Ruscetti has admitted that the two positive lanes in Figure 2c from the Science paper, which contained activated PBMC from CFS patients, were treated with 5AZA. He mentioned only the two CFS patient lanes, and did not say the controls were also treated. This was not mentioned in the paper. Ruscetti says it was a trivial omission, not necessary to the paper. He is wrong, and he must know better. This is why. I'm going to start from the basics, so the story is complete. When a retrovirus infects an animal - lets say, a human - it sometimes integrates a viral DNA sequence into the DNA of infected cells. This is a 'provirus.' The integrated provirus can ay silent, or it can express viral proteins and RNA and make new virus. One mechanism the cell uses to try to turn the provirus off, is to methylate it - add methyl groups to the DNA. Methylation turns off expression of genes. Sometimes a cell infected with a 'provirus' can be passed on through sperm or egg to the person's children. If that happens, the provirus sequence has now become part of the human Genome, part of our genetic heritage. It turns out that a very large part of our human DNA consist of ancient retrovirus insertions, often from millions of years ago or more. They have mutated enough over many generation that they don't make functional virus. They are silenced by methylation. These are called endogenous retroviruses - retrovirus sequences that are not infectious and have become part of our DNA. They are called ERV's for short - and yes, this is where Abbie Smith took the name of her blog. One of her core scientific interests is endogenous retroviruses, which is one reason she is interested in this story. ERVs are not infectious - they can't make functional virus - but many of them are capable of making virus protein if they are turned on. Including among other things, gag protein. Some of these endogenous retroviruses are gamma-retroviruses. So - short version - our DNA is packed with ERV genes that can make retrovirus proteins if they get turned on. They are turned off by methylation. --------------- If a gene - say, an ERV or an integrated provirus from a new viral infection - is turned of my methylation it can be turned back on by demethylation. In her Ottowa presentation where JM showed 5AZA treatment of PBMCs, this is what she was talking about. She was claiming that after the initial infection, HGRVs integrate into the DNA of infected cells, and go silent She claimed that treatment with 5AZA reactivates them by demethylating them, they produce new virus, and then she can detect the virus. That's a reasonable hypothesis. ---------------- But here's the problem Treating the PBMCs with a demethylating agent - 5AZA - can also reactivate all those silent endogenous retroviruses, which are a part of our DNA, of everyone's DNA. Take perfectly healthy cells, demethylate them, and they can start to make gag protein FROM THEIR OWN DNA. 5AZA can cause gag to be made from either ERVs or from infectious viruses. In the Ottowa presentation, JM presented a gel from an experiment that Ruscetti did. She claimed the experiment showed that there were silenced HGRVs in infected cells, and that demethylation could make them start producing virus again. Assuming the labels on the slide she presented actually represent what was on the gel (they don't, she made that up too, but that's another story) they show patient samples that were treated with 5AZA, compared to controls that were not treated with 5AZA. The problem is, she compared CFS-patient, 5AZA treated cells, with healthy-control cells that wereNTo treated with 5AZA, and showed that there was gag protein in the CFS samples. But from that experiment, there is no way to know if she is detecting gag from recent provirus from a fresh HGRV infection, or gag from endogenous gamma-retrovirus sequences of the human DNA. That is, treating the patient cells with 5AZA is enough to make it produce gag from human DNA - even if it does not have a virus infection. ---------- Back to Figure 2c. Ruscetti has now admitted that the patient samples in Figure 2C were treated with 5AZA. That means that the gag they detected, could simply be from endogenous retrovirus sequences, including possible gamma-retrovirus - and have nothing to do with a virus infection. This is really basic virology. I know it as an out-of-date evolutionary geneticist, because the endogenous retrovirus story has been a fundamental story in evolution. Its been a fundamental story in virology,in cell biology, in genetics. Ruscetti and Mikowits had to of known this. Knowing that the patient samples in Figure 2c were treated with 5AZA, there is no way to tell if they are detecting XMRV, some other infections HGRV - or gag from ERVs. Just as bad, even if they are detecting an infectious HGRV, that gets turned on by 5AZA - there is no way to know if that virus might be in the controls as well, because they did not treat the controls with 5AZA. The 5AZA treatment was a major factor in that experiment. There is NO WAY that it was a meaningless trivial detail. Ruscetti and Mikowits had to of known it. When Ruscetti said it didn't matter, every scientist I know of who has followed this, was surprised or laughed out loud. What Figure 2C says, knowing what they did with 5AZA, is no more than - 'hey, we can find gag protein by doing things that are known to cause gag protein from healthy cells.' This is, at best, scientific misconduct in misrepresenting the experiment they did, and what was on Figure 2c. They may not be simply making up data - maybe they do have gels showing gag in patient samples and not in controls, without 5AZA treatment, as they claim in the Science paper. But we don't know -we can't know without someone going through their lab notebooks and results - because they didn't honestly and thoroughly tell us what they did. And at least some of what they did tell us of what they did, was not actually what they did -and they knew it when they told us. They knew that Figure 2C had 5AZA treated patient samples, and didn't tell us. They knew that lanes labeled claimed to be from patient samples in the Ottowa presentation, were actually healthy controls. Once a scientist has shown that they are willing to intentionally say something not true about their methods or their data, they cease to be scientists. They will no longer be trusted in science. It's a career-ender, and it makes all their work suspect, and it means that everyone who was relying on their work for their own research and research decisions cant trust the things they were relying on. Mikowits misled us about their methods and their results, and in the case of Figure 2C and the Ottowa slide, have said they did so. And THAT is why there is so much outrage from scientists about this.