Why Were Only Polytropic Variants Found in the NIH/FDA research?

[Mya Symons]

Could someone with a science or medical background or someone who has done much research on this explain why some scientist claim the NIH/FDA researchers found a different virus than the Whittmore Peterson researchers? As I understand, Whittmore Peterson found XMRV, the xenotropic version, transferred from human to human AND the FDA/NIH study found PMRV, the polytropic version, transferred from mouse to human. Some researchers say this is what you would expect to see from a retrovirus: different variants. This is what I am not understanding: Why didn't the NIH/FDA researchers find different variants of Xenotropic MuLv's? Shouldn't they have found variants of both Polytropic Murine Related Viruses AND variants of Xenotropic Murine Related Viruses. If not, why not? Where are the other Xenotropic viruses? Can you explain this to a person who knows very little about medical research in a way they would understand? Please and Thank You.

 

[Esther12]

I don't think that it is what you would expect. I don't really know what I'm talking about, but from what I've read of the responses of virologists...

You'd expect some variation, but the differences found with XMRV/CFS do seem to be thought of as unusual.

Maybe that's because MRV's are unusual in some way? Someone made a mistake? - could the WPI have misidentified what they found somehow? I think it's a bit of a mystery at the moment.

 

[Hope123]

Perhaps this is a reflection of my knowledge limits but my feeling is that so little is known about XMRVs/PMRVs at the moment that making too much of a fuss (not you, but some researchers) over this type of discrepancy is premature. I agree more with Alter's views about his results being consistent with WPI's even if they are not exact. There are many instances in medicine/ biology where things are classified one way and then re-classified as more is known.

It would be interesting once they isolate the virus from a sick person(s) (which no one has done technically yet) to try to infect mice with them and see if mice can be used as an animal model to figure out X/PMRV pathogenesis and treatments.

[In fact since WPI thinks transmission might be achieve through plasma, why not take some plasma from X/PMRV+ subjects and try to infect mice with it? WPI thinks it's X based on ?sequencing but has anyone actually tried to infect mice?]

People ask good questions on the boards. As you start digging, you realize we know very little and that not all questions can be answered given the current knowledge, time/ staff/ money limits, etc. It's the same throughout medicine; scientists and med professionals certainly know a lot more now than 50 years ago about many illnesses but ask any good doc and you will find they routinely have several questions come up everyday where there is no solid research on it yet.

 

[grant107]

I remember when the Alter paper was leaked it showed him calling it XMRV. XMRV was in his slides. Did he change it to agree with the CDC. I may be mistaken about this because no one has mentioned it.

 

[Mya Symons]

Some fragments of transcription of the tele-press-conference of NIH/FDA that Mark posted on the telebriefing thread:

I think this may answer my question but is a little above my head.

Dr Alter:
"We think, basically, it confirms the findings of the Whittemore-Peterson Group."

"It does, at least, confirm the findings of Whittemore-Peterson...I think one wants to go back to their studies because they've had more time, and they've done extensive work...so their study is more advanced than ours, but -with that as...er...with them having done the groundwork I think our study is highly confirmatory of their work..."

Dr Lo:
"Our findings show that the more diverse group of virus there is compatible with the XMRV, the xenotropic one, but really not identical and more closely related to the polytropic group."

Q:
"Does the finding of a polytropic virus suggest a looser association with CFS than finding a xenotropic would have?...It would seem to suggest more of a process of chance than a firm association."

Dr Lo:
"No, actually this group of virus we call the polytropic MLV - this is actually more characteristic of a retroviral infection...in fact what we found is it actually gives us a very good confidence this can not be an arbitrary artefact from PCR or contamination because the sequences are so varied from one patient to another."

"They are in the same family but...they are compatible with the earlier finding of the XMRV, however they are not identical and they are more diverse."

Q:
And so is it usual that you would find an association with a single disease with this kind of a variety of viruses?

Lo:
"Yes, indeed, that's exactly what we anticipate for retrovirus infection...over time you will see the many different sequences there."

Alter:
"...The retroviruses...exist in big families of viruses...Hepatitis C is a very good example, nobody's infected with one variant... they have a whole huge family of variants...and if you take any given patient the patient will have multiple variants in them, and different patients will have different variants, so it's very characteristic of these kind of RNA viruses."

"Since the original publication the WPI and NCI groups have found that they too are finding greater variability in their patients so it is not just XMRV as in the original cohort of patients."

Lo:
"I think...we can say we found this kind of variability that's actually MORE consistent with the natural form of a retrovirus infection in this group of patients."

Alter:
(regarding the discrepancies between different groups):
"We think it's in the patient populations rather than the laboratory testing although the latter hasn't been completely ruled out...CFS is a syndrome, that's what it is, it's not a specific biopsy-proven diagnosis, and therefore it's subject to variability..."

 

[George]

Plausible deniability!. (big grins)

 

[Otis]

As George said so well.

The problem I have with the PMLVs being in the same family of Gamma viruses they all have similar sequences in the LTR's (long term repeaters). If you don't isolate and cross reference several hundred sequences you really don't know what you've got. I was ecstatic that Dr. Coffin said it out plain and simple that the sequences that Dr. Alter and Dr. Lo pulled tell us absolutely nothing. They may be Polytropic or they may be Xenotropic. There are less than 700nt per each sequence [a full MLV is 8100ns, give or take] and you really can't tell from that. Even going off of the env sequence is questionable because many MLV's in the Gamma grouping share the some of same env sequences.

And Dr. Coffin in the Q&A from the workshop was very clear that we really only have 1 virus - XMRV. The WPI sequenced two distinct and complete XMRV viruses and proved that it could replicate in a human cell line, etc., etc. It really does tell us how good that Science paper really was.

At first I was confounded about Lo/Alter not finding XMRV and the WPI only finding PLMVs but it turns out that the WPI is seeing these as well and that the VIPDx tests find antibodies for this range of "viruses". What I'm really itching for someone to do is sequence a bunch of these darn things and show they can replicate.

So the answer to the original questions is that Lo/Alter have not found and described new "viruses" - yet. Having said that, based on what we hear from the WPI, they are in all likelihood on the right path and from the Q&A session including Lo & Alter they continue to work.

The picture got more confusing with the Lo/Alter paper but all the hints point to the WPI, Ruscetti and possibly others being pretty far out ahead of the published, or even pre-published data.

 

[alex3619]

Hi Hope123

By definition, mice cannot be infected with XMRV, only PMLVs. Mice are immune to XMRV, which is evidence of strong evolutionary pressure - XMRV or something else killed off every mouse that could be infected by it. The survivors were then immune.

We could use another species, or create a genetically engineered mouse to express the receptor XMRV needs, or whatever else we need to do. Then we could infect mice. Since XMRV can infect almost any mammal, we actually have wide range of species to play with. Mice are favoured because they cost little and breed fast, but I think many studies would be better using larger animals.

Bye
Alex

[In fact since WPI thinks transmission might be achieve through plasma, why not take some plasma from X/PMRV+ subjects and try to infect mice with it? WPI thinks it's X based on ?sequencing but has anyone actually tried to infect mice?]

 

[Hope123]

Hi Hope123

By definition, mice cannot be infected with XMRV, only PMLVs. Mice are immune to XMRV, which is evidence of strong evolutionary pressure - XMRV or something else killed off every mouse that could be infected by it. The survivors were then immune.

That's my point, Alex, *by definitiniton*.

Mikovits et al probably classified their virus as XMRV based on sequencing and what we know to date and, as I've said, I'm no bench scientist. But when face with something new, I feel we need to be as open-minded as possible and for me, experimentation always trumps theory although of course, experiments can be guided by theory and vice-versa.

It's been brought up several times but the whole Helicobacter Pylori thing was proven by a treatment experiment while the theory and other basic science experiments had yet to settle anything. Similar with Semmelweis and handwashing -- his ideas on handwashing between examination of patients were ignored because Pasteur's germ theory of disease had not yet been developed. As someone said earlier, only Nature holds the title to the truth.

http://en.wikipedia.org/wiki/Ignaz_Semmelweis

 

[alex3619]

Hi Hope123

There is at least one species of mouse that can be infected by XMRV, but at two months they couldn't find any pathological signs. Mice have been living with this a long time: looking at infection or treatment in mice is probably a lost cause. However, looking at clues on how to fight the virus makes mice very valuable. We should be looking for mice that can be infected, or we can artificially infect, and show no evidence of disease (once we know what to look for, I keep seeing references to IL-8 everywhere lately). If we can understand how mice have immunity, we can develop strategies to give us immunity.

Oh, and you are quite right to question anything that is "by definition". There is another name for anything like that: dogma. Dogma gives us a starting point, but good science should check everything, even things we think are correct.

Bye
Alex

 

[eric_s]

[...]AND the FDA/NIH study found PMRV, the polytropic version, transferred from mouse to human.[...]

I don't think that's correct, but i'm no scientist. I think PMRV can be transferred between mice, from mice to humans (and maybe other species) and probably between humans, too.

 

[Hope123]

Hi Hope123

There is at least one species of mouse that can be infected by XMRV, but at two months they couldn't find any pathological signs. Mice have been living with this a long time: looking at infection or treatment in mice is probably a lost cause. However, looking at clues on how to fight the virus makes mice very valuable. We should be looking for mice that can be infected, or we can artificially infect, and show no evidence of disease (once we know what to look for, I keep seeing references to IL-8 everywhere lately). If we can understand how mice have immunity, we can develop strategies to give us immunity.

Oh, and you are quite right to question anything that is "by definition". There is another name for anything like that: dogma. Dogma gives us a starting point, but good science should check everything, even things we think are correct.

Bye
Alex

The thing about that mouse study is they used XMRV from a prostate cancer line and from an infectious molecular clone. Even though I remember supposedly that the prostate cancer line XMRV is close to the CFS XMRV, is it the same identical one? This is a similar question to using positive controls that are lab produced vs. positive controls from sick patients in the PCR studies. Should they try using XMRV instead isolated from a patient or patient plasma?

Also researchers should try infecting mice with the PMRV+ plasma from CFS subjects as theoretically, those can infect mice.

It's not just an animal model question; it's also a question of transmission. As Eric stated above me, if X/PMRVs in CFS patients are also able to infect wild mice, I can envision a theory whereby mice infected with X/PMRV are able to spread X/PMRV to human subjects. In prior studies of zoonotic disease epidemiology, like we see with West Nile/ hantavirus, etc., a key component of the investigation is looking at animals in the community where humans have become ill. If we understand tranmission better, it might not help those of us already sick with CFS but might help prevent others from getting ill.

 

[George]

Hi Hope

I'm not sure if I'm following you completely, I think you are asking if the X/P-MLV's are transferred directly from mice to humans at the present time. Is that correct? And if so are they looking for the mice that are able to transfer it to humans at this time?

I don't know if this helps but the following study looks at the receptor. The receptor not only decided how many cells and what type that a retrovirus or virus can enter but also by looking at what it can't enter begins to tell the tale of where it came from and how long.

There isn't a definitive answer but they did start gathering information with this study. All really good questions on your part and I sure hope they get some answers this year. (grins)

Evolution of functional and sequence variants of the mammalian XPR1 receptor for mouse xenotropic gammaretroviruses and the human-derived XMRV.

Yan Y, Liu Q, Wollenberg K, Martin C, Buckler-White A, Kozak CA.

Laboratory of Molecular Microbiology, Bioinformatics and Computational Biosciences Branch, Office of Cyber Infrastructure and Computational Biology, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892.
Abstract

Genetic conflicts between retroviruses and their receptors result in the evolution of novel host entry restrictions and novel virus envelopes, and such variants can influence trans-species transmission. We screened rodents and other mammals for sequence variation in the Xpr1 receptor for the mouse xenotropic/polytropic (X/P-MLV) gammaretrovirus family and for susceptibility to mouse-derived X/P-MLVs and to XMRV, an X-MLV-like virus isolated from humans with prostate cancer and chronic fatigue syndrome. We identified multiple distinct susceptibility phenotypes; these include the 4 known Xpr1 variants in Mus and a novel 5th Xpr1 found in M. molossinus and M. musculus. We describe the geographic and species distribution of the Mus Xpr1 variants, but failed to find the X-MLV-restrictive laboratory mouse allele in any wild mouse. We used mutagenesis and phylogenetic analysis to evaluate the functional contributions made by constrained, variable, and deleted residues. Rodent Xpr1 is under positive selection indicating a history of host-pathogen conflicts; several codons under selection have known roles in virus entry. All non-Mus mammals are susceptible to mouse X-MLVs, but some restrict other members of the X/P-MLV family and the resistance of hamster and gerbil cells to XMRV indicates that XMRV has unique receptor requirements. We show that the hypervariable fourth extracellular XPR1 loop (ECL4) contains 3 evolutionarily constrained residues that do not contribute to receptor function, we identify 2 novel residues important for virus entry (I579, T583), and we describe a unique pattern of ECL4 variation in the 3 virus-restrictive Xpr1 variants found in MLV-infected house mice; these mice carry different deletions in ECL4 suggesting either that these sites or loop size affects receptor function.

Mostly it seems that the XMRV virus has some pretty unique and mutated and restrictive factors that make it truly no longer pathogenic in mice but can be carried by some mice (MUS types) and can be carried by all mammals including humans. The PMLV's have some restrictions within both "other" mammals as well as constraints within mouse species. If I'm reading it correctly and I haven't seen the entire thing just the abstract I think they are indicating the house mice are not a potential carrier for any of the strains detected so far. Course I could be wrong, abstracts can be tricky. (big grins)

 

[Dr. Yes]

Hi George!

Mostly it seems that the XMRV virus has some pretty unique and mutated and restrictive factors that make it truly no longer pathogenic in mice but can be carried by some mice (MUS types) and can be carried by all mammals including humans. The PMLV's have some restrictions within both "other" mammals as well as constraints within mouse species. If I'm reading it correctly and I haven't seen the entire thing just the abstract I think they are indicating the house mice are not a potential carrier for any of the strains detected so far.

From the abstract:

We describe the geographic and species distribution of the Mus Xpr1 variants, but failed to find the X-MLV-restrictive laboratory mouse allele in any wild mouse.

That means they found most mouse species can potentially be infected by "X-MLVs", let alone XMRV, and therefore potentially could be carriers. According to the paper, that would include most house mice. (Of course, no one has yet reported finding XMRV in wild mice, though I think Coffin once said his group was looking..)

Also, the reason XMRV is not infective in (most) lab mouse strains is due less to virus mutation than to mutations in the Xpr1 receptor of those mouse strains, which basically 'rendered' certain MLVs xenotropic. XMRV appears to have gotten its envelope sequences from one of these X-MLVs, and thus has the same inability to infect those lab mouse strains.

[Btw Alex, that is why lab mice can't be infected; they don't have 'immunity' but have a mutation in their Xpr1 receptors that denies X-MLVs access to their cells.]

There are three different variants of a key part of the Xpr1 gene in house mice and one variant in lab mice; they are denoted by superscript letters after "Xpr1" in the paper, but here I'll have to settle for a dash...

Xpr1-sxv is found in Mus domesticus (as well as most non-house wild mice) and is permissive of all X and P-MLVs.

Xpr1-c is found in house mouse species M. castaneus. It restricts P-MLV but not X-MLV.

Xpr1-m is found in two house mouse species, including M. musculus (eastern Europe to the Pacific), is restrictive for all P-MLV and restrictive for many, but not all, X-MLV.

Xpr1-n is restrictive for all X-MLV. They found it only in lab mouse strains (which were derived from a few of the house mouse strains).

A very interesting bit of info from Yan et al: the 4 house mouse species (incl. M. domesticus) are the only wild mouse species that carry endogenous X-MLV and P-MLV env sequences. Not surprisingly, they are the ones with the most variants of the Xpr1 receptor! It's likely that these viruses evolved in those species. Interestingly, they entered the germ line of mice around the same time that restrictive variants "c" and "m" evolved, i.e. more recently than 500,000 years ago.

Another interesting observation in the Yan study is that the general geographic areas with the highest incidence of prostate cancer (the U.S. and Western Europe) correspond roughly to the areas inhabited by house mouse species with the most permissive variant of the Xpr1 receptor, while the reverse is true for the places with the lowest prostate cancer incidence (East Asia and Eastern Europe).

And a fun fact:

..the fact that studies on human/hamster Xpr1 chimeras implicate as yet unidentified ECL4 residues in P-MLV entry.. (44)

They have got to stop crossing humans and hamsters! Last time they did that we wound up with that kid from "The Wonder Years".

 

[eric_s]

LOL.. how did they do that? How can you get "chimeric receptors"? Maybe it's better not to know crazy scientists

 

[George]

Go Dr. Yes! (big grins)

 

[Mya Symons]

Dr. Yes, do you happen to remember the names of the species of the mice with the most variant in receoptors for MuLv in that report? It would be interesting to find out what geographic area these particular mice might be located in the United States and England. Would it explain some of the different results in the research they are doing? I am assuming it would have no bearing on why some studies are finding no positives, but maybe in the studies that are finding different percentages of positives this could be the reason? Never Mind. I realize now that you have the scientific names up there. Ooops. I will try to look them up and see if I can find what the ordinary names for those mice are. --One of those days