Retrovirology Publishes Five Papers on XMRV and Contamination

[oceanblue]

If XMRV does not work out I really think more positive will have been done than negative. It really put CFS on the map once again....

Also, however this pans out, the XMRV story has been an amazing ride. It's brought in a lot of very good scientists who've worked very intensely to resolve this and scrutinised each others' work meticulously - and that's just how science should be. So often in this field in the past there's an interesting finding and even the originating researcher doesn't follow it up, let alone other researchers. I do hope that if XMRV turns out to be a dead end, the work we've seen will raise the bar for ME/CFS research.

 

[eric_s]

If XMRV does not work out I really think more positive will have been done than negative. It really put CFS on the map once again....

Bear in mind that all we have gotten those far is the contamination side of the story; we haven't the WPI's reaction or Dr. Singhs or Dr. Alter's reaction. The pMLV studies really are separate from XMRV - they are not necessarily effected by Hue's paper. (He didn't test them)...

I still wonder about XMRV integration into human DNA....even if it was a contaminant - if its integrated into human DNA - its still a virus that has infected a human cell....it's still there....isn't it? Was XMRV integration into human cells proved in the Science paper?

Yes, that's quite "funny". Now all the people who said pMLVs are different from XMRV and for that reason denied that Lo et al. confirmed Lombardi et al. are not able to say that Hue et al. proves that pMLVs are contamination too. Or am i wrong?

And i don't think we should talk like "it's all over" yet. Not at all. We've only heard one side. For me XMRV in ME/CFS is not over until the WPI, Alter/Lo and Hanson say so. Or i have reason to believe they are not telling the truth.

 

[roma]

ME/CFS not caused by XMRV according to researchers from London.

http://www.cbc.ca/health/story/2010/12/20/chronic-fatigue-xmrv-contamination.html#socialcomments

I don't even know what to say anymore, i can't believe there are pseudo scientists out there that still claim is nothing but contamination.

 

[Cort]

Hue explicitly states he did not test the pMLV's in his analysis.....One of the papers - I haven't really read - the McClure paper does suggest that the mtDNA test he used might not be sufficient to test for mouse DNA, though.....that the Intracisternal A Particle (IAP) test is better at that - which obviously could be a problem...

Not a great day for anyone.....

I think the lines with regard to that are drawn now... neither the WPI or Alter did the IAP test; they both have their samples and presumably could retest them...

There is the WPI, Ruscetti, Singh, Alter, Lo - anybody else? (Cheney?, ESME, Hansen) to hear from on all of this..

These papers are really dense...ithey are not easy to understand; its too bad we never got a retrovirologist outside the scene to be our guide to this...

 

[Kate_UK]

We have to fund WPI research (and other good researchers). No-one else will do it for us.

 

[omerbasket]

I really hope that at the end of all that, the journal "Retrovirology" will not be published anymore. They really took a stand against XMRV, and I believe that they are wrong. And anyway, I don't think I'll ever believe any thing negative about XMRV unless it will come from the WPI or from Drs. Alter and Lo. There are people here who are desperate to make XMRV disappear.

Anyway, this thread is already 7 pages long and is continuing to grow. That's not good. If we would not comment about it so much, it will be closed to forgotten very soon, I think.

 

[Jemal]

I don't want to sound too depressing, but if XMRV is proven a contaminant, I think it will do more damage to our cause than good. You would have two retrovirus incidents in the history of this disease, the second time with very reputable scientists involved. I am not so sure many scientists would still want to get involved with us. ME/CFS might be seen as too difficult and unfortunately it's in our nature to take the paths of least resistance. The third time some scientist claims he has found the cause of ME/CFS, he will come under intense scrutiny and his reputation will be in jeopardy. It would probably be easier to just study the mating rituals of hummingbirds.

Still believing in XMRV though and I hope we'll see a response soon from the scientists who think XMRV is for real.

Anyway, this thread is already 7 pages long and is continuing to grow. That's not good. If we would not comment about it so much, it will be closed to forgotten very soon, I think.

Ignoring it won't make it dissappear. More and more news outlets are picking up this story. British, Spanish and now German news outlets have picked up the story. I am sure US news outlets will soon follow.

Hopes of breakthrough dashed as four papers conclude virus originating in mice is not the cause of chronic fatigue syndrome

http://www.guardian.co.uk/society/2010/dec/20/chronic-fatigue-me-research

XMRV definitiv nicht Ursache des Chronischen Mdigkeitssyndroms

http://www.aerzteblatt.de/nachricht...rsache-des-Chronischen-M_digkeitssyndroms.htm

Powerful statements like these tend to stick in people's minds.

Other people on the "other side" of the XMRV debate have also quickly picked up the news and are gloating. Our good friend ERV for example.

 

[oceanblue]

Interesting discussion section from the Robinson paper (which found the mitochondrial mouse DNA test wasn't that sensitive). They point out that if contamination is the cause of the positive, it's unclear where the contamination arises from, though it's probably not from the reagents (unlike the finding from the oakes paper, which showed that the PCR kit they used was contaminated). And they do also politely acknowledge that just because their XMRV positives were all associated with mouse DNA contamination, it doesn't prove that XMRV positives in other cohorts are down to contamination.

Mouse DNA contamination in human tissue tested for XMRV

Discussion

XMRV shares extensive sequence identity with known xenotropic, nonecotropic and
polytropic murine viruses; the first of which is known to infect many common human
tumour cell lines, a phenomenon that has confused retrovirologists looking for disease
associations for over three decades. Most putative associations of new or old human
retroviruses with diseases (including CFS and prostate cancer) have turned out to be
laboratory artefacts [19]. The case of XMRV as a new human pathogen must be judged
against this background [22]. It is true that we cannot formally rule out the possibility
that the samples in question are infected with XMRV and simultaneously contaminated
by mouse DNA, although this is unlikely since we found no IAP-negative samples from
which we amplified MLV-specific sequences (data not shown). Also, the failure to detect
XMRV sequences other than in association with mouse DNA contamination in our cohort
does not mean that the virus is not present in other, unrelated, cohorts.
It is difficult to explain how the contamination may have occurred, especially since the
samples came from three unrelated centres in the UK, Korea and Thailand. As both our
negative tissue and PBS controls treated in parallel with the FFPE were XMRV PCRnegative,
it is unlikely that contamination was introduced via reagents. The UK FFPE
tissue samples were stored boxed and stacked in a cupboard in the histopathology
department for several years; and it is possible that contamination happened during that
time, although why only a few samples (4.8%) were XMRV positive and the remainder
not is difficult to explain. Nor does it explain the Thai and Korean results on tissue
collected prospectively for the study. It does, however, demonstrate the necessity of
controlling by highly specific and sensitive means for mouse contamination.

 

[eric_s]

Don't let this get you down. The last word on XMRV is totatlly not spoken yet. No reason to think you've lost just because the opposite side says so. They have talked enough bs before, we know they're capable of this.

I think if we can pay them we will always have people willing to work on CFS. And there are not only retroviruses to look at.

 

[Jemal]

Going to listen to this song for the rest of the night
http://www.youtube.com/watch?v=OBwS66EBUcY

Where have all the good men gone
and where are all the gods?
Where's the streetwise Hercules to fight the rising odds?
Isn't there a white knight upon a fiery steed?
Late at night I toss
and I turn
and I dream of what I need.
I need a hero. I'm holding out for a hero 'til the end of the night.
He's gotta be strong
and he's gotta be fast

(or replace "he" with a "she")

Also, nice edit omerbasket. Now that sentence is really grabbing attention

 

[KnightofZERO]

I can't comment on the papers--haven't read them, just skimmed the editorial. I do think it is sicking... I don't know which more-- that people would be happy about research findings because such findings could set back the cause for treatment and a cure for suffering patients (I mean: happy about the findings for exactly that reason) or that there *are* such people who are happy. It is sickening.

 

[Cort]

Hue Paper

My take on the Hue paper - a very complex paper - reader beware; this is a laymen's take, of course, on a complicated issue.

WPI Demonstrates XMRV is Not a Contaminant - Demonstrating that XMRV was not likely to be a contaminant was an important of the Science paper. While the WPI was not able to conclusively prove this they provided enough convincing evidence for Science to publish the paper. This following is taken direct from the paper.

Sequences of full-length XMRV genomes from two CFS patients and a partial genome from a third patient were generated (table S1). CFS XMRV strains 1106 and 1178 each differed by six nucleotides (nt) from the reference prostate cancer strain XMRV VP2 (EF185282), and with the exception of one nt, the variant nucleotides mapped to different locations within the XMRV genome, suggesting independent infections. By comparison, prostate cancer-derived XMRV strains VP35 and VP42 differed from VP62 by 13 and 10 nt, respectively. Thus, the complete XMRV genomes in CFS patients are > 99% identical in sequence to those detected in patients with prostate cancer.

To exclude the possibility that we were detecting a murine leukemia virus (MLV) laboratory contaminant, we determined the phylogenetic relationship between endogenous (nonecotropic) MLV sequences, XMRV sequences, and sequences from CFS patients 1104, 1106 and 1178 (fig. S2). XMRV sequences from the CFS patients clustered with the XMRV sequences from prostate cancer cases and formed a distinct branch from nonecotropic MLVs common in cases of inbred mouse strains. Thus, the virus detected in the CFS patients blood samples is unlikely to be a contaminant.

Hue Proposes XMRV is a Contaminant - Now Hue has taken another, more comprehensive look at that question. He appears to have done much the same thing (?) but using more mouse strains.

Hue cloned and sequenced the gag and env genes from the prostate cancer cell line that produced the representative copy of XMRV. (This is the copy the WPI searched for in their initial paper.) Then he compared how closely those genes to those found in endogenous MLV genomes, some other XMRV clones, other MLV genomes and 2 samples from the WPI study.

The phylogenetic analysis suggested that, in contrast to the WPIs findings, that everything, the XMRV from prostate cancer, the CFS patients, the endogenous MLV retrovirus, etc. grouped together; ie. they all appeared highly related to each other. Once XMRV passed into a human host it should developed its own group of viruses that cluster together; that is, in fact what the WPIs analysis indicated. But Hues larger analysis did not - it suggested that have been no outbreaks, genetically speaking, of XMRV; instead of an XMRV branch and an endogenous MLV branch, etc. there was one big cluster.

Statistic Derivation of XMRV's origin - They then ran a Baynesian tree analysis that is designed to statistically determine which part of a group of elements is probably the origin of that group. Every one of 3000 passes of that tree suggested that the entire group was derived from a cell line; ie they are all derived from a laboratory construct.

Hue looked for XMRVs distinctive genetic element - a deletion in the gag sequence - in 12 inbred mouse strains. The fact that he founded it in all of them suggested, I believe that they ALL carried XMRV - a surprise in itself. He also found this specific XMRV element in four endogenous retroviruses. Interestingly, it wasn't easy to find; it was only present in a very low-frequency - which is a little ominous because, as we know, XMRV is not easy to find.

Primers - When they looked at the primers used to screen for XMRV and he found that primers sets as he described as XMRV specific can really amplify MLV sequences that is, if you were to use those primers to search for XMRV in mice in which it is not found - they would tell you that XMRV was there. He seems to be stating that the specific gag deletion researchers are looking for does not necessarily pick up XMRV.

 

[Jemal]

Thanks Cort.

I think this is also a nice take on the recent studies:
http://neuroskeptic.blogspot.com/2010/12/xmrv-innocent-on-all-counts.html

 

[Ronan]

Has anyone actually been cured since all the xmrv stuff started by taking Anti Retrovirals? I know Daffodil has been sharing her experiences with her trials but i dont know of anyone else who has been trying them. If there are people getting well on anti retrovirals it would put alot of this contamination talk to bed in my mind. Also, 3 HIV medications have been shown in vitro to suppress xmrv. Could this be possible if its only a contaminate?

 

[Sushi]

Has anyone actually been cured since all the xmrv stuff started by taking Anti Retrovirals? I know Daffodil has been sharing her experiences with her trials but i dont know of anyone else who has been trying them. If there are people getting well on anti retrovirals it would put alot of this contamination talk to bed in my mind.

Hi Ronan,

If you haven't already been reading it, you might want to check-out Dr. Jamie Deckoff-Jones's blog: http://treatingxmrv.blogspot.com/2010_12_01_archive.html

She and her daughter were disabled by ME/CFS and have both improved a great deal taking a combo of anti-retrovirals. Enough that she is planning to go back into practice on a part-time basis.

She certainly doesn't claim "a cure" but her blog details both her own and her daughter's steady improvement.

Sushi

 

[Cort]

Here's my take on the Robinson study - much shorter and easier to understand and not nearly as comprehensive. This study suggested IAP assays pick up mouseDNA better than the mtDNA tests do.

Mouse Contamination DNA in Human Tissue Tested for XMRV - Robinson, Erlwein, etc.

This study looked at prostate cancer specimens embedding paraffin from the UK, Thailand and Korea. They extracted the DNA and then used primers to test for the XMRV gag sequences that encompass the special deletion that reportedly differentiates XMRV from other MLV’s. They then ran mtDNA and intracisternal A particle (IAP) LTR’s.

Of the 437 samples 21 were XMRV positive. When they sequenced out what they had found they found that the primer had picked up some sequences that did not have the XMRV specific deletion ie; it appeared to pick up something other than XMRV. When they dug deeper they found it was picking up an endogenous retrovirus found in a common laboratory mouse strain. (My understanding is that because the WPI sequences their positive tests - they should be picking these up).

Contamination Tests - they then ran both mtDNA and IAP assays in two cell lines and found that the IAP assay was 100 times more sensitive that mtDNA at picking up genomic mouse DNA. Then they applied both tests to their samples and found that the IAP and mtDNA tests indicated that 26% and 18% of samples were positive for mouseDNA suggesting that the IAP test, if it is more accurate, is about 50% more effective at finding the mouse contamination. ALL the putatively positive XMRV prostate cancer cells were positive for mouseDNA by the IAP test.

XMRV or Comtamination or Both? - They weren’t able to show the XMRV was not there - it and mouseDNA both could conceivably have been present; what they showed was that all the samples that had tested positive for XMRV also tested positive for mouseDNA (ie contamination) using the IAP test.

 

[Cort]

Hi Ronan,

If you haven't already been reading it, you might want to check-out Dr. Jamie Deckoff-Jones's blog: http://treatingxmrv.blogspot.com/2010_12_01_archive.html

She and her daughter were disabled by ME/CFS and have both improved a great deal taking a combo of anti-retrovirals. Enough that she is planning to go back into practice on a part-time basis.

She certainly doesn't claim "a cure" but her blog details both her own and her daughter's steady improvement.

Sushi

That's just another piece of the puzzle isn't it? How do you explain her and her daughter's improvement and Andrea Whittemore's improvement (although we don't know of her treatment regime)...if XMRV doesn't work out? Is there another retrovirus or is it something else?

 

[Cort]

Here is Neuroskeptic's blog

http://neuroskeptic.blogspot.com/2010/12/xmrv-innocent-on-all-counts.html

XMRV - Innocent on All Counts?
A bombshell has just gone off in the continuing debate over XMRV, the virus that may or may not cause chronic fatigue syndrome. Actually, 4 bombshells.

A set of papers out today in Retrovirology (1,2,3,4) claim that many previous studies claiming to have found the virus haven't actually been detecting XMRV at all.

Here's the rub. XMRV is a retrovirus, a class of bugs that includes HIV. Retroviruses are composed of RNA, but they can insert themselves into the genetic material of host cells as DNA. This is how they reproduce: once their DNA is part of the host cell's chromosomes, that cell is ends up making more copies of the virus.

But there are lots of retroviruses out there, and there used to be yet others that are now extinct. So bits of retroviral DNA are scattered throughout the genome of animals. These are called endogenonous retro-viruses (ERVs).

XMRV is extremely similar to certain ERVs found in the DNA of mice. And mice are the most popular laboratory mammals in the world. So you can see the potential problem: laboratories all over the world are full of mice, but mouse DNA might show up as "XMRV" DNA on PCR tests.

Wary virologists take precautions against this by checking specifically for mouse DNA. But most mouse-contamination tests are targeted at mouse mitochondrial DNA (mtDNA). In theory, a test for mouse mtDNA is all you need, because mtDNA is found in all mouse cells. In theory.

Now the four papers (or are they the Four Horsemen?) argue, in a nutshell, that mouse DNA shows up as "XMRV" on most of the popular tests that have been used in the past, that mouse contamination is very common - even some of the test kits are affected! - and that tests for mouse mtDNA are not good enough to detect the problem.

Hue et al say that "Taqman PCR primers previously described as XMRV-specific can amplify common murine ERV sequences from mouse suggesting that mouse DNA can contaminate patient samples and confound specific XMRV detection." They go on to show that some human samples previously reported as infected with XMRV, are actually infected with a hybrid of XMRV and a mouse ERV which we know can't infect humans.

Sato et al report that PCR testing kits from Invitrogen, a leading biotech company, are contaminated with mouse genes including an ERV almost identical to XMRV, and that this shows up as a false positive using commonly used PCR primers "specific to XMRV".

Oakes et al say that in 112 CFS patients and 36 healthy control, they detected "XMRV" in some samples but all of these samples were likely contaminated with mouse DNA because "all samples that tested positive for XMRV and/or MLV DNA were also positive for the highly abundant IAP long terminal repeat [found only in mice] and most were positive for murine mitochondrial cytochrome oxidase sequences [found only in mice]"

Robinson et al agree with Oakes et al: they found "XMRV" in some human samples, in this case prostate cancer cells, but they then found that all of the "infected" samples were contaminated with mouse DNA. They recommend that in future, samples should be tested for mouse genes such as the IAP long terminal repeat or cytochrome oxidase, and that researchers should not rely on tests for mouse mtDNA.

They're all open-access so everyone can take a peek. For another overview see this summary published alongside them in Retrovirology.

I lack the technical knowledge to evaluate these claims, no doubt plenty of people will be rushing to do that before long. But there are a couple of things to bear in mind.

Firstly, these papers cast doubt on tests using PCR to detect XMRV DNA. However, they don't have anything to say about studies which have looked for antibodies against XMRV in human blood, at least not directly. There haven't been many of these, but the paper which started the whole story, Lombardi et al (2009), did look for, and found, anti-XMRV immunity, and also used various other methods to support the idea that XMRV is present in humans. So this isn't an "instant knock-out" of the XMRV theory, although it's certainly a serious blow.

Secondly, if the 'mouse theory' is true, it has serious implications for the idea that XMRV causes chronic fatigue syndrome and also for the older idea that it's linked to prostate cancer. But it still leaves a mystery: why were the samples from CFS or prostate cancer patients more likely to be contaminated with mouse DNA than the samples from healthy controls?

 

[pictureofhealth]

The BBC interviewed Professor Tim Peto for the BBC Report on the contamination studies published in Retrovirology Journal today
http://www.bbc.co.uk/news/health-12041687

Prof Tim Peto told the BBC: It now seems really very, very unlikely that XMRV is linked to chronic fatigue syndrome

Professor Tim Peto, consultant in infectious diseases at the University of Oxford, said the original paper in Science came as a great surprise to experts.

What great timing for a high profile piece by the BBC to have one of the PACE Trial Management team (Professor Tim Peto) announcing that "it now seems very very unlikely that XMRV is associated with Chronic Fatigue Syndrome".

Especially as the PACE Trials are due to be published some time after Christmas - how jolly good for White Sharpe and Chalder to get publicity for scuppering ideas of an infectious cause for ME.

Prof Tim Peto was involved in the CBT/GET PACE Trials as a Centre Leader Leader (the local PACE Research team) at the John Radcliffe Hospital Oxford

Professor Tim Peto is on the [PACE] Trial Management Group (TMG)

http://www.biomedcentral.com/1471-2377/7/6

Trial Management Group (TMG)

The Trial Management Group (TMG) will be responsible for the day-to-day running and management of the trial. It is composed of:

The three principal investigators

1. Professor PD White, Professor of Psychological Medicine, Centre for Psychiatry, Wolfson Institute of Preventive Medicine, Bart's and the London, Queen Mary School of Medicine and Dentistry, Department of Psychological Medicine, St Bartholomew's Hospital, London, EC1A 7BE.

2. Professor MC Sharpe, Professor of Psychological Medicine and Symptoms Research, School of Molecular and Clinical Medicine, Symptoms Research Group, Royal Edinburgh Hospital, Edinburgh, EH10 5HF.

3. Professor T Chalder, Professor of Cognitive Behavioural Psychotherapy Academic Department of Psychological Medicine, Guy's, King's and St Thomas' School of Medicine, Weston Education Centre, Cutcombe Road, London SE5 9RJ

All centre leaders and co-leaders
1. Dr D Wilks
2. Professor S Wessely
3. Dr M Murphy
4. Dr BJ Angus
5. Professor T Peto
6. Dr E Feldman
7. Dr G Murphy
8. Hazel O'Dowd
.

Thanks for the heads up and the reminder Wild Cat.

 

[oceanblue]

My take on the Hue paper - a very complex paper - reader beware; this is a laymen's take, of course, on a complicated issue.

That was quick! And very helpful, thanks Cort.