Switzer - Publishes new XMRV varieties, and challenges Coffin's recombination theory

[Bob]

Which would only be relevant to the Lombardi study and all the negative studies if they had looked for virus DNA in the patients blood. But they didn't. That's my point. Also when Switzer talks about copy number he means cells infected with virus DNA, NOT virus particles, XMRV is a retrovirus (RNA virus) you can't test for virus particles with nested PCR (DNA). This paper basically says that the virus is present but it's not doing anything (latent).

I think my opening post is very clear and speaks for itself.

If you don't understand the possible significance, then I can't really explain it any clearer than I already have.

Obviously, you will draw your own conclusions from the paper.

I don't understand why you are focusing on the phrase 'low copy numbers'. I didn't use that phrase in my opening post, and I have explained my use of that term in subsequent posts.

To clarify, Switzer used RT-PCR to test for XMRV in the blood and failed to detect it: "Plasma from all three persons was negative for viral RNA by RT-PCR."
I believe that this is significant as it demonstrates how little XMRV presence or activity there is in the blood of XMRV infected individuals.

 

[RedRuth]

I haven't really drawn any conclusions beyond the phylogenetics being some convincing evidence that XMRV may not be contamination. I apologise if I sound patronising but you don't seem very clear on what the different techniques are and what they test for.

 

[Bob]

This paper basically says that the virus is present but it's not doing anything (latent).

This opens up a whole new area of interesting discussion.

Switzer concludes that XMRV is a latent (chronic) human retrovirus infection with extremelly low copy numbers in both blood and tissue.

This conclusion challenges the zero/zero studies, and it leads to many many important questions.

Personally, I think that his conclusion that there is no association with prostate cancer is premature based on this study.

 

[Mark]

Switzer concludes that XMRV is a latent (chronic) human retrovirus infection with extremelly low copy numbers in both blood and tissue.

Note that this conclusion applies to the 3 prostate cancer patients in which he detected it. The degree of viremia in the blood may perhaps be extremely low or non-existent in prostate cancer patients but slightly more detectable in ME/CFS patients and even more detectable in the carefully-selected cohort tested by the WPI.

Perhaps XMRV is latent in prostate cancer but sequence fragments (at least) may be present in the blood of ME/CFS patients who are viremic...and the persistent and relapsing/remitting nature of ME/CFS seems to me suggestive of the possibility that XMRV may be most detectable when the patient is relapsing.

I agree with Bob's notes in the first post regarding the significance of Switzer's findings. Specifically...

1. Switzer confirms that XMRV is a real wild human virus.

2. Switzer concludes that the genetic variation is consistent with normal human infection.

4. Switzer confirms that his findings not due to contamination.

All these conclusions are contrary to the claims of the 'contamination theorist' camp and so they do indeed run contrary to the 'case against' XMRV as a factor in ME/CFS.

3. Switzer admits that he cannot detect XMRV in the blood of XMRV-positive prostate cancer patients, using established methodologies. This has relevance for all of the other negative XMRV studies.

Redruth's clarification of the scientific detail here has been illuminating for many I'm sure, and Bob's extensive analysis of the wider body of XMRV research is also logical and valid IMO - so I'd personally be interested to see this discussion continue.

Could we hear more about what the phylogenetics tell us? They seem to be a crucial part of Switzer's findings, and he seems to be saying some very interesting things about complex recombination of sequences, suggesting that sections of these viruses may have been recombining for decades, and, most interestingly of all, that he has identified multiple sequences which could overlap to produce XMRV in a similar way to the sequences that Coffin found in his contamination argument.

This scenario of recombination and (what I would understand as) convergent evolution in this population is something that has been proposed on this forum for over a year, particularly in relation to the relationship between Lo/Alter's "PMLV" sequences and the WPI's "XMRV".

The way I think of this is that if XMRV shows a very high level of sequence conservation, as appears to be the case, then this might suggest that it is the most evolutionarily successful sequence of a family, appearing many times and being very successful when it does so...I also find myself thinking of it as the "hub" of a "hub and spoke" family of viruses.

I appreciate that those ideas are a little vague, and I'm not trained in any of these areas of science, but I'd be very interested in Redruth's thoughts on these ideas - I'm sure you could explain the significance of Switzer's phylogenetic arguments in a way that would be of great interest to many of us here (despite your modesty, it turns out that your writing isn't that bad, as these things go! )

 

[Mark]

PS: Regarding my comment about 'fragments' in the blood, I was also thinking about a very interesting comment in one of the papers at the NIH State of the Knowledge conference, where a surprising finding that generated quite a bit of interest was that fragmentary sequences of certain proteins (?) were found in the blood of ME/CFS patients...my recollection is that the suggestion was that after cells broke up during cell apoptosis these fragmentary chains were left floating around in the blood, and were perhaps capable of interfering with neurological processes...it's a very vague recollection I'm afraid but I wonder if it could have some significance regarding exactly what is being detected in the blood of ME/CFS patients?...

 

[RedRuth]

The degree of viremia in the blood may perhaps be extremely low or non-existent in prostate cancer patients but slightly more detectable in ME/CFS patients

Which is another reason to be cautious about comparing the two, apart from the technical issues. I also think that you have to be cautious about the results of any paper, the most you can say is that this paper lends support ot the theory that XMRV is an infective virus that isn't just present in tissue culture cells. I don't think anyone is denying that it's a 'real virus'.

A brief explanation of the phylogentics is that when the retrovirus is part of the host cell genome (this is when it's a provirus) it replicates via the host cell machinery and doesn't pick up many mutations. When the virus infects a cell and copies itself into DNA it uses it's own enzyme (reverse transcriptase) that is far more error prone, therefore the virus picks up mutations. This is one reason why it's so difficult to make vaccines against retroviruses.

So, very little variation in the sequence of a retrovirus would be evidence of it being a cell culture contaminant and variation is evidence of an infective virus. Also, the mutations are used to construct phylogenetic trees which show the relationship between viruses (or any other species). Specifically, nested hierarchies (shared differences) show which viruses are ancestral or related. This study found a reasonable amount of variation between the sequences and the variation falls into nested hierarchies with other sequences. However, this is only 3 sequences so you have to be careful not to overinterpret the results.

 

[Mark]

I don't think anyone is denying that it's a 'real virus'.

Nobody has quite said that, but the contamination argument of Coffin et al argues that (all?) purported detection of XMRV has been of contamination from cell lines, and not detection of something that was actually in humans. That's why any suggestion that XMRV is a virus that is actually infecting humans, and not just infecting cell lines, is significant.

With that context in mind I think your conclusion in your first paragraph above is basically agreeing with what Bob has been saying here, just in more cautious language. It does seem quite significant to us in the context of the wider picture, but I wouldn't disagree with the cautious way you've expressed it - I guess this tentative support is just a little more exciting to us given the wider political context - especially the fact that this research comes out of the CDC!

Thanks for the explanation of the phylogenetics...I'd like to hear more about the details of what Switzer found in this respect. You said earlier that they were "some convincing evidence that XMRV may not be contamination" - and against a background where one side of this argument seems to be emphatically claiming that the case for contamination is proven and the book should now be closed, that's what makes the research seem so significant.

Do we have the percentages of variation here? I guess what I'm curious about boils down to some pretty raw maths at the end of the day: the sequences themselves and the phylogenetics of their relationships are the key really?

Also, it may not be in this paper, but I'm sure Switzer has also suggested that he has found multiple MLV sequences which could combine to produce XMRV in the same way as Coffin's "pre-XMRV1 and pre-XMRV2" sequences did. I think he said he found groups of 3 sequences which could do so? That seemed to be the most significant bit of Switzer's findings, to me (again partly because it confirmed what some posters here predicted)...so it might be worth digging into those details to see if there are indeed other ways to make XMRV...

 

[Bob]

I apologise if I sound patronising but you don't seem very clear on what the different techniques are and what they test for.

RedRuth, please don't accuse me of being unclear about techniques when I haven't actually mentioned any, and I haven't made any comparisons.
Any confusion seems to be your own.
The only technique I have mentioned in this thread is RT-PCR, which Switzer used, which I sloppily referred to as 'PCR' in one post. (Sorry for the confusion, but you knew that I meant RT-PCR.)
I think if you read any of my blogs on the forum, you'll realise that I have an understanding of the basics (at least) of retroviruses.

I haven't made any direct comparisons between Switzer's study and other studies.
I simply said that Switzer's study was significant, and relevent to other negative studies, for a number of different reasons, and clearly explained why.

I think your approach to the studies is different to mine.
You seem to be looking for specific conclusions that Switzer makes in his studies.
Whereas I am looking at the study in a slightly wider sense, as an ME patient, and drawing my own logical conclusions that are relevent for the ME patient community, from the results of the study.

As an ME patient, I am interested in whether XMRV is a human virus, and whether all of the negative studies were adequate enough to detect XMRV in the blood.
The Switzer study demonstrates how difficult XMRV is to detect in the blood (looking for both antibodies and plasma viremia), and so I am suggesting that all XMRV activity in the blood may be at very low levels. For this reason only, I am suggesting that, whatever technique is used to detect XMRV in the blood, it might not be adequate. This is a perfectly valid issue to raise, and requires further investigation.

Also, Switzer specifically compares the behaviour of XMRV in the blood in his study to the behaviour of the virus (and simian retroviruses) in macaque studies, where the virus and antibodies quickly disappear from the blood after infection. He refers to SRV macaque studies and an XMRV macaque study where they looked for provirus in PBMCs rather than viremia in plasma. This is a direct comparison to other studies that Switzer himself makes (i.e. other techniques may be inadequate/inappropriate to detect XMRV in the blood due to low copy numbers), and so adds further reason for Switzer's study to have relevance for other negative XMRV studies.

"Loss of antibody during a latent infection, while atypical of most retroviral infections, has been described previously for natural infection of macaques with simian type D retroviruses (SRV) [27]. SRV in macaques is associated with outbreaks of severe immune deficiency in primate colonies and latent SRV infection in these antibody negative animals is confirmed with greater sensitivity using PCR analysis [27]. Our results are also consistent with those seen recently in macaques experimentally infected with XMRV in which tissues at necropsy are PCR-positive but viremia and detection of provirus in PBMCs disappear quickly, followed by loss of antibody detection [28], [29]. "



Switzer also demonstrates how difficult XMRV is to detect in tissue.
Another question which the study raises is: were Switzer's techniques adequate to detect XMRV in most of the prostate cancer samples?
He struggled to detect XMRV in 3 samples, in very low copy numbers. Were there more positive samples but with even lower copy numbers?
Maybe he will develop his techniques.

 

[RedRuth]

I think you have to remeber that if you have 2 scientsts you'll get 3 opinions (to paraphrase), you also have to bear in mind that science is all about replication and conscilience, it's extremely foolhardy to draw firm conclusions from one paper. So a few scientists will be convinced XMRV is a cell culture derived virus that gives false positives in blood samples (because of reagent contamination), a few scientists will be convinced that XMRV is an infective virus present in the blood of CFS patients. Most scientists will be somehwere in between until the evidence has stacked up on one side of the debate. These kinds of arguments are par for the course in science, it's just that they're not usually of any great interest to the wider public.

As for the paper in the OP, I think its only relevance to the XMRV/CFS argument is the phylogenetics data which is fine (but it's not really my field) but limited. He does elsewhere I think outline a possible scenario for the evolution of XMRV from other Mouse retroviruses but I haven't read the paper so I can't say. It also has a bearing on the Paprotka paper which I have read and which I thought had some convincing evidence that XMRV is a cell culture infection. I think it will be interesting to see what each of them has to say about the other's paper. But it's far too soon to say that Switzer has confirmed anything about the emergence of XMRV.

 

[RedRuth]

I simply said that Switzer's study was significant, and relevent to other negative studies, for a number of different reasons, and clearly explained why.

And I disagree and I explained why. He didn't do the one thing that one of the biggest arguments is over - testing for virus DNA in blood. His comments about not finding protein or RNA are just that, comments about why there's no viremia. I can't remember now, did Lomabardi et al do reverse trancriptase PCR? I know they did some western blotting.

I realise (and sympathise) that you're a CFS sufferer and therefore see this research in this light but I think you are overinterpreting the results.

 

[eric_s]

Ruth, i don't think it's necessary to use the term "CFS" here. The US DHHS's advisory committe proposed using the term ME/CFS and the US NIH are also using this one. It's how this illness is called in the Canadian Consensus Criteria, which are the most up to date and probably best definition. Most people here would certainly appreciate you using another term than "CFS". I think your input is interesting and this could prevent some unnecessary tensions. If you're from the UK, you might also just use the term that is widely used there and has been around for much longer than "CFS", which is "ME".

 

[markmc20001]

I haven't really drawn any conclusions beyond the phylogenetics being some convincing evidence that XMRV may not be contamination. I apologise if I sound patronising but you don't seem very clear on what the different techniques are and what they test for.

Yes, you do sound patronizing Redruth.

I realise (and sympathise) that you're a CFS sufferer and therefore see this research in this light but I think you are overinterpreting the results.

It comes across as speculation to me when you claim to know what "light" Bob is interpreting these papers through.

I happen to agree with Bob's point of view here and he hasn't made any leaps of logic in my opinion.

It is safe to say everybody on these forums sees things in their own light.

 

[RedRuth]

Ruth, i don't think it's necessary to use the term "CFS" here. The US DHHS's advisory committe proposed using the term ME/CFS and the US NIH are also using this one. It's how this illness is called in the Canadian Consensus Criteria, which are the most up to date and probably best definition. Most people here would certainly appreciate you using another term than "CFS". I think your input is interesting and this could prevent some unnecessary tensions. If you're from the UK, you might also just use the term that is widely used there and has been around for much longer than "CFS", which is "ME".

Point taken, I didn't know why the term had changed from ME to CFS, I didn't realise it had unfortunate connotations. ME, it is.

 

[RedRuth]

I'm just reiterating what he said himself Whereas I am looking at the study in a slightly wider sense, as an ME patient, and drawing my own logical conclusions that are relevent for the ME patient community, from the results of the study.

 

[Bob]

I think you have to remeber that if you have 2 scientsts you'll get 3 opinions (to paraphrase), you also have to bear in mind that science is all about replication and conscilience, it's extremely foolhardy to draw firm conclusions from one paper.

I realise (and sympathise) that you're a CFS sufferer and therefore see this research in this light but I think you are overinterpreting the results.

Yes, I agree that definite conclusions should not be made from this one study. (Please tell that to the numerous researchers who cannot detect XMRV and then, after carrying out a single study, declare that XMRV is a contaminant, or that it isn't associated with ME.) And this is why I did not make any conclusions in my opening post. They were all Switzer's conclusions, taken from his paper. And I think if you re-read my opening post and read Switzer's study, you'll see that I didn't do any interpreting at all, let alone over-interpreting. The comment that I made, regarding the relevance to other negative XMRV research studies, is based on Switzer's own discussion and interpretations.

With regards to that comment (re relevance to other research), Switzer himself discusses this issue, where he compares his research to the research in macaques, where viral DNA and antibodies disappear from the blood quickly after infection. This is Switzer's discussion and interpretation, not mine. And of course his discussion and paper has relevance to other negative blood studies. How can it possibly not? If viral DNA and RNA and Antibodies disappear from the blood, then they won't be easily detectable in the blood. I have not done any over-interpreting here.

I have considered all of the other research when making the comments that I have made, including the macaque studies that Switzer references.
I have not considered this paper in isolation.

If you would like to see a more comprehensive discussion of some of these issues, then please see my recent blog entry:
http://phoenixrising.me/forums/entry.php?959-XMRV-The-case-against-contamination

 

[Mark]

As for the paper in the OP, I think its only relevance to the XMRV/CFS argument is the phylogenetics data which is fine (but it's not really my field) but limited.

I must say I'm personally unconvinced by your rebuttal of Bob's points, Redruth. In the context of the wider arguments about XMRV/CFS and contamination, the detection of XMRV as a real human infection (not a laboratory contaminant) and undetectable (by him) in the blood when nevertheless present as a latent infection, has to have relevance.

Scientifically, you've pointed out well the appropriate caution to be taken towards these findings, and been more precise about specifically what was and was not tested for and found...but Bob has been following the whole of the political and scientific arguments about XMRV very closely over the last year or two, and in the context of that bigger picture, these results do seem quite significant to me too...though that's of course a subjective term, "quite significant", so perhaps it's a bit daft to argue about it...

He does elsewhere I think outline a possible scenario for the evolution of XMRV from other Mouse retroviruses but I haven't read the paper so I can't say. It also has a bearing on the Paprotka paper which I have read and which I thought had some convincing evidence that XMRV is a cell culture infection.

IMO, that part of Switzer's research is the most significant, and if we've interpreted what he's saying correctly it's big news. If he's even suggesting that there may be other ways for XMRV to evolve, and sequences that demonstrate that possibility, and evidence of recombination in this family of viruses, then that is a real blow to Coffin and Paprotka's argument. So I really hope you (or somebody else here) will read that paper and report back - because it really does seem crucial: on the face of it, it has the potential to completely undermine Paprotka and Coffin's contamination theory.

Regrettably, I don't often have the time these days to dig into the details of these papers, so I just keep up with the bigger picture...and like Bob, that tells me that, here, Switzer (specifically because he's from the CDC) finding evidence of real human XMRV infection is significant.

I think it will be interesting to see what each of them has to say about the other's paper.

It most certainly will!

But it's far too soon to say that Switzer has confirmed anything about the emergence of XMRV.

Well now, who said that he had? Are you over-interpreting Bob's comments, perhaps? This seems a bit like a straw-man comment to me.

 

[Mark]

I realise (and sympathise) that you're a CFS sufferer and therefore see this research in this light but I think you are overinterpreting the results.

RedRuth, this comment, and one or two others, has annoyed some members. Perhaps you don't understand why, but perhaps I can illustrate. Perhaps you would be annoyed if someone were to say:

"I realise that you're not an ME/CFS patient, have only tangential interest in XMRV research, have no particular reason to care about us, and are not familiar with the history and politics surrounding ME/CFS, and that this lack of motivation and background knowledge limits your ability to grasp the context and significance of these results."

That would be an insulting and personal way to express things, but similar in attitude to what you wrote. I really don't think it will be positive if we engage on that level on this thread, so everyone: please try to stick to the science.

We all have different perspectives, and if this conversation is going to be worthwhile and illuminating, we need to respect each other's backgrounds. RedRuth is a researcher, with relevant technical knowledge which many of us here lack: we could all benefit from that perspective. Bob (like many forum members) doesn't just have the perspective of a patient, he has also been following and studying XMRV science with a strong focus for quite a long time now, and he has followed the wider world of ME/CFS science and politics for longer still, and that broad and relevant knowledge also gives him an important wider perspective which the majority of researchers lack.

So please, everyone, try to respect each others' knowledge and expertise. Of course we have different perspectives that colour our thinking and may skew our interpretation - even scientists may perhaps make assumptions and assume too much trust in their colleagues' objectivity, neglecting the political factors - but we're all intelligent adults here, so we ought to be able to have a decent conversation and learn from each other respectfully.

 

[Bob]

Mark, that perfectly explains the perspective of an ME patient. Thanks for that.

To be fair to RedRuth, I did say that I was approaching the science from the 'wider' perspective of an ME patient.

I can see that it wasn't a helpful comment.

 

[eric_s]

Point taken, I didn't know why the term had changed from ME to CFS, I didn't realise it had unfortunate connotations. ME, it is.

Thank you. It's a long and complicated story and i don't know all about it either. The thing is that it used to be called ME for decades and then, in the 1980s in the USA, after the Lake Tahoe outbreak, the US CDC introduced the term CFS, as far as i know. The problem is that this term is misleading, since it omits the fact that people with ME/CFS have many other symptoms in addition to fatigue and also fatigue is not really an accurate description of what we experience. It leads to people feeling this illness is not serious, when in reality it has a very big impact on patient's lifes. Also it's the favourite term used by those groups who believe CFS should be treated with a psychological approach.

 

[RedRuth]

Mark, that perfectly explains the perspective of an ME patient. Thanks for that.

To be fair to RedRuth, I did say that I was approaching the science from the 'wider' perspective of an ME patient.

I can see that it wasn't a helpful comment.

There was nothing wrong with your comment! I understood exactly what you meant, that your interest in the research is its relevance to ME/XMRV. At least that's how I took it and that's what I meant. I think whoever has got upset by it has entirely misunderstood my intent. I think a more accurate analogy would be ............I'm not a CFS/ME patient or retrovirologist but I am a cell biologist and see the research in this light I'm only cautioning against overinterpeting results, which is something we're all guilty of.