XMRV and Culturing, HERV's and more

[Esther12]

From what I remember reading on hearing at the time of the Science paper, I think this could well have confused a fair few academic virologists.

I'm not really able to follow this discussion properly, but it sounds like different procedures were followed than a lot of the summaries I heard and read at the time thought. I'd have assumed that those running replication studies would be more thorough than those writing blog posts, etc - but it still seems to imply there was some confusion here.

 

[Gerwyn]

no it would not have confused them the only plausible explanation is the one given by Rich assuming no malintent

 

[ukxmrv]

Esther,

If they were confused they should have asked. The worst thing to do was blunder in (with their recipe for "fruitcake) and do the test as per HIV (because that is what they know).

The post-Science published papers squandered money, made outrageous claims and didn't attempt to follow even the most basic of the Science protocols.

Frozen blood - possibly well past it's "sell by date" in the case of the Dutch.

Were they confused over the blood, the cohort - no.

Once they have started in that manner, it may well be irrelevant what they did next but the intent of what they did is clear. They never intended to follow any procedure in Science or to do a good, thorough, well defined job. They made an error of judgment.

If I acted like that in my profession I would be fired.

Deliberate and willful "ignorance" is no excuse

Then to blame the WPI and hope for a retraction or "clarification" in Science.

It's back to the "We are Sick not Stupid" motto

 

[Cort]

Like I said we'll see what happens. Hopefully we'll have some good news soon.

 

[Gerwyn]

Like I said we'll see what happens. Hopefully we'll have some good news soon.

i hope so to cheers!

 

[kurt]

hervs do not carry the genetic equipment to manufacture Sag full stop

Gerwyn, do you have references on that? Maybe an older view? Yes, not all HERVs are viable but some appear capable of protein manufacture due to recent studies. Here is a study into the real evidence that HERVs have some SAg production capability, correlated with EBV:

http://www.ncbi.nlm.nih.gov/pubmed/11672540?dopt=Abstract

And this one is more detailed, using a mouse model (full paper here):

http://www.jimmunol.org/cgi/content/full/177/5/3178

Also, please note that my discussion of HERVs has always been in the context of the antibodies used in the WPI study. Not the PCR testing. WPI's ruling out of HERV involvement was for the PCR portion of the test. What interests me is the high rate of antibody positives, and what that might mean IF those were hits on HERVs.

 

[kurt]

cort a competent virologist could easily follow those instructions they were not constructed for laymen but proffessional retrovirologists

it may well "seem" messy to a layman but to the science peer reviewers it was clear enough as indeed it is to me.Now Mclure and Groom have many o rders of magnitude more knowledge in this area than me.You tell me why the methods and proceedures were not clear to them.

the WPi said that their patients were typical of patients with ME/cfs using their diagnostic criterea NOT typical of the patients labelled with CFS by the Oxford mythodology.The two patient cohorts are clearly different and Dr Vernon should be repeatedly stating that

The professional retrovirologists understood the paper well enough, although I have been told it was not easy even for them due to the dense writing. But they did not follow the paper exactly because they were conducting validation and not replication, thus using their own study designs. What they got from the paper was the main idea which is simple enough, cells were found to be infected with XMRV virus by PCR and there are antibody and WB positives for MuLV. How they designed their own attempts to validate that finding is up to them. These are professionals with more knowledge than we on this forum, I think they know how to construct a validation study. Somebody has to find XMRV using tests different from WPI, otherwise false positives are suspect in the original Science paper. That is just the way this type of science works.

 

[Gerwyn]

The professional retrovirologists understood the paper well enough, although I have been told it was not easy even for them due to the dense writing. But they did not follow the paper exactly because they were conducting validation and not replication, thus using their own study designs. What they got from the paper was the main idea which is simple enough, cells were found to be infected with XMRV virus by PCR and there are antibody and WB positives for MuLV. How they designed their own attempts to validate that finding is up to them. These are professionals with more knowledge than we on this forum, I think they know how to construct a validation study. Somebody has to find XMRV using tests different from WPI, otherwise false positives are suspect in the original Science paper. That is just the way this type of science works.

no you cant validate untill you have calibrated your method agaist a known positive.The writing was standard whatever you were "told".No your attempt at representing the science study is a gross over simplification and factually incorrect.

I know facts are inconvenient try reading the paper it sometimes helps.

No "type of science" works this way and never has.Repeating the same tired old mantra does not make it true.

You might think they knew how to construct a study attempting to find xmrv unfortunately the facts prove you wrong.

If the english retrovirologists understood the study well enough(what evidence have you to back up that claim by the way) why are they now asking for clarifycation.

The point you made about false positives indicates to me that you have not read the study or failed to understand it or both

 

[ukxmrv]

I wonder if Science works in the way that the McClure crowd did it.

Announce a press conference and tell everyone that there is no XMRV in UK CFS.

Maximum publicity and a "1000 per cent certain that there is no XMRV in UK CFS".

Where does the line between Science, misinformation and unsubstantiated claims start? It's been all spin from them.

After all they (the UK researchers in the McClure group) now know that people with CFS have been tested in the UK and are positive. So what do they do? Retract the paper, recall the press conference, issue an apology, make sure that they get treatment - no.

The charade goes on here. It's not Science, it's politics.

 

[kurt]

You might think they knew how to construct a study attempting to find xmrv unfortunately the facts prove you wrong.

Exactly which facts prove this point wrong? Has someone else validated the WPI finding that I have not heard of?

Do we know with certainty that XMRV was in that sample cohort and the European tests could not find it?

Perhaps you are referring to the anecdotal reports of positive WPI tests among Europeans. But none of that validates, that would be anecdotal replication, but certainly not proof that the European tests were invalid. Speculation, as you like to say, is not proof.

 

[Gerwyn]

Exactly which facts prove this point wrong? Has someone else validated the WPI finding that I have not heard of? Do we know with certainty that XMRV was in that population and the European tests could not find it?

the cleveland clinic you cant validate untill you calibrate your assay otherwise you produce unfalsyfiable results . That is very simple basic science.

you cant prove that the european tests were invalid:they designed a study with unfalsifiable results:that is proof that they did not know how to design a validation study

That is not speculation but fact.

 

[kurt]

the cleveland clinic you cant validate untill you calibrate your assay otherwise you produce unfalsyfiable results . That is very simple basic science.

Is this what you are referring to? (From the Science paper)

Detection of XMRV was confirmed in 7 of 11 WPI CFS samples at the Cleveland Clinic by PCR-amplifying and sequencing segments of XMRV env [352 nucleotides (nt)] and gag (736 nt) in CFS PBMC DNA (Fig. 1A) (6). In contrast, XMRV gag sequences were detected in 8 of 218 (3.7%) PBMC DNA specimens from healthy individuals. Of the 11 healthy control DNA samples analyzed by PCR for both env and gag, only one sample was positive for gag and none for env (Fig. 1B). In all positive cases, the XMRV gag and env sequences were more
than 99% similar to those previously reported for prostate tumor–associated strains of XMRV (VP62, VP35, and VP42) (fig. S1) (5).

So are you saying that the labs should have sequenced detected XMRV? How could they do that when their PCR test, using the identified sequence with only slight variation from VP62, found no positive hits? Not sure what you are referring to regarding calibration. They acquired positive cells and calibrated their probes, they had to in order to construct a PCR, right?

 

[Gerwyn]

Is this what you are referring to? (From the Science paper)



So are you saying that the labs should have sequenced detected XMRV? How could they do that when their PCR test, using the identified sequence with only slight variation from VP62, found no positive hits? Not sure what you are referring to regarding calibration. They acquired positive cells and calibrated their probes, they had to in order to construct a PCR, right?

you cant use an assay without first calibrating it afainst a known positive blood sample containing the virus that you are looking for.To do therwise produces results which are unflsifyable.Simple basic science

they did not aquire a positive blood sample. nor positive cells of anykind for the PCR. Calibration of an assay method and validation of an assay method are entirely different.

 

[Gerwyn]

In case anyone does not know there are two kinds of validation
"Validation of an analytical method is the process that establishes, by laboratory studies, that the performance characteristics of the method meet the requirements for the intended analytical applications."

Before the english could go about validating the wpi study they first had to validate their own proceedures

 

[kurt]

In case anyone does not know there are two kinds of validation
"Validation of an analytical method is the process that establishes, by laboratory studies, that the performance characteristics of the method meet the requirements for the intended analytical applications."

Before the english could go about validating the wpi study they first had to validate their own proceedures

Yes, validation of methodology and then validation of a finding are definitely two different levels of validation.

I thought all of those research teams reported acquiring a positive cell line (have not re-read the studies though). And they would have run positive and negative controls with each batch. They knew their resolution. What other type of method validation are you referring to?

Also note that this is still research and not clinical. The original finding is not yet validated by outside labs using different tests on different patient samples that match the original criterion. Therefore, 'whose test is right' is still in question. If the WPI test is for some reason NOT finding XMRV, if there was some type of systematic error, then neither will any validation studies.

 

[Cort]

I know facts are inconvenient try reading the paper it sometimes helps.

Can we PLEASE dispense with these remarks!

My understanding was that they all validated their ability to find XMRV by running a test sample first. First they show they can find XMRV at a certain resolution or sensitivity. Then once they've proved that they find XMRV at 'X' level then they look for it in the patient samples. In the first study it was a water sample I remember - I don't remember the other ones.

Isn't that kind of a given in these types of studies? They use a standard sample to show they can find it - and then they look for it?

 

[Gerwyn]

Can we PLEASE dispense with these remarks!

My understanding was that they all validated their ability to find XMRV by running a test sample first. First they show they can find XMRV at a certain resolution or sensitivity. Then once they've proved that they find XMRV at 'X' level then they look for it in the patient samples. In the first study it was a water sample I remember - I don't remember the other ones.

Isn't that kind of a given in these types of studies? They use a standard sample to show they can find it - and then they look for it?

The points kurt keeps raising are clear in the science paper either he has not read the paper or he does not understand what he read.

They did not validate their assays with an known positive blood sample

what you are referring to is calibration proceedures not validating that the method is capable of detecting XMRV if present.the latter is validation.it is confusing and an easy mistake to make



perhaps we can also dispense with these remarks

"The professional retrovirologists understood the paper well enough, although I have been told it was not easy even for them due to the dense writing." pure speculation

and these

" What they got from the paper was the main idea which is simple enough, cells were found to be infected with XMRV virus by PCR and there are antibody and WB positives for MuLV. How they designed their own attempts to validate that finding is up to them."

Both factually inaccurate scientists dont get a main idea from a paper: they "get" the detailed protocol as anyone with a claim to be a scientist knows.If Kurt is right then such a gross departure from the scientific method and they only took the "idea" from the paper and decided their protocols on a whim would explain why the study designs produced unfalsyfiable results and conclusions.One Scientific "sin" lead to another.

The points KURT keeps raising are not supported by any observed data published in any of the papers or even contrary to such observations

 

[Gerwyn]

Yes, validation of methodology and then validation of a finding are definitely two different levels of validation.

I thought all of those research teams reported acquiring a positive cell line (have not re-read the studies though). And they would have run positive and negative controls with each batch. They knew their resolution. What other type of method validation are you referring to?

Also note that this is still research and not clinical. The original finding is not yet validated by outside labs using different tests on different patient samples that match the original criterion. Therefore, 'whose test is right' is still in question. If the WPI test is for some reason NOT finding XMRV, if there was some type of systematic error, then neither will any validation studies.

no none of the english researchers used a positive cell line in fact McLure et all used a negative cell line

As you said yourself Kurt professional retrovirologists know a lot more than you in these matters and none have raised the possibility of a systematic error of any kind

 

[Gerwyn]

Yes, validation of methodology and then validation of a finding are definitely two different levels of validation.

I thought all of those research teams reported acquiring a positive cell line (have not re-read the studies though). And they would have run positive and negative controls with each batch. They knew their resolution. What other type of method validation are you referring to?

Also note that this is still research and not clinical. The original finding is not yet validated by outside labs using different tests on different patient samples that match the original criterion. Therefore, 'whose test is right' is still in question. If the WPI test is for some reason NOT finding XMRV, if there was some type of systematic error, then neither will any validation studies.

you cant validate a study without validating your assay proceedures without producing unfalsifyable results

 

[fairlight]

I'm a little confused about the focus of this thread but my stomach biopsy tested positive to Dr. Chia's enterovirus. Blood tests also positive. I have a lot of stomach issues. I'll get my test results tomorrow from a 24-hour ph acid test (horrible test where they put a tube down your nose and send you home for 24 hours) and a barium swallow and ct scan.

I feel much beter if I don't eat because my food just seems to sit in my esophageous and doesn't digest. I also have gastorparesis.. I also could have a vagus nerve problem. Are these problems from the enterovirus?