Retrovirology Publishes Five Papers on XMRV and Contamination

[Countrygirl]

http://7thspace.com/headlines/36741...of_xmrv_like_sequences___.html?forumid=331851


http://7thspace.com/headlines/36740...an_tissue_tested_for_xmrv.html?forumid=331851

Two new articles, one including the names of Coffin and mcClure and the other, Coffin and Huber, which have found mouse DNA contamination in XMRV samples.

Can our scientific folk here evaluate the papers, please?

Thanks.

C.G.

 

[pictureofhealth]

I don't think its fair to keep publishing negative and contamination papers and not publish the WPI's UK study and others positive papers.

Still at least Myra is keeping up the studies and not just squawking from the sidelines.

Also this does help to explain Coffin's position (and perhaps, as a result, Stoye's 'contamination stance at the latest BWG meeting too - although he is not a co author).

 

[Countrygirl]

Third paper today - XMRV consistent with mouse contamination

http://www.retrovirology.com/content/7/1/111

This is not turning out to be a good day for XMRV research. This is the third paper/article claiming XMRV is possibly/probably the result of mouse contamination.


C.G.

 

[Jemal]

During the BWG meeting Coffin also announced he had 2 papers in the pipeline that would be published this month. This does look like quite a blow to XMRV research...

Alter and the WPI do seem pretty sure XMRV was not caused by contamination though.

 

[Jemal]

I read on the Niceguidelines blog that 5 papers about contamination were published on the same day !?

http://niceguidelines.blogspot.com/2010/12/publishing-5-papers-on-same-day.html

 

[Sam Carter]

Links for cited papers:

Contamination of clinical specimens with MLV-encoding nucleic
acids: implications for XMRV and other candidate human retroviruses

Robert A Smith

Retrovirology 2010, 7:112 (20 December 2010)

http://www.retrovirology.com/content/pdf/1742-4690-7-112.pdf


Disease-associated XMRV sequences are consistent with laboratory contamination

Stphane Hu, Eleanor R. Gray, Astrid Gall, Aris Katzourakis, Choon Ping Tan, Charlotte
J. Houldcroft, Stuart McLaren, Deenan Pillay, Andrew Futreal, Jeremy A. Garson1, Oliver G. Pybus, Paul Kellam, Greg J. Towers

Retrovirology 2010, 7:111 (20 December 2010)

http://www.retrovirology.com/content/pdf/1742-4690-7-111.pdf


An Endogenous Murine Leukemia Viral Genome
Contaminant in a Commercial RT-PCR Kit is Amplified
Using Standard Primers for XMRV

Eiji Sato, Rika A Furuta , Takayuki Miyazawa

Retrovirology 2010, 7:110 (20 December 2010)

http://www.retrovirology.com/content/pdf/1742-4690-7-110.pdf


Contamination of human DNA samples with mouse DNA can lead to false
detection of XMRV-like sequences

Brendan Oakes, Albert K Tai, Oya Cingz, Madeleine H Henefield, Susan Levine, John M
Coffin and Brigitte T Huber

Retrovirology 2010, 7:109 (20 December 2010)

http://www.retrovirology.com/content/pdf/1742-4690-7-109.pdf


Mouse DNA contamination in human tissue tested for XMRV

Mark J Robinson, Otto W Erlwein, Steve Kaye, Jonathan Weber, Oya Cingoz,
Anup Patel, Marjorie M Walker, Wun-Jae Kim, Mongkol Uiprasertkul, John M
Coffin and Myra O McClure

Retrovirology 2010, 7:108 (20 December 2010)

http://www.retrovirology.com/content/pdf/1742-4690-7-108.pdf

 

[Esther12]

It would be really strange if all these different studies were finding similar levels of conatmination in healthy controls, and much more in prostate cancer and CFS patients. But I also feel that some of the pro-XMRV camp have been too casually dismissive of worries about contamination... stange things can happen in science labs that don't get fullyt reported in papers.

Has McClure now moved from being pretty sure XMRV is linked to prostate cancer to now thinking it's probably all the result of problems with contamination?

I was hoping the BWG would sort this out... but it seems like that's not their priority. I wonder if publication of these papers was delayed until the BWG released results? (They wouldn't want to do so prior to a big splash confirming the association).

 

[Esther12]

Would it be a good idea to have one of the mods combine the various threads on these?

 

[Jemal]

I wonder if publication of these papers was delayed until the BWG released results? (They wouldn't want to do so prior to a big splash confirming the association).

I don't think that's likely. Getting these kind of papers published is a process of months. Also Coffin said during the BWG meeting that he 2 of his papers would be published this month. I think he would rather have seen those papers published already, as that would have strengthened his cause?

 

[eric_s]

I'm not a scientist, but i'll give my take on this.

The Huber paper is, if i'm not mistaken, nothing new. This study was presented at the 1st Int'l XMRV Workshop, we knew her results for a long time. Since Coffin was a co-author there, this might explain his "cautious" stance.

In the "McClure paper" they are talking about prostate cancer. I think so far Myra McClure has hinted that she believes XMRV in prostate cancer is real, but XMRV in ME/CFS is not. So that would me she's contradicting herself.
And the paper finds that many of the "negatives" show evidence for contamination. So either contamination is not the reason why a sample is found to be positive or you need a certain degree/quality of contamination.

Either way, i don't have a problem with those papers. We just should not let ourselves be discouraged by them and the public should not draw any wrong conclusions, that's my only concern.
They are scientists and they publish, other scientists can read their papers and maybe learn something. The more information there is, the better.
For exmaple the "McClure paper" shows which assay seems to be best to check for that sort of contamination, so this can be used by the WPI, Alter/Lo and other teams to make their papers more "bullet-proof" (if they haven't already used that IAP assay). And if they have, this paper is a validation for them.

And none of this can explain why the positive studies find different percentages of positives in cases vs controls (unless they used different kits or something of that sort).

 

[WithHope]

This seems like really unsettling news at Xmas!

Does anyone know when Ila Singh's XMRV study will be released? And does anyone have any solid idea that it is indeed positive news - for those of us hoping XMRV is real?

Thanks for any update.

 

[alex3619]

Hi, these papers are talking about contamination of samples with mouse DNA (I have only read the abstracts at this point). Since mouse DNA contains endogenous MLVs, selective amplification of mouse DNA will amplify the retrovirus segments, and result in a false positive. However, I recall that Lo and Alter have stated on Dec 14 that they have tested for mouse DNA and not found it. I also know that it would be easy enough for the WPI and others to go back and retest all (or some of ) their positive samples for mouse DNA. This will end the speculation.

However this still doesn't explain why so many controls were not positive for MLVs as if they were handled the same then nearly all would be MLV positive - if the contamination theory is valid. If the controls were handled differently, we need to know that because it violates the concept of a control. If this is contamination, it is due to repeated failings, by multiple experts, at different labs, on multiple occasions, to hold to the expected standard of care with controls. I find this very unlikely at this point.

There is also this: if a lab contaminates samples with mouse DNA, test samples and controls will all be contaminated, and it may no longer be possible to reliably determine if XMRV was present in the first place. This does not mean it wasn't there, it only means that the tests need to be redone with fresh samples at a mouse free lab.

It is far more likely that the labs that did these test are contaminated, and therefore should not be doing any more XMRV experimentation. If I were a researcher who believed in contamination, I would be asking for samples from WPI etc for independent testing for mouse DNA, and I would want to do this at new labs that have never worked with mouse samples or live mice. I would also be taking fresh draws from CFS patients and testing their blood all over again at labs thought to be mouse clean.

A bigger risk, as has been pointed out by others, is that the cell line used to culture XMRV might be contaminated. However, that is one of the first things the WPI should have checked. Does anyone know if they have? Also if controls were also cultured, then the same arguments about controls arise as I have discussed for blood testing.

All of this speculation rests on this: endogenous MLV sequences can be confused with XMRV. Whereas some cases of PMLV and XMLV might be endogenous false positives, XMRV is not present in most murine genomes. It is unclear how it could be found falsely positive using MLV sequences.

Then there is this. Singh uses an antibody staining technique that clearly shows XMRV infected tissues. While it is possible that this is due to cross-reactivity (the stain picking up something else) I find it unlikely that this could be true here, and that XMRV could be mistakenly identified from MLV sequences, and that all the labs could have failed to properly process controls. This are too many coincidences: this doesn't prove it isn't the case, but on current evidence (these studies notwithstanding) I would still back XMRV over contamination theories.

Bye
Alex

 

[eric_s]

Here they show that some type of primers seem to also pick up contamination. I don't have a problem with that. It's the primers Coffin/Huber used. I don't know wheter the WPI, Alter/Lo, Hanson used those too. Would be interesting to know.
They say that XMRV doesn't acquire diversity in patients. Lo et al. demonstrated exactly the opposite, if i remember correctly...
They conclude that XMRV is not a human pathogen at all. That's pretty bold, i think.

I don't have a problem with those negative papers, the only thing i don't like is that they are all published simultaneously and that there seems to be reluctance to publish positive ones. Retrovirology seems to publish more or less anything, as long as it's negative. I find it very hard to understand that.
To be honest this smells rotten to me.

Either way, the others are continuing their work, so we don't have to care about those papers too much, i think. In the end they will help bringing out the truth, because they provide some new information, unless they lead to funds being cut or people turning away from research into XMRV. Those papers will probably make the authors of positive papers want to prove they were right, which will help us.

 

[Esther12]

There was that discussion between Coffin and Alter about different techniques for detecting contamination. I didn't really understand it well enough to know if it was really settled there or not.

The ambiguous results from the BWG and false positives that turn up mean that I'm still worried about the possibility that the XMRV link is entirely down to some bizarre contamination.

 

[eric_s]

Let's hope we get a reaction by the authors of positive studies to those papers. If they show that they have not used those kits, primers, etc. and have done the IAP test, it's one step further towards certainty that contamination is not the explanation for their results.

 

[pictureofhealth]

Thanks Eric and Alex! Aren't the Ruscetti's MLV experts already anyway? If so, surely they would know the difference between regular MLV's, contamination and a new finding.

Re contamination in ME/CFS patients v control samples - I think it is possible that the collection techniques are different. Patient samples may be 'live' and fresh or recently harvestd, whereas control samples may be held for many years in blood repositories like a blood bank, in tubes containing god knows what and under a variety of different protocols and temperatures - which would be available as control samples to any researcher studying blood samples for anything.

Perhaps to avoid confusion, both ME/CFS patient samples and healthy donor samples could be collected from live patients IN THE SAME WEEK ie over the space of several hours and stored and treated IDENTICALLY and PCR, culture and serology run an equal number of times for patient and healthies. Not impossible with careful and thoughtful planning.

 

[Dx Revision Watch]

Press Release: Wellcome Trust Sanger Institute

Wellcome Trust Sanger Institute.


For immediate release – Monday 20 December 2010

Chronic Fatigue Syndrome is not caused by XMRV
New research shows XMRV virus is a lab contaminant

A virus previously thought to be associated with chronic fatigue syndrome is not the cause of the disease, a detailed study has shown. The research shows that cell samples used in previous research were contaminated with the virus identified as XMRV and that XMRV is present in the mouse genome.

XMRV was first linked to chronic fatigue syndrome – also known as myalgic encephalomyelitis (ME) – in a study published in October 2009, where blood samples from chronic fatigue syndrome patients were found to have traces of the virus. XMRV had also been identified previously in samples from certain prostate cancer patients.

The new study, published in Retrovirology, identifies the source of XMRV in chronic fatigue syndrome samples as being cells or mouse DNA rather than infection by XMRV. The research does not rule out a virus cause of chronic fatigue syndrome - it is simply not this virus.

The research team developed improved methods to detect XMRV against the genetic noise of other sequences and make recommendations for future study of virus causes of human disease.

“Our conclusion is quite simple: XMRV is not the cause of chronic fatigue syndrome,” says Professor Greg Towers, a Wellcome Trust Senior Research Fellow at University College London (UCL). “All our evidence shows that the sequences from the virus genome in cell culture have contaminated human chronic fatigue syndrome and prostate cancer samples.

“It is vital to understand that we are not saying chronic fatigue syndrome does not have a virus cause – we cannot answer that yet – but we know it is not this virus causing it.”

The team, from University College London, Wellcome Trust Sanger Institute and University of Oxford, showed clearly that the experimental design of previous studies would pick up sequences that resembled XMRV; however, in this improved study, they could prove that the signal was from contamination by a laboratory cell line or mouse DNA. The sequences from the contaminated cell line and chronic fatigue patient samples were extremely similar, contrary to the pattern of evolution expected during the infectious spread of a virus in a human population.

They also showed that the existing methods would indicate that one in fifty human cell lines they examined were infected with XMRV-related viruses: they showed that contamination of human tumour cells with XMRV-related viruses is common and that a principal prostate cancer line used is contaminated.

“When we compare viral genomes, we see signs of their history, of how far they have travelled in space or time,” says Dr Stphane Hu, Post Doctoral Researcher at UCL. “We would expect the samples from patients from around the world, collected at different times, to be more diverse than the samples from within a cell line in a lab, where they are grown under standard conditions. During infection and transmission in people, our immune system would push XMRV into new genetic variants.

“Viral infection is a battle between the virus and the host and XMRV does not have the scars of a virus that transmits between people.”

Together the results demonstrate that XMRV does not cause chronic fatigue syndrome or prostate cancer in these cases. The team’s methods suggest ways to ensure that virus contamination does not confound the search for a cause of disease in future work.

The authors propose that more rigorous methods are used to prevent contamination of cell and DNA samples. They also suggest that consistent and considered standards are needed for identifying viruses and other organisms as cause of a disease.

“Increasingly, we are using DNA-based methods to accelerate our understanding of the role of pathogens in disease,” explains Professor Paul Kellam, Virus Genomics group leader from the Wellcome Trust Sanger Institute. “These will drive our understanding of infection, but we must ensure that we close the circle from identification to association and then causation.

The strongest lesson is that we must fully use robust guidelines and discriminatory methods to ascribe a cause to a disease.”

###ends###

Notes to Editors
Chronic fatigue syndrome
Chronic fatigue syndrome (CFS) is also known as myalgic encephalomyelitis (ME). CFS is characterised by long-term tiredness or fatigue that affects the everyday life of patients. There is no known cure for CFS.
For more information, visit: http://www.nhs.uk/Conditions/Chronic-fatigue-syndrome/Pages/Introduction.aspx

Publication Details
Hu S, Gray ER, Gall A et al. (2010) Disease-associated XMRV sequences are consistent with laboratory contamination. Retrovirology.

Participating Centres
MRC Centre for Medical Molecular Virology, Division of Infection and Immunity, University College London, London, United Kingdom
Wellcome Trust Sanger Institute, Hinxton, Cambridge, United Kingdom
Department of Zoology, University of Oxford, Oxford, United Kingdom

About UCL (University College London)
Founded in 1826, UCL was the first English university established after Oxford and Cambridge, the first to admit students regardless of race, class, religion or gender, and the first to provide systematic teaching of law, architecture and medicine. UCL is among the world's top universities, as reflected by performance in a range of international rankings and tables. Alumni include Marie Stopes, Jonathan Dimbleby, Lord Woolf, Alexander Graham Bell, and members of the band Coldplay. UCL currently has over 13,000 undergraduate and 9,000 postgraduate students. Its annual income is over 700 million.
www.ucl.ac.uk

The Wellcome Trust Sanger Institute, which receives the majority of its funding from the Wellcome Trust, was founded in 1992. The Institute is responsible for the completion of the sequence of approximately one-third of the human genome as well as genomes of model organisms and more than 90 pathogen genomes. In October 2006, new funding was awarded by the Wellcome Trust to exploit the wealth of genome data now available to answer important questions about health and disease.
http://www.sanger.ac.uk/

The Wellcome Trust is a global charitable foundation dedicated to achieving extraordinary improvements in human and animal health. We support the brightest minds in biomedical research and the medical humanities. Our breadth of support includes public engagement, education and the application of research to improve health. We are independent of both political and commercial interests.
http://www.wellcome.ac.uk/

Contact details
Don Powell
Press Officer
Wellcome Trust Sanger Institute
Hinxton, Cambridge, CB10 1SA, UK
Tel +44 (0)1223 496 928
Mobile +44 (0)7753 7753 97
Email press.officer@sanger.ac.uk

Craig Brierley
Senior Media Officer
Wellcome Trust
Tel +44 (0)20 7611 7329
Mobile +44 (0)7957 468218
Email c.brierley@wellcome.ac.uk

Ruth Howells
Media Relations Manager
UCL Corporate Communications
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Tel +44 (0)20 7679 9739
Mobile +44 (0)7990 675 947
Email ruth.howells@ucl.ac.uk

UCL out of hours contact: 07917 271 364

 

[aruschima]

http://www.retrovirology.com/content/7/1/111

We propose that XMRV might not be a genuine human pathogen.

Hi Eric,

How is Barcelona ? Must be freezing cold too?

Thanks for trying to calm us down, however, I think the statement above is reason to worry. Considering though from whom it is coming from not surprising, as you said.

However, I say it again; I think from what I gathered from the Spain news (by the way Eric, anything new there?) they proceed in a much smarter way. They getting the immune markers figured out; cannot contaminate those, can you?
Associate those with the retrovirus (more research needed), and you have a strong argument.

Also, I always said concerning advocacy and XMRV: use it as a tool, but do put all eggs into this basket.
Do not forget 3000 research studies up to this date, which all point to an acquired neuro-immune dysfunction.

As Alter said: Absolutly convinced when you define this by proper criteria, it's a very serious, medical disease. Chars of a viral disease. If it's NOT XMRV, we must continue the research to find out what is.

Aruschima

 

[eric_s]

I think it would be best if we just bought Retrovirology or another paper, then the "blockade" would be over
Just making fun at this point, but i think it would be possible, given how many we are.

The WPI will get it's chance in the BWG and Lipkin study and if they manage to prove they are able to reproduce their results there, their work should be published again. I don't see why it wouldn't. Ok, i also don't see why it isn't now...

 

[Esther12]

That Press Release is pretty forthright. Wow.

I wish this had come out prior to the BWG and it could have been discussed there.

I don't really know how they can be so confident, but that doesn't mean the must be wrong. I don't understand the WPI's confidence, and one of them must be right.