is specifically referring to household dust. The potential for dust being an issue in forensic DNA testing first came to light with the publication of a paper from Bonnie Brown and coworkers: Toothman et al., “Characterization of human DNA in environmental samples,” Forensic Science International 178 (2008) 7–15. These workers sampled dust from offices, research laboratories, and classrooms.

Figure 3 in this paper is an electropherogram of dust from a classroom, and it shows peaks of 1000 to 3500 RFU corresponding to 3-6 alleles in some loci associated with shorter pieces of DNA. As the length of the DNA fragments increases (moving from left to right on the elecropherogram), the height of the peaks decreased. The authors point out that this is consistent with DNA template that is partially degraded. The combination of a sample’s being a mixture and its yielding only partial profiles makes it very difficult to identify individual profiles.

The authors wrote, “Results of this study have implications regarding the processing of forensic samples. First, the presence of genotypeable human DNA in dust illustrates a significant
potential contamination source in forensic investigations. Twenty-five of 36 samples contained sufficient input human DNA for STR analysis using the AmpFlSTR® Profiler PlusTM assay (~1.0 ng), and 36% (including low-input samples) produced alleles at multiple loci. These results demonstrate that even though anti-contamination measures may be in place at a crime scene and the laboratory, trace DNA derived from dust in the vicinity of other evidence is capable of producing signals higher than background noise in STR analyses.”

Trace DNA and the environment
Secondary transfer is the movement of DNA from a donor to an intermediate object then to another object from which it is collected. Some forensic scientists only classify it as contamination when it occurs subsequently to the object’s being taken into custody; however, other workers prefer to treat innocent secondary transfer as equivalent to contamination on the basis that neither is relevant to the investigation. Both secondary transfer prior to an object’s being collected and contamination need to be understood more thoroughly for trace DNA to be used routinely in DNA forensics. Van Oorschot and colleagues wrote, “Greater effort needs to be made by police/crime investigators to investigate how a DNA sample arrived at the location where it was found, as well as by scientists to better understand the impact of activities on the relative amounts of DNA from particular sources at a crime scene… Some preliminary contributions to our knowledge of transfer in relation to residential burglary and street robbery have recently been made [67].

In their review of trace DNA in forensics Van Oorschot and colleagues suggested six remedies to address the problem of contamination:
“1. perform more studies similar to those of Raymond et al. [67], Cook and Dixon [202], Dowlman et al. [203] and Toothman et al. [201] in order to learn more about the occurrence and persistence of DNA on particular surfaces in different environmental conditions” Understanding how prevalent and persistent background DNA is in the environment is far from being completely understood. Dr. Novelli declined to comment for this report, but his comparison with a meteorite is premature, at best.

The relationship between DNA sample size and the ease of transfer
In their 2010 review article Van Oorschot and colleagues wrote, “Contaminant DNA may appear as either the major or minor sample within a mixture or, alternatively, may overwhelm the target DNA completely.” One explanation for this somewhat counterintuitive statement is that contamination in the laboratory may introduce DNA from previous PCR amplifications by a number of possible routes. However, the two main DNA profiles of interest (Meredith’s on the knife blade and Raffaele’s putative profile on the bra clasp) both involve relatively small amounts of DNA; therefore, we will focus on small samples.

The discussion of low copy number (LCN) testing from the Crown Prosecution Service noted, “This increased sensitivity means ultra-clean laboratories are needed for the testing to minimise contamination of the sample by DNA from any other source.” The New Zealand Institute of Environmental Science and Research has spent $1 million building anticontamination areas for low copy number (LCN) DNA forensics. The New Zealand Herald wrote, “The bogey is contamination. The very sensitivity of the technique which enables it to extract a DNA profile from the tiniest sample also makes it extremely vulnerable to contamination. Stringent measures are needed to minimise that risk… We live in a ‘soup’ of DNA, explains ESR forensic programme manager Keith Bedford. ‘If I were to shed dandruff, massive amounts of dna could fall ... hair could carry DNA. The way I am speaking at the moment, we could probably detect DNA on this pad in front of me.’”

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 responded, “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 ([Sara] Gino's point) and because it is easier to transfer (and to transfer without knowing) smaller amounts of DNA than larger amounts of DNA.”

Some examples of DNA contamination
Farah Jama was a young man accused of rape on the basis of his DNA seemingly being found on the alleged victim. Mr. Jama is black and at 21 was too young to have entered the club at which the incident occurred, which catered to people over 28. Moreover, the alleged victim did not recall seeing a black man at the club that night. Yet, as Milanda Rout wrote, “But the judge and the jury did not buy his alibi, despite supporting evidence from his father, brother and friend. Instead, they believed the forensic scientist who testified there was a one in 800 billion chance that the DNA belonged to someone other than the accused man.” After Mr. Jama spent more that eighteen months in prison, he was released because prosecutors said that they could not rule out contamination. The contamination event may have occurred during two forensic medical examinations, one of the victim and the other of Mr. Jama on an unrelated matter that occurred one day earlier.

Perhaps the most thoroughly studied case of contamination is that which occurred in the Jaidyn Leskie case. This blog has covered the Leskie case on two previous occasions. The DNA of a woman who probably never left her village was found on the clothing of the submerged body of a toddler. She was a mentally challenged woman who may have been raped, which is why her DNA was being examined. Because the woman was such an exceedingly unlikely suspect, the only reasonable explanation was contamination. Contamination has been documented on several occasions when evidence items from unrelated cases are examined within a few days in the same lab.

Russell John Gesah was charged with the rape and murder of a mother and child. Kathleen Skeen wrote, “A Victorian Police Forensic Services Centre review found clothing with Mr Gesah's DNA from an unrelated offence had been examined on the same day and same surface as clothing from the Tapp case." The Gesah case, and the murders of Jane Mixer and Jane Durrua (see below) are all examples of DNA cold hits. The Gesah case prompted the State of Victoria to reexamine thousands of cases (see below).

Gregory Turner might have been convicted of murder on the basis of DNA evidence. However, a forensic worker contaminated a key piece of evidence with his and her DNA. She also acknowledged contamination in two other cases on which she had worked. An interesting aspect of the Turner case is that the DNA from the victim came from her fingernails, and Mr. Turner’s DNA came from his wedding ring. These facts suggest that the presence of liquids is not necessary to bring about cross-contamination, in contrast to the implications of Patrizia Stefanoni’s testimony in the present case.

The murder of Jane Mixer was initially attributed to a serial killer. When Gary Leiterman’s DNA was found on the decades-old evidence, he was convicted. However, the presence of the DNA matching then four-year old John Ruelas on the same item of evidence (despite Ruelas’s living in another city) strongly points to this being another example of contamination. This illustrates another important principle. One does not always know the precise moment that contamination occurred, but one can infer contamination when the direct deposit of DNA is shown to be highly unlikely.

A seemingly solved cold case that turned out to be contamination involved the 1968 murder of Jane Durrua. Jerry Lee Bellamy’s DNA was found when the evidence was tested in 1999. Evidence against Mr. Bellamy in an unrelated case was tested on the same day as items from the Durrua case. The actual evidence of contamination was not conclusive, but despite this, charges against Mr. Bellamy were dropped. Alleged serial killer Robert Zarinski was later arrested, but he died before he could be tried.

The difficulties in quantifying the frequency of DNA contamination
Not all labs document contamination events. Some labs argue that contamination that is identified with the use of negative control experiments does not count as contamination. Negative controls will spot wholesale contamination events but will not necessarily catch sporadic contamination. These facts make it difficult to quantify how frequently contamination occurs. However, it does not seem to be an especially rare event. Professor Thompson is a lawyer who specializes in probability theory as it relates to DNA profiling. Maura Dolan reported that he is among the leading authorities on laboratory errors in the United States. In response to a request from the Los Angeles Times to review the records from some California forensics labs, Thompson said, “’on a regular basis, laboratory personnel make mistakes that could lead to false identifications’ of suspects.” He also indicated that what has emerged in recent years is just “the tip of the iceberg.”

In 2008 Professor Thompson wrote an article, “The Potential for Error in Forensic DNA Testing (and How That Complicates the Use of DNA Databases for Criminal Identification"). “Doubt was also cast on a number of convictions in Queensland when a forensic scientist who had previously worked for a state forensic laboratory publicly expressed concerns about the reliability of the lab’s work. He told The Australian newspaper that it was not uncommon for the lab to mix up DNA samples from different cases.[62] For example, he said that analysts’ own DNA, from blood samples used as analytical controls, often was mixed up with (or found its way into) casework samples, creating false matches: “[Q]uite often my (colleague) would walk down the aisle and say, ‘I’ve just committed another rape on the Gold Coast.’”[62] The analyst said that while many such errors were caught, sample limitations made it impossible to resample or retest in some questionable cases.” These remarks underscore the notion that DNA contaminations are not a rare event.
[62. A. McDonald, “DNA evidence claim clouds Australian convictions,” The Australian, July 8, 2006.]

In response to the Russell John Gesah contamination incident (see above), the Victorian police reexamined their cases involving DNA forensics. During the period from 1988 to 2008 the Victorian police service handled 7000 cases involving DNA. According to Peter Gregory and coauthors, “In 2003, Mr Scheffer told an inquest on Moe toddler Jaidyn Leskie that since late 1999, 39 cases had been identified as requiring "diagnostic and corrective action", with most involving contamination.

Finally, testimony reported by Annabelle McDonald (in The Australian) implied that mixing up samples is a not uncommon event. Although mislabeling of samples (if that is what mixing up means) is not itself contamination, it has the potential to lead to the same erroneous judicial result. A mislabeling in Nevada was uncovered during an independent review of the Lazaro Sotolusson case. In addition Dwayne Jackson was also the victim of a similar mistake at the Las Vegas forensics lab.

Conclusions
There is not yet enough information on environmental contamination to make conclusive statements about how common environmental contamination is; however, DNA is deposited routinely in all sorts of ways that are unrelated to criminal activity. The authors of a recent study believe that environmental dust can give rise to extra alleles in evidence samples. The frequency of contamination is difficult to quantify, but it is not an especially rare occurrence. The chances of contamination are greater for DNA in the low template range than they are for larger samples. Historical examples of contamination suggest that it is more likely to occur when items of evidence are processed closely in time. Contrary to the implication of Dr. Novelli's remarks, it is rarely the case that the exact mechanism of contamination is proven.