1. Patients launch $1.27 million crowdfunding campaign for ME/CFS gut microbiome study.
    Check out the website, Facebook and Twitter. Join in donate and spread the word!
Knitting Equals Pleasure, Despite ME/CFS
Jody Smith loves knitting. Again. She thought her days of knitting and purling were long over but ... she's back ...
Discuss the article on the Forums.

Another negative XMRV CFS study - is this new? looks to be from CDC

Discussion in 'XMRV Research and Replication Studies' started by Jemal, Feb 22, 2011.

  1. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    I think looking to the XMRV/MLV antibodies was a strength of the study, not a weakness. The more recently infected patients should have them for certain, and the older patients might as well. WPI says they find antibodies.

    But I think your second comment is right on target. The authors raised a point I have not seen before in print (might have been discussed at a conference though), and that is the possibility of human cell line contamination. Apparently this is known to be possible, it has happened (sorry, I definitely read/heard that somewhere but forget the source right now). No, that does not prove human infection in the patient population. Infected cell lines means that human cells grown for many generations as part of lab reagents (commercial or proprietary) have at some point been contaminated, and the contaminant continues to grow and stay in the HUMAN cells in the reagent. There are lots of human cell lines preserved and used in lab testing (we just don't hear about that much, so it sounds strange). Apparently WPI's and some of the other XMRV tests use human cell lines at some point in the assay. If some of those cell lines are contaminated, the testing for mouse DNA would be insufficient. They would have to screen every reagent used for contaminated human cells. Given that human cell line contamination is possible, this just seems prudent and I am surprised this issue has not come up before.
     
  2. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    I don't know many details of the antibody test, only that they acquired LIVE samples (I believe the source is mentioned at the end of the paper) and literally broke them in fragments and put them directly into the patient samples. They then looked for every XMRV protein. This is a very thorough test. If there was any antibody response to XMRV in their samples they would have seen it.
     
  3. omerbasket

    omerbasket Senior Member

    Messages:
    510
    Likes:
    13
    Your last sentence is totally a guess of yours, especially when we know that you have no training in biological sciences.
    Now, I wonder - why did they try so hard to get this "very thorough test" when we actually want them not to do this very thorough test, but the probably-not-very-thorough-test-in-your-opinion that was developed by the WPI and was proven to be able to find XMRV antibodies in clinical samples?

    Yes, we really needed another study that won't reproduce the "Science" study methods.
     
  4. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    No that is not a guess, that is what I was told when I spoke with one of the study authors. And since when is training in the biological sciences a prerequisite to post here? As for the motives of the study authors, I am convinced after many interactions that it is honest, to find XMRV if they can. And nobody has the right to tell these researchers what they can do, they all are interested in using their own tools and methods, including WPI. This study was an attempt to go far beyond WPI's level of resolution, they had more sensitive tests. And they found no evidence of any MuLV type infection.

    The Science 'methods' included using PCR without culturing and finding XMRV sequences in 67% of banked CFS samples. This study was equal to that method and goes well beyond.
     
  5. Esther12

    Esther12 Senior Member

    Messages:
    5,296
    Likes:
    5,606
    Hi Kurt.

    Do you know what this author thinks about prostate cancer? Could XMRV be circulating in the human population in a way that's undetectable by the methods they're using? Ta.
     
  6. alex3619

    alex3619 Senior Member

    Messages:
    7,495
    Likes:
    11,978
    Logan, Queensland, Australia
    Hi Kurt, I too am worried about human cell line contamination, but that was not my point. Non-murine sources include other animals - and possibly humans. We are everywhere, and it is very likely XMRV is in the human population, even if it is not associated with ME/CFS due to contamination. Hence, a likely source of contamination is lab staff. It is not enough to discredit this by saying XMRV is not in patients, they have to prove its not in the population too - and that is very much harder to do. But if staff are the source, how come they don't find it in the controls? Some of them should have it too. (I have not though much about the possibility that people carry mouse and other contamination around with them, because mouse contamination is looking less likely.)

    It is very much a case of XMRV not being a real virus as one option, but we have cloned it and used it to infect animals. If it were not real, it would not be in the population, it is all a mistake - background rates, prostate cancer, ME/CFS, all a mistake together.

    On the other hand, the mistake may lie in their testing, or something about the virus we do not understand.

    A position in between these two extremes is looking less and less likely.

    Bye
    Alex
     
  7. omerbasket

    omerbasket Senior Member

    Messages:
    510
    Likes:
    13
    Kurt, it is not a requirement to have a training in biological sciences in order to write here - but one can expect that a person who feel comfortable telling everyone that "If there was any antibody response to XMRV in their samples they would have seen it" would have a training in biological sciences at least, and would probably be god himself.
    You talked to one of the study authors? Wow, surprising that they said that. It's not like every other person who was involved in negative studies said that. So why does the NIH, FDA, NCI, BSRI and even the CDC continue making those tests, if so many studies that would have found XMRV if it were there are not finding it?

    That is a disgrace to them as people who call themselves "scientists". Reproducibility is an basic principle in science - and saying that a result is incorrect without trying to reproduce the methods that led to that result is 100% unscientific and requires a lot of arrogance - which these people surely have.

    At the first XMRV workshop Mindy Kitey said something like: "Why not, Mr. Switzer, instead of going to the RKI, who couldn't find it (XMRV) in prostate cancer, why not go to the Cleveland Clinc, who could find it (XMRV) in prostate cancer?". The arrogant Bill Switzer heared it, and decided to publish a study in which he cooperates with Cooperative Diagnostics - a company that couldn't find even one clinical sample until now, out of hundreds, to be positive for XMRV. Why not cooperate with an institute that found samples to be positive for XMRV, and just make sure to control well enough against contamination? Why did he use in his previous XMRV study the 20 positives from the WPI to test his serology method, didn't find any positive and decided not to publish it in his study? Why did he not check his PCR with those samples as well, to see if it's just as good as his serology test?

    Mr. Switzer is not a physician, so I guess he didn't say the hipocratic oath. But he is in a far more important position - as a governemt scientist he is the executioner - and while it may very well be that he did want to find XMRV had it been there (although it should be noted that the organization in which he came from, and also he himself as a researcher dealing with zoonotic problems - have every reason to dismiss the whole XMRV thing), he is doing a lot of obvious mistakes that are causing his hand to get closer and closer to pusing the stick down and send at least 17 million people to life of misery, or to death.

    Kurt, altough I don't like everything that is going on in the other ME/CFS-XMRV forum, reading your messages reminds me how anxious and angry one can get from reading the unbelievable, distructive things that some are allowing themselves to write here.
     
  8. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    To my knowledge, the authors do not believe they are missing XMRV due to detection problems of their methods. I think they were just as surprised as we all were to have negative results after the promising Science study article.

    Anyway, I just spoke with one of the authors on the phone and asked your question (he literally called me on other business while I was reading your post). He said they did not have anyone in their pool with prostate cancer. It is possible that someone with prostate cancer could have XMRV, even with their negative study, but he thinks that is unlikely. What it's coming down to is that there is either a group having false positives, or a group having false negatives. It is either present and significant or not present at all. Whatever happens in the CFS studies will probably be eventually reflected in the prostate cancer studies. We are hearing more about CFS right now because of all the publicity, but really what goes for CFS will likely go for prostate cancer regarding XMRV.
     
  9. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    I agree, a position between the extremes is looking less likely now. You make a good point that other animals could be sources of contamination, that is certainly possible, don't know if that has been talked about. As for whether XMRV is in the population, has anyone here discussed this study yet? (I don't spend a lot of time on the forum these days, am partly recovered now and working part-time again)

    I know that is a bit hard to wade through, but basically, this study found that XMRV is easily defeated by a cellular anti-viral mechanism, where viruses are over-stimulated to mutate until they are unviable to infect other cells. This makes the extreme possibilities even more striking (either it is present everywhere and being missed or a totally false finding). The reason this is striking for me is that the XMRV can spread but the dose required is enormous (10^6 virons if you read the whole article), and the virus will mutate so much that there is no way we would be seeing the identical gene sequences that the positive XMRV studies have shown.
     
  10. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    Sorry my posts make you anxious and angry. I am just sharing what I know about this study. It was a negative study, it was well constructed, and that's all there is to say really. I don't see all the drama in this situation you are finding, and am not looking at XMRV as a black and white final straw for CFS. I don't think failure of XMRV to be proven the cause for CFS will mean sentencing 17 million people to a life of misery or death. To the contrary, finding out the truth quickly is what we need. If the evidence is going against XMRV, that should be known sooner than later so other research directions can receive support. I fully expect those heavily invested in XMRV such as WPI to continue trying to prove their beliefs, but also expect that if they find they were wrong they will admit that quickly and move on. The same goes for the negative studies. If WPI or anyone can tell them how they can find XMRV, I would expect those labs to admit THEIR mistakes. This is science and not a courtroom where motives matter. A scientist's motives are almost irrelevant, what counts is their data. Besides that I think your analysis of motives is incorrect anyway from what I know of these people. We just have to go with the ups and downs of this evidence-driven process. If all we look at is the way we HOPE things go, then we are not in a scientific process, but something else. I think we should show some respect for every scientist seriously studying CFS as a biological illness, whether we personally like their findings or not.
     
  11. omerbasket

    omerbasket Senior Member

    Messages:
    510
    Likes:
    13
    Look, I stopped reading after you saying that you "just ahre what youy know about this study". Saying that "If there was any antibody response to XMRV in their samples they would have seen it" is not "saying what you know about the study", it is stating an opinion - a very arrogant opinion too, considering that it's a new virus that has no test that is agreed-upon, and the tests that were proven to be able to find the virus in clinical samples were not used.
     
  12. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    That was simply what I was told, but the wording was mine. Maybe better would have been to say 'they should have seen it, based on the parameters of their test'. And I definitely agree that no test is proven at this point, including WPIs. So we are working through a drawn-out consensus process.
     
  13. Esther12

    Esther12 Senior Member

    Messages:
    5,296
    Likes:
    5,606
    Ta Kurt.

    It seems like they're further ahead with PC virology stuff... examining how XMRV integrates into human cells, etc. And it's hard to imagine the PC link holding up without their being a problem with a lot of the 0/n studies... but it's all so uncertain. And still possible the link with PC will hold up, but not the link with CFS.

    PS: Hadn't we heard about the study on XMRV being unable to overcome some of our defenses from ages ago? Maybe it's only just been published, but I think there were responses to it. Not sure how convincing they were.
     
  14. dannybex

    dannybex Senior Member

    Messages:
    2,247
    Likes:
    562
    Seattle
    "Easily defeated"?

    Hi Kurt,

    Can you explain this part a little further for those of us with half a brain?

    Thanks,

    d.
     
  15. Esther12

    Esther12 Senior Member

    Messages:
    5,296
    Likes:
    5,606
    I'll have a go to test myself. (I've no idea what I'm talking about, so don't believe me until someone's graded my work):

    Our cells have defenses against various intruders. Much of our DNA is made up of old viruses that got into our genome and use us to replicate... it's important that we're able to restrict these sort of things. All viruses that survive by infecting us have clever ways of getting past our defenses. XMRV is lacking some of the tricks you would expect to see in a human retrovirus. While it is able to infect our cells, it did so (in one experiment) only by sheer numbers and luck. Our cell's defenses were able to push XMRV to mutate at so high a rate that it largely ended up destroying itself, with only a few bugs able to get past our defenses in a form that allowed it to infect and replicate. They also didn't seem to replicate much either.

    That they were forced to evolve so rapidly would also seem problematic given the Wellfare paper, which made much of the low levels of mutation reported in XMRV samples found in human populations. They'd said that being passed from human to human should have forced much, much, much higher levels of mutation to occur.

    All looks bad.

    My hypothetical, ignorant reply would be... maybe XMRV is deeply opportunistic, and is easily spread/highly prevalent, but only able to survive in a form very like that reported. When it gets mutated a bit, it starts to go wrong so that branch of the family tree dies out, and only 'pure' XMRV is able to thrive?

    This would be a very strange way for a virus to behave though (I think).

    It already looks like either XMRV is a contaminant, or a very strange virus though, and it seems like a number of top viroligsts are quite happy to consider the possibility of it being a very strange virus. At this point, I'm not comfortable making predictions, and hope we'll get some solid new from the BWG soon.
     
  16. alex3619

    alex3619 Senior Member

    Messages:
    7,495
    Likes:
    11,978
    Logan, Queensland, Australia
    Restriction factors can indeed stop XMRV, including APOBECs, but this just slows it down. One reason why it can take decades to cause disease. It does not remove the infection. Bye, Alex
     
  17. jace

    jace Off the fence

    Messages:
    855
    Likes:
    170
    England
    Satterfield et al - look at the paper.

    Satterfield et al 2011 (the paper under discussion, available here http://www.retrovirology.com/content/8/1/12) referenced Danielson et al 2010 at no [17] in their paper. They said they knew, from Danielson, that 600ng of prostate DNA was necessary to be able to find XMRV with their PCR assay.

    But what did they do? they used 0.25 ug of DNA (from plasma). Aha! Now can you see there's a problem? They also used the VP62 clone to calibrate, 10 times less sensitive than using live virus.

    Satterfield also, just to make sure this would be another negative paper to outweigh the real stuff, used the cycling conditions and primers from Switzer et al 2010.

    The serology assay that was used in the macaque monkey study found XMRV in the blood of their subjects two weeks after infection. They then found, one month later, that they could no longer detect the XMRV using the VP62 assay. But, as we know, on sacrificing the poor animals (I have problems with this arm of science :worried: *crying*) they found that XMRV had hidden itself away in internal organs. I would suggest that the subjects of Satterfield's study were rather further than two weeks from infection, but I'm glad that no more primates were sacrificed in order to autopsy.
     
  18. alex3619

    alex3619 Senior Member

    Messages:
    7,495
    Likes:
    11,978
    Logan, Queensland, Australia
    Jace, we need biopsy studies. Why is it still all blood? It is not only about the blood banks. Bye, Alex
     
  19. jace

    jace Off the fence

    Messages:
    855
    Likes:
    170
    England
    Good question, Alex. I agree, the blood is not the best place to look by a long chalk, although HGRV's can reliably be found there with long culturing and the right PCR protocols (primers, wild type positive sample for calibration, correct annealing temperatures, etc etc) as Lombardi used. When are we going to get a replication study, instead of these lame attempts to divert the science away from retroviruses?

    I'd rather have a muscle biopsy than a lumber puncture :eek: :headache:
     
  20. kurt

    kurt Senior Member

    Messages:
    1,132
    Likes:
    176
    USA.Earth
    I think Esther did a good job of explaining that study. The title really says it all...

    From reading the study, I believe the NCI study authors were originally attempting to prove XMRV was infectious, and found nearly the opposite, that it has very week defenses against PBMC's hyper-viral-mutation defense mechanism. They had to inject high amounts of the virus (10^6) just to get a little transmission. I don't know how this plays out in epidemiology, but certainly does not make a good case for contagion.

    Probably analysis and discussion of that study belongs in its own thread, if people are interested let me know and I'll move it.
     

See more popular forum discussions.

Share This Page