PMRV joins XMRV as possible etiologic agent of chronic fatigue syndrome

The new human retrovirus XMRV, first detected in malignant prostate tissue, was subsequently identified in a high percentage of patients with chronic fatigue syndrome (CFS). The virus was not detected in four independent studies of CFS patients in Europe or the United States. The results of a second American study, whose publication was blocked for two months, provide support for the involvement of murine retroviruses in CFS.

The new study, a collaboration among scientists at the Food and Drug Administration, the National Institutes of Health, and Harvard Medical School, utilized samples from 37 CFS patients obtained in the mid-1990s. A key difference from earlier studies is that some repeat samples were used: four obtained two years later and frozen, and eight taken in 2010 and processed without freezing. The patients were from New England, New York State, and North Carolina and were not part of the previous study in which XMRV was detected. Control samples were obtained from 44 healthy blood donors.

Two kinds of polymerase chain reaction (PCR) assays were used to determine the presence of XMRV. In the first, DNA was extracted from PBMC to search for the presence of proviral DNA (e.g., viral DNA integrated into the host chromosome). In the second, RNA was extracted from plasma, and subjected to reverse-transcriptase PCR to detect viral RNA.

The authors found that 32 of 37 samples from CFS patients (86.5%) were positive in the PCR assay of PBMC DNA, compared with 3 of 44 samples (6.8%) from healthy controls. All four samples taken 2 years later, and 7 of the 8 samples taken 15 years later from CFS patients were also positive in this assay. The latter is an important result because it shows presence of virus in patients for long periods. Viral RNA was detected in plasma of 42% of the samples.

The authors performed additional experiments to rule out false positive results. The nucleotide sequences of all PCR products were determined to ensure that they represented the expected target sequence. Furthermore, contamination with mouse DNA was excluded by assaying for murine mitochondrial DNA – it was not found in any of the samples tested.

Perhaps the most interesting finding, aside from the detection of viral RNA in a large fraction of CFS patients, is the observation that the virus detected is not XMRV. Nucleotide sequence analysis of PCR amplified DNA revealed 96.6% nucleotide identity with XMRV, but they are clearly different viruses. The RNA genome of XMRV is a recombinant between the genomes of xenotropic* and polytropic* murine leukemia viruses (Figure). The genome of the viruses detected in the new study appears to be derived from polytropic MLVs, and could be called PMRV. This observation indicates that both xenotropic and polytropic MLV can infect humans in North America.

What is the significace of the observation that both XMRV and PMRV have been associated with CFS? Some investigators have suggested that variation of XMRV is minimal, based on the close identity of sequences obtained from prostate cancer and CFS patients. The finding of a different virus, PMRV, suggests greater genetic variation than previously thought, and could in part explain previous failures to detect MLV-related viruses in the previous four studies of CFS patients. Other explanations, including geographic differences and patient heterogeneity, are also plausible.

I find it very interesting that the authors readily detected PMRV sequences in both PBMC DNA and plasma. The authors of the 2009 Science paper have suggested that these approaches are not sufficiently sensitive to detect viral nucleic acid, and that culturing with susceptible cells must be done to amplify the virus. That explanation, which was used to explain the failure to detect XMRV in the previous four studies, is not likely to be correct. The authors of those studies might want to repeat their assays using primers more likely to detect PMRV.

Where do these findings lead next? The authors of an accompanying commentary have two appropriate suggestions:

At this juncture, it would seem reasonable to conduct extensive case-control studies in North America…using coded samples from subject with inflammatory disease to determine the frequency of MLV infection in patients with CFS. The potential transmission of MLV-related sequences from human to human should also be epidemiologically evaluated. […]it would also be appropriate to conduct interventional studies.

It is possible that these studies, which are needed to determine if MLVs cause human disease, might not have been done without a second independent confirmation of the association of MLV-related viruses with CFS.

*Xenotropic murine leukemia viruses infect non-mouse cells, while polytropic murine leukemia viruses can infect the cells of mice and other mammals.

Lo SC, Pripuzova N, Li B, Komaroff AL, Hung GC, Wang R, & Alter HJ (2010). Detection of MLV-related virus gene sequences in blood of patients with chronic fatigue syndrome and healthy blood donors. Proceedings of the National Academy of Sciences of the United States of America, 107 (36), 15874-9 PMID: 20798047

FDA and NIH confirm WPI XMRV findings

A press release from the Netherlands indicates that the FDA and NIH have independently confirmed the association of XMRV with chronic fatigue syndrome as published last fall in Science. Apparently two journalists for the Dutch magazine ORTHO obtained a copy of a lecture by Dr. Harvey Alter in Zagreb which confirms these findings. According to

The ORTHO journalists were able to obtain a pdf document of the lecture given by Harvey Alter at the IPFA/PEI 17th Workshop on ‘Surveillance and screening of Blood Borne Pathogens’ in Zagreb. The International Plasma Fractionation Association (IPFA) represents the not-for-profit organizations around the world involved in plasma fractionation. The IPFA is based in Amsterdam, the Netherlands.

The highly-experienced Dr. Harvey Alter is Clinical Studies Chief at the Infectious Diseases and Immunogenetics Section of the Department of Transfusion Medicine at the NIH Clinical Center in Bethesda. “The data in the Lombardi, et al Science manuscript are extremely strong and likely true, despite the controversy”, was one comment on the XMRV findings reported by Alter in Zagreb. “Although blood transmission to humans has not been proved, it is probable. The association with CFS is very strong, but causality not proved. XMRV and related MLVs are in the donor supply with an early prevalence estimate of 3%‐7%. We (FDA & NIH) have independently confirmed the Lombardi group findings.”

ORTHO contacted Dr. Harvey Alter today for a reaction. He did not want to comment, but confirmed that a paper is soon to be published.

I’m not entirely sure what it means to have confirmed the Lombardi group findings. Did the FDA and NIH use the same clinical specimens, or independently collected ones? We’ll have to wait for the article to appear to find out.