Development and application of a high-throughput microneutralization assay: Lack of xenotropic murine leukemia virus-related virus and/or murine leukemia virus detection in blood donors

Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California, United States
Transfusion (Impact Factor: 3.23). 02/2012; 52(2):332-42. DOI: 10.1111/j.1537-2995.2011.03519.x
Source: PubMed

ABSTRACT Xenotropic murine leukemia virus (MLV)-related virus (XMRV) and other related MLVs have been described with chronic fatigue syndrome and certain types of prostate cancer. In addition, prevalence rates as high as 7% have been reported in blood donors, raising the risk of transfusion-related transmission. Several laboratories have utilized microneutralization assays as a surrogate marker for detection of anti-MLV serologic responses--with up to 25% of prostate cancer patients reported to harbor neutralizing antibody responses.
We developed a high-throughput microneutralization assay for research studies on blood donors using retroviral vectors pseudotyped with XMRV-specific envelopes. Infection with these pseudotypes was neutralized by sera from both macaques and mice challenged with XMRV, but not preimmune serum. A total of 354 plasma samples from blood donors in the Reno/Tahoe area were screened for neutralization.
A total of 6.5% of donor samples gave moderate neutralization of XMRV, but not control pseudotypes. However, further testing by Western blot revealed no evidence of antibodies against MLVs in any of these samples. Furthermore, no evidence of infectious virus or viral nucleic acid was observed.
A microneutralization assay was developed for detection of XMRV and can be applied in a high-throughput format for large-scale studies. Although a proportion of blood donors demonstrated the ability to block XMRV envelope-mediated infection, we found no evidence that this inhibition was mediated by specific antibodies elicited by exposure to XMRV or MLV. It is likely that this moderate neutralization is mediated through another, nonspecific mechanism.

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Available from: Imke Steffen, Mar 05, 2014
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    • "The reasons of failure to detect XMRV sequences in our specimens are unlikely due to the methods used, such as DNA extraction and PCR amplification, because both beta-actin in all prostate specimens and the env region of MLV in YAC-1 cells were successfully amplified. Because of the high degree of homology between XMRV and related MLV provirus sequences present in mouse genomic DNA, it has been shown that XMRV specific PCR assays can cross-react with MLV provirus sequences (Urisman et al., 2006; Silverman et al., 2010; Zhou et al., 2012; Rezaei et al., 2013). PCR assays described herein detected MLV provirus env sequences present in mouse genomic DNA (YAC-1 cells) agrees with result of other studies (Danielson et al., 2010; Tang et al., 2012; Rezaei et al., 2013). "
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    ABSTRACT: XMRV (xenotropic murine leukemia virus-related virus) was initially discovered in association with prostate cancer and later with chronic fatigue syndrome (CFS). Its association with CFS is now largely discredited, and current results support a laboratory origin for XMRV with no reproducible evidence for infection of humans. However, some results indicating the presence of XMRV in prostate cancer are difficult to attribute to sample contamination. Here we have sought biological evidence that might confirm the presence of XMRV in prostate cancer samples previously having tested positive. We have tested for infectious XMRV and neutralizing antibodies against XMRV in blood plasma from 29 subjects with prostate cancer, and for infectious XMRV in prostate secretions from another five prostate cancer subjects. Nine of these subjects had previously tested positive for XMRV by PCR or by virus assay. We did not detect XMRV or related retroviruses in any sample, and the neutralizing activities of the plasma samples were all very low, a result inconsistent with XMRV infection of the plasma donors. We find no evidence for XMRV infection of any human subject tested, either by assay for infectious virus or for neutralizing antibodies. Our results are consistent with the majority of published studies on XMRV, which find that XMRV is not present in humans. The observed low to undetectable XMRV neutralization by human plasma indicates a lack of innate restriction of XMRV replication by soluble factors in human blood.
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