Former Visiting Professor Patrick Holford has such a sensitive moral barometer that he is constantly pointing out his perception of the failings of actual researchers and scientists and questioning their integrity. Operating in this parallel world that he does (see Part 1 and Part 2), perhaps it is not surprising that he continues to avoid issuing an update on what the findings from the Autism Omnibus and the stark revelations of the fraud and deliberate manipulation that underpins Dr Andrew Wakfield’s research means for his marketing and promotion of unevidenced diagnostic tests and diet strategies for the ‘treatment’ of autism (assessed here).
You may remember Patrick Holford’s keen support for Andrew Wakefield and his research (see, also, Patrick Holford and Andrew Wakefield’s Discredited Findings: Part 1 and Part 2). we have previously noted that Holford espouses support for Wakefield and his research allows some entrepreneurs to sell unevidenced diagnostic tests (both Wakefield and Holford continue to support the use of Secretin despite not only the absence of efficacy but the indication that it may be less efficacious than a saltwater placebo), promote consultation for difficult-to-follow diets and sell supplements.
The idea that some behaviours result from opioid peptides derived from incompletely digested proteins is built on Panksepp’s speculations concerning his observations of rats and their response to morphine.
We have approached the possible neurochemical causes of autism by assuming that the fundamental problem of the autistic child is emotional. Some of the earliest observed symptoms of autism include a lack of crying during infancy, a failure to cling to parents, and a generally low desire for social companionship, which, we believe, shows that the autistic child is constitutionally unable to feel properly the emotions arising from social relationships. Injections of low doses of morphine can generate such behaviour patterns in animals..
The speculation was combined with several other reports and received undue prominence in the Wakefield et al. 1998 paper.
The “opioid excess” theory of autism, put forward first by Panksepp and colleagues and later by Reichelt and colleagues and Shattock and colleagues proposes that autistic disorders result from the incomplete breakdown and excessive absorption of gut-derived peptides from foods, including barley, rye, oats, and caesin from milk and dairy produce. These peptides may exert central-opioid effects…
The typical explanation for the sequence of events is as follows. An insult injures the gut wall. The gut wall becomes ‘leaky’ and there is a general disruption caused by inflammation that also dysregulates the digestion of dietary proteins. These partly-digested proteins are opioid-like; they escape into the blood-stream and are transported across the blood-brain barrier where they intoxicate the brain, resulting in damage that manifests as a spectrum of symptoms labelled ASD.
Thus, the Gluten-Free, Casein-Free (GFCF) diet for children with ASD was born and it continues to be recommended despite the lack of any scientific support for its theoretical underpinnings. However, Patrick Holford writes about it regularly and it is the bedrock of his recommendations for restrictive diets and supplements.
So, with Brian Deer’s recent revelations that must remove any remaining shard of plausibility for the 1998 paper even for Wakefield’s most ardent supporters, and the recent rulings from the Special Court for the Vaccine Injury Compensation programme (aka, the Autism Omnibus hearings) that reserved harsh criticism for doctors who mislead parents as to the causes of their children’s health issues and was strongly critical of Wakefield’s research. The evidence at the hearings pitilessly exposed the inadequate conditions at the laboratories that supposedly confirmed Wakefield’s hypothesis: see, e.g., Special Master Hastings’ findings (ftp and pdf).
[T]he petitioners have failed to demonstrate that thimerosal-containing vaccines can contribute to causing immune dysfunction, or that the MMR vaccine can contribute to causing either autism or gastrointestinal dysfunction… [pg. 2]
I have not required a level of proof greater than “more probable than not,” which has also been described as “50 percent plus a feather.” I understand fully that petitioners are not alleging that the MMR vaccine was the sole cause of Michelle’s disorders, but are alleging only that the vaccine contributed to the causation of her disorders, possibly in concert with an underlying genetic vulnerability…
This case, however, is not a close case. The overall weight of the evidence is overwhelmingly contrary to the petitioners’ causation theories…The petitioners’ evidence has been unpersuasive on many different points, concerning virtually all aspects of their causation theories, each such deficiency having been discussed in detail above. The petitioners have failed to persuade me that there is validity to any of their general causation arguments…
In short, this is a case in which the evidence is so one-sided that any nuances in the interpretation of the causation case law would make no difference to the outcome of the case…
After studying the extensive evidence in this case for many months, I am convinced that the reports and advice given to the Cedillos by Dr. Krigsman and some other physicians, advising the Cedillos that there is a causal connection between Michelle’s MMR vaccination and her chronic conditions, have been very wrong. Unfortunately, the Cedillos have been misled by physicians who are guilty, in my view, of gross medical misjudgment. [pp. 172-173; Cedillo v. Secretary of Health and Human Services (ftp and pdf)]
The Special Masters’ findings clearly indicate that:
- the basis for the claim that vaccine damage contributes to autism has never been based on more than speculation and correlation
- the Petitioners’ Steering Committee asked for and received many time extensions to allow them to assemble the best case that they could
- Petitioners’ Steering Committee vacillated as to the format of the hearings but settled on one that they believed would best suit them and they evidence they believed that they had
- Petitioners’ Steering Committee not only received assistance from the court to gather evidence but were admonished for not being more effective in pursuing it. The Special Masters and Respondents both volunteered to support Petitioners’ Steering Committee in obtaining more evidence but the PSC failed to act in a timely manner.
The nett result of both this and the review that determined the discontinuation of the UK Legal Aid case is that the people who are arguing for vaccine injury as a general causation theory for autism have no supporting evidence despite generous financial support to discover and assemble it.
To conclude that Colten’s condition was the result of his MMR vaccine, an objective observer would have to emulate Lewis Carroll’s White Queen and be able to believe six impossible (or, at least, highly improbable) things before breakfast. The families of children with ASD and the court have waited in vain for adequate evidence to support the autism-MMR hypothesis. [pg. 278; Snyder v. Secretary of Health and Human Services (ftp and pdf)]
All of the Special Masters address the issue of Andrew Wakefield and his research. Singling out Special Master Vowell, there is a dedicated section (Section VI) that provides a very clear summary of the evidence that relates to the ‘Measles Theory’.
The MMR vaccine is usually administered to children in the U.S. between 12-15 months of age,  shortly before parents first begin to notice the behavioral symptoms that eventually lead to their children’s ASD diagnoses. It is not surprising that some parents considered the vaccine to be causal. The 1998 publication of a paper  suggesting a temporal relationship, and implying a causal one, between the MMR vaccine and onset of autistic symptoms in a group of 12 children being treated for gastrointestinal complaints–a symptom not uncommon in autistic children–added fuel to smoldering suspicion. The paper’s primary author, Dr. Andrew Wakefield, later advanced a hypothesis as to how measles virus could cause both the gastrointestinal complaints and autism. Although Dr. Wakefield’s hypothesis differs in some respects from the theories advanced in this case, petitioners’ theories are its linear descendants… [pg. 113]
[T]heories rely upon a link between gut disorders in autistic children (“autistic enterocolitis” ) and the presence of measles virus genomic material in their gut tissue…The evidence supporting the presence of measles virus in the gut tissue was provided primarily by Dr. Wakefield’s research and by the testing performed at Unigenetics laboratory… [pg. 114]
Respondent mounted a vigorous challenge to the scientific validity of reports of measles virus genomic material in autistic children in addition to challenging the biological plausibility of, and the logical connection between, the theories. Challenging the scientific validity both of Dr. Wakefield’s research and Unigenetics’ testing, respondent relied upon experts in many disciplines.
This section begins with a discussion of how the MMR theory first arose, followed by a more detailed explication of the current theory. The discussion of “autistic enterocolitis” is followed by the evidence concerning the wild strain measles virus, covering how the vaccine strain of the virus differs from the wild-type virus, their particular effects on the immune system, and the diseases commonly recognized as being caused by the virus strains. Finally, this section examines PCR testing, problems commonly encountered in such testing, and the problems demonstrated in the Unigenetics testing program in particular. [pg. 115]
Special Master Vowell’s summary of the evidence and testimony (ftp and pdf) makes painful but essential reading. It is clear that there have been grave reservations and even well-grounded indications that Wakefield should have redacted his research findings from the 1990s. The extracts that cover the comprehensive rebuttal of Wakefield’s work and the conditions at the testing laboratories are in the appendix. They are well worth reading if only because it is rare to see such candour permitted. There is frequent reference to “fraud” and “deception” as well as the frank description of Wakefield et al’s 1998 paper as, “probably the worst paper that’s ever been published in the history of [Lancet]” [pg. 118]. Dr. Wakefield’s theory was described by verifiable experts, with supporting evidence for their descriptions as “incredible,” “fantastic,” “improbable,” and “not based on any data”. If anything, the assessment for the 2002 paper was even worse than for the 1998; it was characterised as “scientific deception”.
There is a timeline that indicates that there were problems with testing samples in Wakefield’s work even when he was working on the hypothesis that measles virus is causative for Crohn’s disease (see, e.g., Dr Rima’s testimony in the appendix below and we shall publish a full timeline in the near future).
The results from the Unigenetics and Kawashima testing laboratories could not be replicated by researchers in laboraties with high standards. Investigations of the conditions at the Unigenetics laboratory were over-whelmingly negative and there are indications of negligent practices including faulty machinery and more than a hint of fraud and deceptive practices, including “planted evidence” [pg. 188]. The Kawashima testers agreed to participate in a bench-marking exercise with other laboratories that are known to have a fine reputation for the probity of their results: the Kawashima results were discordant with those from other labs.
The key part of Wakefield’s research that depends upon the assertion that the measles virus persists in the gut disappeared. It also took with it the elaboration of his research by others who claimed that measles virus could be detected in cerebro-spinal fluid. Unfortunately, the same laboratories were responsible for those results.
Experts who examined the Wakefield data pointed out that there were gross mistakes in interpreting the diagnostic photographs from the endoscopies: even mistakes as basic as ‘confusing’ the ileum and cecum (the witness in that instance characterised this as “something of a deception”, pg. 121). Taken all together, there is no support for the existence of Wakefield’s new pathology of ileal lymphonodular hyperplasia (ILNH).
There is no evidence to support any of the more elaborate theories that other researchers developed from Wakefield’s research. Dr Dave Gorski has written up a detailed account both of the Deer revelations and the findings from the Autism Omnibus: 2009: Shaping up to be a really bad year for antivaccinationists. He gives a comprehensive and accessible explanation for the implications of the findings of error, manipulation and fraud.
All of the above addresses Wakefield’s own research which relied upon the testing facilities that he used. His ‘in-house’ expert, Nick Chadwick had informed him that there were problems with the testing facilities. Many other experts informed Wakefield that there were substantial problems and did so in a timely fashion (see appendix). However, Wakefield persisted in relying upon these results that are described in some of the clear and most unequivocal terms as ‘unreliable’ to even ‘fraudulent’.
Patrick Holford, for one, relies upon unevidenced laboratory tests to establish food ‘allergies’ and to establish heavy metal toxicity. Special Master Vowell details some of these tests that were run on Colton Snyder and emphasises that the tests, like the treatments that were administered, are without independent validation, far less an evidence base, they have even been known to kill.
Until the ASA put a stop to Patrick Holford’s self-aggrandising flights of fancy about himself, he used to describe himself as “Britain”s most informed, independent health expert”. We side with the ASA on the actual veracity of that claim but if Holford wishes to confer such status on himself, it is time he accepted responsibility for his pretensions to leadership.
It is two weeks since Brian Deer’s revelations and 10 days since the Special Masters published their findings. As a matter of some urgency, Patrick Holford to issue an update of his previous work and to withdraw his current recommendations for ‘treatment’ plans for autism. If he does not, he gives up any pretence that he has to staying ‘up to date’ with research literature and delivering a distillation of ‘research you can use to improve your health and that of your family’. If he decides to continue his current support for Andrew Wakefield, then he owes it to the people who pay him an expensive subscription for his advice, or who have purchased his books, that he should explain why he is doing this in the face of what has been described as “overwhelming evidence” to the contrary. Plus, anyone who followed his posturing over his absurd claims about the efficacy of AZT as compared to vitamin C in the treatment of Aids should know how long it takes him to issue a non-clarifying clarification that he somehow thinks exonerates his egregious errors.
Because there were no studies to detect measles virus in gut tissue before Dr. Wakefield’s research, two steps were necessary to develop evidence that measles virus might cause gut disorders. First, Dr. Wakefield and his team needed to establish the presence of persistent measles virus in gut tissue. If present, they still needed to establish that its presence was abnormal and associated with a disease process. To do so, they needed to show that it was present significantly more often in those with gastrointestinal disorders than in those without such disorders. They failed on both accounts.
Doctor Wakefield published other papers, in 1995  and 1997,  stating that measles virus causes Crohn’s disease. Concerns developed about Dr. Wakefield’s claims when a French researcher noted that the antibody Dr. Wakefield used to identify measles virus reacted with all tissues, not just inflamed bowel tissue. A Japanese research group used PCR techniques on tissue samples from Crohn’s disease patients, and was also unable to detect the virus.  A Japanese research group also examined the second antibody Dr. Wakefield used to detect measles virus, and the researchers determined that it was reacting with a human protein.  These reports were followed by a number of articles, published from 1996-2000, demonstrating that the measles virus was not present in Crohn’s disease. In 1998, Dr. Wakefield appeared as the senior researcher on an article describing Dr. Nicholas Chadwick’s unsuccessful attempts to find the measles virus in the gut tissue of Crohn’s disease patients. [245 Hazlehurst Tr. at 632A-34A.] Why Dr. Wakefield appeared as an author on an article casting doubt on his claims of measles virus involvement in Crohn’s disease is unclear…
In 1998, Dr. Wakefield published a paper in Lancet describing 12 autistic children with symptoms of abdominal pain and food intolerance.  During colonoscopies on the children, Dr. Wakefield found lymphonodular hyperplasia [“LNH”] in their small intestines (a finding often referred to as ileal lymphonodular hyperplasia [“ILNH”]  and mild nonspecific colitis. Doctor MacDonald characterized this paper as “probably the worst paper that’s ever been published in the history of [Lancet].” Hazlehurst Tr. at 633A-34A. Ten of the paper’s 12 authors later filed a “Retraction of an Interpretation” of the paper…  [pp. 117-8]
Respondent’s witnesses were highly critical of Dr. Wakefield’s hypothesis, and of Dr. Wakefield personally. Doctors MacDonald and Fombonne were both part of a U.K. Medical Research Council investigation into Dr. Wakefield’s claims. Doctor MacDonald found evidence suggesting fraud. Doctor Rima examined some of the evidence Dr. Wakefield relied upon and found that it was not what Dr. Wakefield claimed it to be.
Doctors Ward and Griffin both had personal experiences with Dr. Wakefield that left them highly skeptical of his claims. [Bold emphasis added.][pp. 119-120]
Doctor MacDonald criticized the 1998 Wakefield paper for its lack of proper controls, but focused much of his criticism on the misleading statements it contained. Doctor Wakefield reported that the children had food intolerance and abdominal pain. However, several of Dr. Wakefield’s co-authors,  who were the physicians caring for the children, reported in a March 21, 1998, letter to Lancet, that the children suffered from severe constipation. Hazlehurst Tr. at 637A-38. The significance of this additional information is that lymphonodular hyperplasia is often caused by constipation. Hazlehurst Tr. at 628A. Doctor MacDonald characterized Dr. Wakefield’s theory as “incredible,” “fantastic,” “improbable,” and “not based on any data.” Hazlehurst Tr. at 642A-43A.
Doctor MacDonald was even more critical of the Wakefield 2002 paper.  [Bold emphasis added.] At the Hazlehurst hearing, he characterized the Wakefield 2002 paper as scientific deception. Hazlehurst Tr. at 646A-47A. He noted that the rates of inflammation documented in the pathology reports for the colon were similar for the autistic children and the control children. Although more of the autistic children had pathological findings in the ileum, Dr. MacDonald criticized the pathology reports for inventing new conditions and characterizing normal lymphoid follicles as pathologic abnormalities. Doctor MacDonald also pointed out that the presence of LNH is neither evidence of inflammatory bowel disease nor inflammation.  …
During his testimony in Hazlehurst, Dr. MacDonald used four endoscopy photographs that appeared in Dr. Wakefield’s 2002 paper as examples. Although the four photographs purportedly show ileum, Dr. MacDonald explained that the time stamps that appear on them indicated that Panel A was cecum, not ileum. He characterized this as “something of a deception.”…
With 15 years of experience in measles virology, Dr. Rima was very interested in Dr. Wakefield’s first reports of an association of the MMR vaccine with gastrointestinal disease. He met Dr. Wakefield at a conference in 1992, where a number of measles virologists looked at the material Dr. Wakefield had produced. Doctor Rima attended two meetings, and concluded that the material produced in support of Dr. Wakefield’s claims was highly selective, and that Dr. Wakefield was not responsive to criticisms of his findings. The cellular material that Dr. Wakefield claimed was measles virus was not measles virus. Snyder Tr. at 843A-46A.
In 1995, one of Dr. Wakefield’s students approached Dr. Rima about coauthoring a paper. After examining the data supplied by the student, Dr. Rima concluded that the findings of measles virus were based on contamination from a measles virus clone he had previously supplied to Dr. Wakefield as a positive control for his research. When an abstract concerning positive results for the presence of measles virus was not retracted after Dr. Rima informed them of the contamination, Dr. Rima formally withdrew from his collaboration with Dr. Wakefield…
According to Dr. MacDonald, the MRC investigation concluded that Dr. Wakefield was using reagents to identify measles virus that were not specific for that virus, and that important control measures were not being used. As a result, the Royal Free Hospital asked Dr. Wakefield to repeat the studies. No repeated study results have been published. Hazlehurst Tr. at 629A-632A. [pp. 120-122] [Bold emphasis added.]
When Dr. Wakefield first began to implicate MMR as a cause of autism, he invited Dr. Griffin to the U.K. as a consultant, presumably based on her expertise with measles virology. Cedillo Tr. at 2832A. She spoke with people from his laboratory at an open scientific meeting where they indicated they were having problems getting their PCR testing to work. Cedillo Tr. at 2861A-62. It was quickly apparent to Dr. Griffin that Dr. Wakefield’s laboratory personnel did not know how to perform PCR testing and analysis. Based on her personal interactions with Dr. Wakefield, she was suspicious of the research he did, and she declined the consultation offer. Cedillo Tr. at 2832A-33.
Doctor Ward testified that Dr. Wakefield presented data from an abstract of work done by Dr. Ward’s laboratory as supportive of Dr. Wakefield’s MMR-autism hypothesis. Doctor Ward personally cautioned Dr. Wakefield against relying on this data because what was presented initially in the abstract turned out to be wrong.  Cedillo Tr. at 1864A-65. [pp123-4] [Bold emphasis added.] [pp. 120-4.] [Bold emphasis.]
The testing was performed by Unigenetics, an uncertified laboratory…
Respondent mounted an overwhelming challenge to the reliability of Unigenetics’ test results for measles virus. [pg. 159-60.]
The Unigenetics laboratory had several of the hallmarks of unreliability noted in Daubert. It was established, primarily, if not solely, for the purpose of supporting the claimants in the U.K. MMR litigation.  [Unigenetics] results were not reproducible by independent investigators, and its quality control problems were so pervasive that they suggested gross negligence, if not outright scientific fraud.…
[A]n analysis of the public evidence alone clearly demonstrates that the results from Unigenetics cannot be relied upon as evidence of the persistence of measles virus in children with autism. When considering the U.K. litigation information, the evidence that Unigenetics’ results are not reliable is overwhelming. Unigenetics’ operations reflect unsound applications of the sound scientific process of PCR testing… [pp. 168-9.] [Bold emphasis added.]
Between 2000-2006, Dr. Afzal published a series of papers involving PCR testing of the Wakefield hypotheses of measles virus causation of both IBD and autistic enterocolitis.  The 2000 article  began with a summary of previous attempts to identify measles virus in biopsies from patients with IBD by various histological means, noting issues concerning the specificity of reagents and the difficulty in distinguishing virus structure from normal cellular structures. The study itself involved testing PBMCs and biopsies of inflamed bowel tissue from Crohn’s disease patients, using primers specific for three different measles genes (N, M, and H). No measles RNA was found in any of the samples. This result was consistent with reports from other researchers, using different PCR methodologies, all of whom had failed to detect measles RNA. The report also refuted a number of hypotheses put forth by Dr. Wakefield’s group from the Royal Free Hospital to explain why other researchers could not detect measles virus.…
In a 2002 publication, Dr. Afzal summarized the evidence for and against the IBD/autistic enterocolitis and measles virus hypotheses.376 He proposed an initiative to supply measles virus samples to a number of laboratories, including Unigenetics, to determine whether the laboratories could reliably detect measles virus in tissue. Unigenetics declined to participate. [pp. 176-7] [Bold emphasis added.]
Prior to Dr. Wakefield’s association with Dr. Kawashima, Dr. Chadwick (who was then working on his Ph.D thesis with Dr. Wakefield as his primary supervisor) had attempted to detect the presence of measles virus in PBMCs, but was unable to do so. He used PCR to test PBMCs of children with autism for the presence of both the H and N gene, but did not find the virus. He was surprised that Dr. Kawashima was able to detect the presence of the H gene in PBMCs. Cedillo Res. Ex. QQ at 2-3.
During the course of Dr. Wakefield’s association with Dr. Kawashima, Dr. Chadwick sent samples to Dr. Kawashima for testing. On one occasion, Dr. Kawashima reported positive results for measles virus in samples that Dr. Chadwick had previously tested with negative results. Doctor Chadwick performed gene sequencing on the positive samples, and discovered the sequence was identical to that of the positive control SSPE samples he previously provided to Dr. Kawashima. The patient samples were contaminated with material from the positive control. [pg. 178] [Bold emphasis added.]
Dr. Oldstone was approached about a collaboration with Dr. O’Leary, but, before he would agree to collaborate, he wanted to ascertain the O’Leary laboratory’s ability to detect reliably measles virus. He therefore prepared tissue and cell cultures at various levels of infection, and sent blinded samples to the O’Leary laboratory. After the laboratory tested them, Drs. Oldstone and O’Leary unblinded the specimens and determined that the laboratory results were only about 80% accurate, including about 10% false positive results. Snyder Tr. at 952A-56A. According to Dr. Ward, this result would be unacceptable in a research setting, and was wildly inappropriate for a diagnostic lab, such as Unigenetics. Cedillo Tr. at 1846; Snyder Tr. at 956A.
Doctors Oldstone and O’Leary agreed to try again, and Dr. Oldstone prepared another set of samples. After testing, the results were jointly unblinded, with a similar level of inaccuracy of around 20%. Some of the samples that were false positives or false negatives in the first round of testing were resubmitted in the second round with new code numbers. In several instances, samples that tested as positive during the first round tested negative in the second, and vice versa. At that point, Dr. Oldstone decided against any further collaboration, and suggested publishing the results. However, the study’s sponsor would not grant permission for publication. [pg. 180] [Bold emphasis added.]
Doctor Griffin summed up the criticisms when she testified that the reputation of the O’Leary laboratory in the scientific community was “not very good.” Cedillo Tr. at 2866.
The evidence that Unigenetics was not reliably detecting measles virus is highly probative. The lack of specificity in the F gene primers and the resultant amplification of DNA from sources other than the measles virus, the discordant results for H and N genes, the failure to sequence the results from the F gene positives, and the inability of other laboratories to reproduce its results all cast considerable doubt on the reliability of Unigenetics’ reports. Therefore, based on evidence not derived from the U.K. litigation materials, I conclude that Unigenetics’ reports cannot be considered at face value and are entitled to little evidentiary weight, absent some indication that a specific test result is corroborated by evidence from sources other than Unigenetics. [pg. 181] [Bold emphasis added.]
The evidence from the U.K. MMR litigation established that Unigenetics experienced all of the problems commonly encountered in PCR testing, and that it failed to take adequate measures to detect and correct them. In addition to the problems in the primers used to select the amplification target addressed by Dr. Ward, other problems with the primers were discovered. Unigenetics had problems in RNA extraction and sample quality, poor internal controls and poor adherence by laboratory personnel to Unigenetics’ own SOP, unblinded testing that permitted data manipulation, and a critical problem with contamination. Some of the data discovered during Dr. Bustin’s review suggested a fraudulent manipulation of laboratory notebooks and machine settings. Other evidence generated during the U.K. litigation demonstrated anew that Unigenetics’ results could not be reproduced…
Doctor Bustin assessed the Unigenetics practice with regard to quality and quantity assessment of RNA in samples as inconsistent. He found very little quality assessment was performed. Even when quality assessment was done, it was often done improperly, resulting in the testing of samples that should have been discarded. Data from the Uhlmann paper indicated that, in approximately one-third of the samples tested, the RNA quality was unacceptable for PCR. Cedillo Tr. at 1996-97…
Additionally, the conversion of RNA to cDNA had hallmarks of “operator error,” as the conversion step was sometimes skipped, rendering any results useless. During Dr. Bustin’s review of the Unigenetics laboratory data, he began to think that the positive laboratory results for measles virus genes were the result of contamination, rather than the actual presence of the virus in the unknown samples. His slides, Cedillo Res. Tr. Ex. 13 at 8-9, reflected Unigenetics laboratory data that demonstrated the reverse transcription step had not been performed. The results on page 9 of this exhibit indicated that in four of the lead cases in the U.K. litigation, the laboratory failed to perform reverse transcription. The positive results could not have come from the patient samples because any RNA present in them was not converted to DNA. Because PCR cannot amplify RNA, whatever was amplified had to be a contaminant introduced in the laboratory. Cedillo Tr. at 1980A-82. …
Approximately one-third of Unigenetics’ runs had positive results for the negative controls, a result indicative of a significant problem with contamination. In at least one case, an environmental control (a tube left open to the air in the laboratory) tested positive, a strong indicator that the laboratory itself was contaminated. Cedillo Tr. at 1995-96.
The concerns Dr. Bustin developed about contamination at Unigenetics were heightened when he visited the laboratory and discovered that the PCR portion of the laboratory was next to a room labeled “Plasmid Room.” Plasmids are DNA molecules used to replicate large quantities of DNA in a bacterium. These bacteria offer massive potential for DNA contamination, and standard scientific practice would be to place the plasmid facility as far from the PCR laboratory as possible. Plasmids get onto hair, hands, and clothes, and are easily transferred throughout the facility, and extremely difficult to eradicate. Although Dr. Sheils assured Dr. Bustin that they did not use the plasmid room for growing F gene targets, he remained concerned it was the source of the contamination he already had identified in the assays. Cedillo Tr. at 2021-23.[pg. 182-4] [Bold emphasis added.]
Unigenetics frequently reported positive results from samples that should have been read as negative. Cedillo Tr. at 2005-07. Unigenetics’ practice on reporting discordant results from the same sample was decidedly unusual. It is standard practice to test samples at least twice. If the results are genuine, they will be the same. If there are discrepancies, the test should be repeated…
The results of one repeated assay are displayed in the slide appearing at page 16 of Cedillo Res. Tr. Ex. 13. The open bars reflect the assay run on March 21, 2003, and the solid bars reflect the repeated assay done on March 26, 2003. Doctor Bustin noted the massive variability in the cycle numbers obtained on the different dates for the same samples. He pointed out that some of the samples tested positive on the first run and negative on the second, and vice versa. Unigenetics reported the samples that tested positive on either day as positives, ignoring the discordant results. Cedillo Tr. at 2014.
Doctor Bustin also testified that an ABI 7700 machine used by Unigenetics was defective…The machine that was used on most of Unigenetics’ runs had huge variations in the heating and cooling across the block. Depending on where the tube was placed, the temperature fluctuations caused variability in the results because both reverse transcription and PCR are sensitive to temperature. Unigenetics was unaware of the problem with the ABI 7700 machines. Cedillo Tr. at 1998-2000. [pp. 186-7.] [Bold emphasis added.]
[Evidence of fraud in laboratory books.] Laboratory notebooks are used to record what is done in an experiment, and are later used to write papers on the experiment. Laboratory notebooks are generally available for inspection. In standard practice, notebook entries are usually made relatively contemporaneously and are not, thereafter, changed…
The later addition of this parenthetical suggests something less benign than such an error. This was only one of several examples of alterations Dr. Bustin found in laboratory notebooks…
Unigenetics submitted testing data for the U.K. litigants to the court with each case in the litigation assigned a unique number. When documents such as the page appearing on page 22 of Cedillo Res. Tr. Ex. 13 were submitted as evidence, Unigenetics personnel would mark the results relevant to a particular litigant by boxing them in and writing the litigant number next to the result. In this particular exhibit, the results relevant to wells C1 and C2 were boxed in, with a handwritten “# 8″ inside the box, indicating that these two wells pertained to Patient Number 8 in the litigation.389 The handwritten notation “viral cells 1/100″ was also written by Unigenetics personnel, reflecting that these were positive controls–cells known to be infected with measles virus at a 1/100 level of infection. The entry in column three for these two wells is “Positc,” an abbreviation for “positive control.” Because Dr. Bustin received this document from evidence relating to Patient Number 8 in the U.K. litigation, he interpreted the entries as an attempt to report the results from a positive control as a litigant’s result. Cedillo Tr. at 2031-32. In essence, Unigenetics took a control sample known to contain measles virus and reported it as a litigant’s sample, the PCR testing equivalent of “planted evidence.”
The bottom slide on Cedillo Res. Tr. Ex. 13 at 14 reflected another problem with Unigenetics’s reporting. The baseline on slide B reflected a range of 2-13, rather than the 3-15 range recommended by the manufacturer. The effect of using the altered range was to change the results from positive to negative. This is significant, because the sample in question was a negative control. If it tested positive, suggesting contamination, it would raise doubts as to the other results from the same run. Using the manufacturer’s settings, as Slide A of this exhibit demonstrated, the negative control sample should have returned a positive result. Cedillo Tr. at 2008-10A.
Although Dr. Bustin stopped short of saying the alteration in the settings was a deliberate manipulation of the data, he testified that the unusual settings were a matter of curiosity for him, until he obtained access to the raw data, which showed this particular sample was a negative control. The clear implication from his testimony was that the raw data provided the motive to manipulate the machine settings, as a positive result for a negative control reflects a contamination problem that should cause all of the samples from that run to be discarded. [pp. 187-9.] [Bold emphasis added.]
[Analysis of the Evidence Regarding MMR Causation of Autism] [T]he problems with the case presented by petitioners for general causation are overwhelming. The quality of the petitioners’ experts paled in comparison to the world-class experts proffered by respondent. The theories petitioners’ experts advanced lacked support in both logic and research. As Dr. Ward testified, an hypothesis has a life span. An hypothesis may be biologically plausible at the time it is first advanced. As evidence accumulates, the hypothesis may be strengthened or weakened. The MMR hypothesis may have appeared biologically plausible at its inception, but the accumulating body of scientific evidence has tipped the scales decisively against it. Snyder Tr. at 975. The weight of the scientific evidence is that the measles vaccine virus plays no role in the pathogenesis or triggering of autism. I thus conclude that petitioners have failed to demonstrate that the MMR vaccine can cause autism, even in the highly circumscribed subset of children with regressive ASD and gastrointestinal symptoms.
Aside from the flaws evident in the theories of causation advanced, there are
other reasons for concluding that MMR vaccine causation of autism is improbable. [pg.196] [Bold emphasis added.]
[Measles Virus Has Not Been Reliably Detected in Children with ASD.] The only evidence that truly distinguished children with “regressive autistic enterocolitis” from other children was the finding of measles virus in gut or CSF. Was this evidence sufficiently reliable to suggest that the virus was causal of ASD and/or gastrointestinal disorders? I conclude that it was not.
The bedrock of the scientific process is reproducibility.  If a result or effect is real, it can be repeated. If a different result obtains from efforts to duplicate it, the experiment must, at a minimum, be repeated. Cedillo Tr. at 2014-15A. Unigenetics’ results could not be duplicated by any other researchers…Unless measles virus can be detected in greater numbers of children with ASD than in typically developing children, the preliminary findings are largely irrelevant in establishing a causal connection between measles genomic material and gastrointestinal complaints, much less a connection with ASD. The mere presence of measles virus in gut tissue would be extremely weak circumstantial evidence of MMR causation of autism, particularly in view of the paucity of evidence that “autistic enterocolitis” is a separate phenotype of autism. The presence of measles virus in CSF, if reliable, would present a different, and far stronger, case for causation, even if petitioners could not demonstrate such a finding more frequently in children with ASD than in their typically developing peers. Unfortunately, the evidence for the presence of measles virus in CSF is not reliable, because of the flaws in Unigenetics’ testing…
Doctor Rust used the term “scientific fraud” in describing the information upon which the MMR theory of causation is based. While noting that scientists are very careful about using that term, he testified that there was “abundant evidence” of scientific fraud in the body of evidence developed to support the MMR-autism hypothesis. Hazlehurst Tr. at 506A-07A. Sadly, the petitioners in this litigation have been the victims of bad science, conducted to support litigation rather than to advance medical and scientific understanding of ASD.
The evidence in support of petitioners’ causal theory is weak, contradictory, and unpersuasive. This is particularly apparent when considering the impressive body of epidemiologic evidence contradicting their theories.
The “logical connection” between MMR vaccination and onset of regression was also undercut by a considerable body of evidence showing onset of regression can be triggered by gene expression. Children with Rett’s disorder, which is entirely genetic in nature, evince loss of skills at specific times in their development, without triggering events. Other genetic conditions manifest when a particular gene is expressed; for example, Huntington’s chorea manifests decades after conception, without a triggering event. Given the complex genetic basis for ASD and the epidemiologic investigations into regression, gene expression is a more likely explanation than the MMR vaccine for the manifestation of regressive ASD symptoms.
Doctor Fombonne summed up the body of scientific research into ASD’s causes and the petitioners’ TCVs-MMR vaccine hypothesis, saying the possibility that some children are genetically predisposed to abnormal reactions to TCVs and the MMR vaccine so as to cause autism was less likely than the possibility of the earth being the center of the solar system. Cedillo Tr. at 1486A-88. His statement is an exaggeration of the evidence (or lack thereof), but is a concise and pithy expression of the general scientific disapproval of petitioners’ theories. [pp. 207-9.] [Bold emphasis added.]
Special Master Vowell reports Dr Rima’s testimony that he would have been delighted if Unigenetics had been capable of producing the results that they claimed.
Unigenetics’ rate of error was unacceptable. Doctor Rima’s response to one of my questions provided a cogent reason for so concluding. He noted that Unigenetics was the only laboratory commercially testing CSF or blood for the presence of measles virus. Snyder Tr. at 927A. He then commented:
Let me explain this. I mean, if the technology had been validated, then Dr. O’Leary would have found me and Oldstone and several other people interested in measles virus at his door saying, can you help us resolve issues about not only this disease. I can give you other diseases where there is a question about the formation of measles virus in – – disease, in otoclerosis. And I’m involved in several of these instances where people are struggling to find a link or an etiology for a disease which has no known etiology. And so, if indeed that technology had been validated, if that indeed had been the circumstance, a lot of people would have knocked on O’Leary’s lab and said you can do something which we can’t do. And there would have been a flood of people coming to him independent of the litigation of some. But that flood hasn’t taken place for the very simple reason that everyone who has looked at it said, no, actually, this technology does not work. What he claims he can do he cannot do. What he claims, he simply has not been able to give us the sort of confidence in his technology that would allow us to start looking at it from a research perspective. That’s a research perspective. That is a very different perspective even from the perspective of a diagnostic lab that is going to test children for pathological conditions that there are. So I would have said I would have been the first at his door…But it was clear that the company that was set up by Unigenetics had only one trading activity and that was to test measles presence in samples from the litigants in the U.K…We came quickly to the conclusion that some of the practices that I described here, some of the sloppiness, some of the inconsistencies in the data were there and they led us to the conclusion that this simply does not work.
Snyder Tr. at 927A-29.
 Panksepp J. A neurochemical theory of autism. Trends Neurosci 1979; 2: 174-77.
 Wakefield AJ, Murch SH, Anthony A, Linnell J, Casson DM, Malik M, Berelowitz M, Dhillon AP, Thomson MA, Harvey P, Valentine A, Davies SE, Walker-Smith JA. Ileal-lymphoid-nodular hyperplasia, non-specific colitis, and pervasive developmental disorder in children. Lancet. 1998 Feb 28;351(9103):637-41.
 Reichelt KL, Hole K, Hamberger A, Saelid G, Edminson PD, Braestrup CB, Lingjaerde O, Ledaal P, Orbeck H. Biologically active peptide-containing fractions in schizophrenia and childhood autism. Adv Biochem Psychopharmacol. 1981;28:627-43.
 Shattock P, Kennedy A, Rowell F, Berney TP. Role of neuropeptides in autism and their relationships with classical neurotransmitters. Brain Dysfunction 1991; 3: 328-45. [as cited in ]