Abstract
Gammaretroviruses related to murine leukemia virus (MLV) have variously been reported to be present or absent in blood from chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) patients and healthy controls. Using subjects from New York State, we have investigated by PCR methods whether MLV-related sequences can be identified in nucleic acids isolated from whole blood or from peripheral blood mononuclear cells (PBMCs) or following PBMC culture. We have also passaged the prostate cancer cell line LNCaP following incubation with plasma from patients and controls and assayed nucleic acids for viral sequences. We have used 15 sets of primers that can effectively amplify conserved regions of murine endogenous and exogenous retrovirus sequences. We demonstrate that our PCR assays for MLV-related gag sequences and for mouse DNA contamination are extremely sensitive. While we have identified MLV-like gag sequences following PCR on human DNA preparations, we are unable to conclude that these sequences originated in the blood samples.
Highlights
The 2009 report [1] that XMRV was associated with CFS sparked our interest in examining additional populations to determine whether we could replicate the results and observe variability from the originally reported sequences
peripheral blood mononuclear cells (PBMCs) were isolated and examined directly or cultured in order to determine whether murine leukemia virus (MLV)-related viruses could be detected
PCR was performed with genomic DNA (gDNA) and cDNA produced from uncultured and 5-day and 9/10-day cultured PBMCs and plasma-inoculated LNCaP cells
Summary
The 2009 report [1] that XMRV (xenotropic murine leukemia virus-related virus) was associated with CFS sparked our interest in examining additional populations to determine whether we could replicate the results and observe variability from the originally reported sequences. CFS/ME is a debilitating illness without a known cause and no generally effective treatment [2,3,4,5,6]. Aspects of the illness, including a number of outbreaks [7,8], are consistent with involvement of a virus. Our initial attempts to utilize published primers in PCR assays to detect XMRV failed; some experiments resulted in identification of gag sequences similar to MLV. With the report in 2010 [9] of detection of MLV-like gag sequences in CFS patient blood samples, we decided to explore our findings further. As reports of laboratory [10,11] and reagent contamination [12] began to appear, we investigated the possibility of spurious results and the possible sources of the sequences we observed
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