Is a mouse leukemia virus contaminating vaccine reagents the cause of Chronic Fatigue Syndrome?
source: TheShiningLight’s Blog
New York Times:
Virus Is Found in Many With Chronic Fatigue Syndrome
By DENISE GRADY
Published: October 8, 2009Many people with chronic fatigue syndrome are infected with a little known virus that may cause or at least contribute to their illness, researchers are reporting.
The syndrome, which causes prolonged and severe fatigue, body aches and other symptoms, has long been a mystery ailment, and patients have sometimes been suspected of malingering or having psychiatric problems rather than genuine physical ones. Worldwide, 17 million people have the syndrome, including at least one million Americans.
An article published online Thursday in the journal Science reports that 68 of 101 patients with the syndrome, or 67 percent, were infected with an infectious virus, xenotropic murine leukemia virus-related virus, or XMRV. By contrast, only 3.7 percent of 218 healthy people were infected. Continuing work after the paper was published has found the virus in nearly 98 percent of about 300 patients with the syndrome, said Dr. Judy A. Mikovits, the lead author of the paper.
XMRV is a retrovirus, a member of the same family of viruses as the AIDS virus. These viruses carry their genetic information in RNA rather than DNA, and they insert themselves into their hosts’ genetic material and stay for life.
Dr. Mikovits and other scientists cautioned that they had not yet proved that the virus causes the syndrome. In theory, people with the syndrome may have some other, underlying health problem that makes them prone to being infected by the virus, which could be just a bystander. More studies are needed to explain the connection.
But Dr. Mikovits said she thought the virus would turn out to be the cause, not just of chronic fatigue, but of other illnesses as well. Previous studies have found it in cells taken from prostate cancers.
“I think this establishes what had always been considered a psychiatric disease as an infectious disease,” said Dr. Mikovits, who is research director at the Whittemore Peterson Institute in Reno, a nonprofit center created by the parents of a woman who has a severe case of the syndrome. Her co-authors include scientists from the National Cancer Institute and the Cleveland Clinic….
Oops: Contaminated cell-lines from the NIH
Category: HIV/AIDS
Posted on: January 5, 2009 11:15 AM, by ERV…Well, heres where the ‘oops’ comes in. Every HIV-1 lab in the country uses TZM-bl cells for something. All from the same stock from the NIH reagent bank. Turns out this stock has been contaminated with another retrovirus- murine leukemia virus.
This doesnt effect me at all, personally. MLV is a gamma retrovirus, so it has no effect on the blue/glow assay I do every day (MLV doesnt have the right proteins to interfere with that assay, which is HIV specific), BUT it could be screwing over other labs if they are doing something different (like measuring reverse transcriptase activity levels).
This paper brings up a very, very important lesson: ALWAYS DO ALL THE RIGHT CONTROLS IN YOUR EXPERIMENTS, INCLUDING MOCK/NEGATIVE CONTROLS!!!
This lab discovered that something was ‘wrong’ with their cells (which turned out to be ALL of our cells) by doing all of the appropriate controls… and getting unexpected results. If they had skipped their negative controls, they wouldnt have noticed that they were getting reverse transcriptase activity off of their uninfected cells.
Can never have too many controls.
More contamination by MLV here, and here This reference specifically mentions vaccines:
Viral Evaluation of Animal Cell Lines Used in Biotechnology
By: Alasdair J. Shepherd2, Kenneth T. Smith
Abstract
Animal cell culture is being increasingly used for production of therapeutic reagents such as monoclonal antibodies, recombinant proteins, viral vaccines and replication incompetent viral vectors for gene therapy. Material derived from a variety of biological processes has been associated in the past with incidents involving the transmission of infectious agents, principally viruses (1). Thus, virological evaluation of animal cell substrates for use in the manufacture of biologicals is essential to ensure the safety of products for pharmaceutical use.
Affiliation(s): (2) Q-One Biotech Ltd., Glasgow, Scotland, UK
Book Title: Animal Cell Biotechnology: Methods and Protocols
Series: Methods in Biotechnology | Volume: 8 | Pub. Date: Feb-22-1999 | Page Range: 23-36 | DOI: 10.1385/0-89603-547-6:23
It seems the highly inbred standard laboratory mouse is prone to retroviral infections.
Xenotropic/Ecotropic Murine Leukemia Virus Testing
The risk of viral contamination is a feature common to all biotechnology products derived from cell lines. Murine cells used to produce biopharmaceutical products are known to harbor infectious retroviruses due to the breeding program (loss of a suppressor function on endogenous virus sequences) when used to produce the standard laboratory mouse. Retroviruses in these murine cells pose a particular problem in assessing the safety of cell substrates, as they may be transmitted in the germ line, since the viral genome persists within the cell. These endogenous retroviruses may be expressed without deleterious effects on the cells, and could be infectious to human cells….
The clincher is that this mouse leukemia virus sounds similar to a genetically engineered virus that was recently discovered to have escaped a research lab. Wonder what affect this one will have on us?
Retrovirology 2009, 6:86doi:10.1186/1742-4690-6-86
Unintended spread of a biosafety level 2 recombinant retrovirus
Received: 23 April 2009
Accepted: 22 September 2009
Published: 22 September 2009
Abstract
Background
Contamination of vertebrate cell lines with animal retroviruses has been documented repeatedly before. Although such viral contaminants can be easily identified with high sensitivity by PCR, it is impossible to screen for all potential contaminants. Therefore, we explored two novel methods to identify viral contaminations in cell lines without prior knowledge of the kind of contaminant.
Results
The first hint for the presence of contaminating retroviruses in one of our cell lines was obtained by electron microscopy of exosome-like vesicles released from the supernatants of transfected 293T cells. Random amplification of particle associated RNAs (PAN-PCR) from supernatant of contaminated 293T cells and sequencing of the amplicons revealed several nucleotide sequences showing highest similarity to either murine leukemia virus (MuLV) or squirrel monkey retrovirus (SMRV). Subsequent mass spectrometry analysis confirmed our findings, since we could identify several peptide sequences originating from monkey and murine retroviral proteins. Quantitative PCRs were established for both viruses to test currently cultured cell lines as well as liquid nitrogen frozen cell stocks. Gene fragments for both viruses could be detected in a broad range of permissive cell lines from multiple species. Furthermore, experimental infections of cells negative for these viruses showed that both viruses replicate rapidly to high loads. We decided to further analyze the genomic sequence of the MuLV-like contaminant virus. Surprisingly it was neither identical to MuLV nor to the novel xenotropic MuLV related retrovirus (XMRV) but showed 99% identity to a synthetic retrovirus which was engineered in the 1980s.
Conclusion
The high degree of nucleotide identity suggests unintended spread of a biosafety level 2 recombinant virus, which could also affect the risk assessment of gene-modified organisms released from contaminated cell cultures. The study further indicates that both mass spectrometry and PAN-PCR are powerful methods to identify viral contaminations in cell lines without prior knowledge of the kind of contaminant. Both methods might be useful tools for testing cell lines before using them for critical purposes.
….
The present report extents these studies by identifying for the first time a presumably synthetic chimeric retrovirus as a contaminant. This gene-modified organism seems to have replicated and spread intensely in a broad set of cell lines for several years without being noticed. This hybrid amphotropic/Moloney murine leukemia virus was engineered in the 1980s [7,8] and neither the virus itself nor the plasmid (pAMS) containing its proviral genome were ever used in our laboratory. Although the precise source for the contamination could not be traced back, sharing cell lines with other laboratories seems the most likely explanation. A frozen aliquot of 293T cells (HEK 293tsA201), which we obtained from ECACC, was not contaminated. While SMRV contaminations were detected in different laboratories, testing of three other laboratories’ cell lines did not reveal contaminations with the hybrid amphotropic/Moloney murine leukemia virus (data not shown).
In this document, Stanford says this bioengineered virus will infect humans via the injection route:
Moloney Murine Leukemia Virus (MoMuLV) (9)
Virology: Retroviridae; subfamily oncovirinae type C, enveloped, icosahedral core, virions 100 nm in diameter, diploid, single stranded, linear RNA genome. MoMuLV integrates into the host genome and is present in infected cells as a DNA provirus. Cell division is required for infection.
Virus is not lytic.
Data suggests a pathogenic mechanism in which chronic productive retroviral infection allowed insertional mutagenesis leading to cell transformation and tumor formation. The nature of a transgene or other introduced genetic element may pose additional risk.
The host range of recombinant MoMuLV vectors is dependent on the specificity of the viral envelope. The ecotropic env gene produces particles which infect only murine cells. Amphotropic env allows infection of murine and nonmurine cells, including human cells. VSV-G envelope allows infection in a wide range of mammalian and non-mammalian cells.
Clinical features: None to date.
Epidemiology: MoMuLV infects only actively dividing cells. In mice, the virus is transmitted in the blood from infected mother to offspring. Transmission may also occur via germline infection. In vivo infection in humans appears to require direct injection with amphotropic or pseudotyped virus.
Treatment: No recommended treatment.
Though the researchers that discovered the contaminated cell lines classified this virus as one that should be worked with under the guidelines of biosafety level 2 precautions, the Standford document says that the mice may be housed at only biosafety level 1.