XMRV in monozygotic twins discordant for chronic fatigue syndrome

[Jemal]

Diagn Microbiol Infect Dis. 2011 Jul 25. [Epub ahead of print]
Xenotropic murine leukemia virus-related virus in monozygotic twins discordant for chronic fatigue syndrome.
Jerome KR, Diem K, Huang ML, Selke S, Corey L, Buchwald D.
Department of Laboratory Medicine, University of Washington, Seattle, WA, USA; Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.

Abstract
A recent report suggested an association between xenotropic murine leukemia virus-related virus (XMRV) and chronic fatigue syndrome (CFS). If confirmed, this would suggest that antiretroviral therapy might benefit patients suffering from CFS. We validated a set of assays for XMRV and evaluated the prevalence of XMRV in a cohort of monozygotic twins discordant for CFS. Stored peripheral blood mononuclear cell (PBMC) samples were tested with 3 separate polymerase chain reaction (PCR) assays (one of which was nested) for XMRV DNA, and serum/plasma was tested for XMRV RNA by reverse transcription (RT)-PCR. None of the PBMC samples from the twins with CFS or their unaffected co-twins was positive for XMRV, by any of the assays. One plasma sample, from an unaffected co-twin, was reproducibly positive by RT-PCR. However, serum from the same day was negative, as was a follow-up plasma sample obtained 2 days after the positive specimen. These data do not support an association of XMRV with CFS.

Copyright 2011 Elsevier Inc. All rights reserved.

http://www.ncbi.nlm.nih.gov/pubmed/21795004

 

[Jemal]

One plasma sample, from an unaffected co-twin, was reproducibly positive by RT-PCR. However, serum from the same day was negative, as was a follow-up plasma sample obtained 2 days after the positive specimen.

This is weird? If I understand correctly they did find XMRV, but when they couldn't find it again they just gave up? Shouldn't they be worried about their detection methods?

 

[eric_s]

Or contamination. But then please find it.

 

[FancyMyBlood]

This is all they said in the FT about the one 'positive':

Only a single plasma sample, from one
unaffected co-twin, had detectable XMRV, at 5000 copies/
mL using the X2 RNA assay. A second 10-?L RNA aliquot of
this sample was run and was again positive. However, a serum
sample obtained on the same date was negative and another
plasma sample obtained from the same person 2 days later was
also negative. Primers for the integrase region confirmed the
positive sample (negative controls included with this run were
negative). Efforts to sequence the amplicons were not
successful, and thus we cannot confirm or rule out the
possibility that the single positive sample was contaminated
with control or other exogenous material.

 

[Jemal]

This is all they said in the FT about the one 'positive':

They cannot confirm or rule out... so basically they have no idea what they found. Could be XMRV, could be contamination, yet they assume it was contamination I guess. Why haven't they done more research? I know I would like to find out what was going on, if I found something like this.

 

[RedRuth]

They assume it's contamination. The fact that they couldn't sequence is odd.

 

[FancyMyBlood]

They assume it's contamination. The fact that they couldn't sequence is odd.

Why is it odd? What could this imply? (total layman)

 

[eric_s]

Primers for the integrase region confirmed the
positive sample (negative controls included with this run were
negative).

What could we conclude from this?
Does this mean it was XMRV or could it also have been mouse material?

 

[RedRuth]

What could we conclude from this?
Does this mean it was XMRV or could it also have been mouse material?

The integrase is part of the Pol gene and codes for the integrase enzyme. It means the positive probably wasn't the result of non specific priming but doesn't rule out contamination.

 

[RedRuth]

Why is it odd? What could this imply? (total layman)

Well sequencing can fail for a number of reasons and they don't say how many times it failed but it could mean a mixed PCR product (non specific plus specific priming). These days sequencing is really reliable and I've only had problems when I've had contamination or non specific priming.

 

[eric_s]

The integrase is part of the Pol gene and codes for the integrase enzyme. It means the positive probably wasn't the result of non specific priming but doesn't rule out contamination.

What i would like to know is if in the case of contamination it would have to be contamination with XMRV or wheter it could be contamination with mouse material that contains mouse endogenous retroviruses or anything else that could give a false positive.

 

[RedRuth]

What i would like to know is if in the case of contamination it would have to be contamination with XMRV or wheter it could be contamination with mouse material that contains mouse endogenous retroviruses or anything else that could give a false positive.

That depends on the specificity of the primers and the annealing temp but I guess the most obvious answer is that it's contamination by their positive control.