Enterovirus Followup Study?

[energyoverload]

Does anyone know when and if there will be a followup study to try and replicate John Chia's work? I seem to remember Enlander mentioning something about this some time ago, but I'm not sure. Anyone?

 

[Seven7]

I tested positive for Coasaxie b2 in serum general GP run test. I got ok and got it back a second time (I could tell because gives me really bad Acid reflux) but I am clean now using his equilibrant.

 

[halcyon]

Does anyone know when and if there will be a followup study to try and replicate John Chia's work? I seem to remember Enlander mentioning something about this some time ago, but I'm not sure. Anyone?

I'm not aware of anything concrete at this point. A fellow patient reported that Dr. Chia told him the CDC had requested some of his samples for further study but I haven't seen anything further about that since.

 

[aimossy]

The IOM report mentions something about Chia and the enterovirus work. I think it said something like was unable to be replicated. Dont quote me but something like this was in there.

 

[halcyon]

The IOM report mentions something about Chia and the enterovirus work. I think it said something like was unable to be replicated. Dont quote me but something like this was in there.

It's on page 160:

Enterovirus A few studies focused on the role of persistent enterovirus
infection in ME/CFS (Chia et al., 2010; Galbraith et al., 1995; Lane et al.,
2003). Chia and Chia (2008) showed that in a subset of ME/CFS patients
who reported significant gastrointestinal complaints, the prevalence of
enterovirus infection as demonstrated in stomach biopsy samples was significantly
higher compared with control subjects. Other investigators failed
to reproduce an increased incidence of enterovirus infection in ME/CFS
patients (Lindh et al., 1996).

One negative study on a total of 34 patients, not too convincing. They also used a number of methods of detection that are prone to error.

 

[Hip]

One negative study on a total of 34 patients, not too convincing. They also used a number of methods of detection that are prone to error.

Enteroviruses were routinely found in ME/CFS patients in the early British research performed in the 1980s and 1990s. However, in a video presentation by Dr Chia, he said that when American researchers at the time tried to replicate the British work, they could not find enteroviruses in their ME/CFS patients.

I am not sure if that failure to replicate was because the US researchers used different viral testing methods, or used different ME/CFS inclusion criteria, or whether at that time there may have been an epidemic of enterovirus infections in the UK, but not the US (so that in the US in that era, ME/CFS cases might have been caused by a different virus, like EBV for example).

I have not actually looked at these US studies that were unable to reproduce the results obtained by the British researchers.


Anyway, Dr Chia's seminal work in the mid 2000s finally replicated, and advanced, the early work from the UK. Though it took 20 years for this to happen. So it is a very slow pace.

How long will it take before Chia's own excellent work is replicated? Well, it is now already 10 years since Chia published his enterovirus studies. So I really hope that someone takes up the challenge soon.

I understand that Dr Chia himself is very saddened by the fact that nobody has taken any interest in reproducing his work. And for me as an ME/CFS patient, I am deeply frustrated by this also.



However, not everything in the enterovirus world has come to a complete standstill. The same sort of chronic smoldering enterovirus infections that you find in ME/CFS patients have also been observed in other diseases: chronic coxsackievirus B myocarditis, dilated cardiomyopathy, and possibly in type 1 diabetes.

Research into the possible enteroviral basis of these diseases is ongoing, albeit a bit slowly, and any advances and treatments developed for these diseases may well be applicable to ME/CFS.

Some further information on the enterovirus research on these diseases is to be found here:

Human Enteroviruses and Chronic Infectious Disease. Steven Tracy and Nora M. Chapman

Replication Defective Enterovirus Infections: Implications for Type I Diabetes

It would be particularly pleasing if they can obtain good evidence to support the theory that chronic enterovirus infection causes type 1 diabetes. If these researchers can get this evidence, I think research into chronic enterovirus infections will explode, because type 1 diabetes is a very prevalent and costly disease, so if it can be cured by effective anti-enteroviral drugs, I am pretty sure those drugs will soon be developed.

 

[halcyon]

I have not actually looked at these failed US replication studies.

The failed study noted in the IOM report was from Sweden. He talks about it briefly in this paper:

Several other studies have failed to demonstrate amplifiable enteroviral RNA in blood or in other tissue,24,25 although two recent papers again demonstrated the presence of enteroviral RNA in muscle biopsies of patients with CFS and not in the controls.26,27 The discrepancy between the various studies is not well explained, but could result from differences in specimen collection and RNA preservation, the time to processing of the biopsies or blood samples, the primers used, and the sensitivity of the assay.

He also touches on it in one of his talks. I forget the exact details, but he talks about the difficulties in getting the primers right for PCR testing.

 

[Never Give Up]

In the fall Dr. Chia told us that several virology researchers had requested some of his samples and that he chose a lab at Cornell University. Later in the year he told us that the CDC was going to look at his samples as well. He asked for permission to send my son's biopsy samples to the CDC and we agreed. The next time we saw him he said that the CDC had his samples, but was putting his project on hold as enterovirus D68 was becoming a more urgent problem. He was fine with this, saying that any entro virus research was good for ME/CFS patients.

 

[Hip]

The failed study noted in the IOM report was from Sweden. He talks about it briefly in this paper:

OK.

In the Invest in ME London 2010 ME/CFS Conference Dr John Chia said this:

I looked at the papers done by the British investigators I found these to be really interesting. Unless they blatantly lied about their findings, I can't believe why these were denied in the first place.

...

If I quickly review some of the literature that the British investigators generated. This slide is important, because there was a 30 year old female that developed ME/CFS for about five years, and committed suicide. Before she died and was . . . the family gave doctors the chance to find the virus in the tissues.

So they were able to find enterovirus RNA in the muscles, heart, hypothalamus, and in the brainstem, and the RNA sequence shows about 83% homology to what we call Coxsackie B3. That is only one patient, published 1994. That has not been done again. But you don't get a chance like this very often. But this clearly shows the virus can go to the various areas where patients are complaining about symptoms.

The other study that needs to be quoted is this blinded control studies done by Clements, et al, and they were able to find enterovirus RNA in the CF patient's serum, about 41% and the control serum was only 2%.

Unfortunately our American investigators could not reproduce this finding. So basically this was left, in 1995, as probably a disease of the mind. And because of some the special comments made, but I'm not going to go over that issue right now.

Video Timecode: 12:31.

When Chia says in the quote above that "our American investigators could not reproduce this finding", I am not sure which papers he is referring to.

It's interesting that in the Clements, et al 1995 study Chia mentions in the quote above, they used the Oxford criteria, and even with those rather imprecise criteria, they still found enterovirus RNA in about 41% of ME/CFS patients, and in 2% of controls.



I do find the postmortem brain autopsy studies particularly interesting:

Enterovirus in the chronic fatigue syndrome (full text here)
McGarry F, Gow J, Behan PO. Ann Intern Med. 1994 Jun 1;120(11):972-3.
➤ This study details an autopsy of a deceased ME/CFS patient. Enteroviral RNA was found in the heart, muscles, hypothalamus and brainstem of this patient, and this RNA showed an 83% similarity to coxsackievirus B3. Control tissue samples taken from four patients who died of cerebrovascular diseases, and another four who had depression and committed suicide, showed no evidence of enteroviral RNA.

Viral Isolation from Brain in Myalgic Encephalomyelitis (A Case Report)
Richardson, J. Journal of Chronic Fatigue Syndrome, Vol. 9(3/4) 2001, pp. 15-19.
➤ This study examined the brain of an ME patient who died through suicide, and found enteroviral VP1 protein in the fibroblasts of small vessels in the cerebral cortex, plus some patchy distribution of enteroviral VP1 protein in a small fraction of glial cells.

In the latter brain autopsy study, the fact that they found that the glial cells of the brain were infected is extremely interesting. Most glial cells are astrocytes, and astrocytes are intimately involved in brain inflammation.

It seems to me that the enterovirus infection of the brain astrocytes in this patient may be directly responsible for the continued brain inflammation found in ME/CFS.



I wish more ME/CFS postmortem brain studies were conducted, looking for enterovirus RNA or enterovirus VP1 protein in the astrocyte cells.

The enterovirus coxsackievirus B is able to enter and infect astrocytes because these cells possess a coxsackievirus and adenovirus receptor (CAR) on their surface. Coxsackievirus B can use the CAR receptor to enter a cell.

What's more, in a murine model it has been shown that when astrocytes are infected with coxsackievirus B, they release the pro-inflammatory cytokines IL-1β, TNF-α and IL-6. Now it just so happens that these 3 pro-inflammatory cytokines are precisely those which cause sickness behavior, and sickness behavior strongly resembles many symptoms of ME/CFS.

You will recall that Michael VanElzakker hypothesizes that ME/CFS is caused by these 3 sickness behavior cytokines when they are secreted by an infection of the vagus nerve. Well, a very similar hypothesis might be that that ME/CFS is caused when these 3 sickness behavior cytokines are secreted by coxsackievirus B-infected astrocytes in the brain.

 

[halcyon]

Later in the year he told us that the CDC was going to look at his samples as well. He asked for permission to send my son's biopsy samples to the CDC and we agreed. The next time we saw him he said that the CDC had his samples, but was putting his project on hold as enterovirus D68 was becoming a more urgent problem.

It turns out there is a CDC CFS conference call coming up on Monday, including a Q&A segment. I will submit a question regarding the status of this.

 

[Hip]

It turns out there is a CDC CFS conference call coming up on Monday, including a Q&A segment. I will submit a question regarding the status of this.

It will be very interesting to hear what they say in response.

 

[Sidereal]

It's a travesty that no one's followed up on the early British enteroviral research and Chia's work. I am very worried about the current autoimmune direction of research which will probably end up leading us down the same blind alley as multiple sclerosis.

 

[halcyon]

In the Invest in ME London 2010 ME/CFS Conference DVD Dr John Chia said this:

Starting at time 14:30 in that talk is kind of what I was thinking about. What I was thinking of more was from this 2008 talk. At 12:44, he talks about the PCR test being difficult to get right.

When Chia says in the quote above that "our American investigators could not reproduce this finding", I am not sure which papers he is referring to.

Again, from the above 2008 talk, at 9:07 I believe this is the American study that could not reproduce the findings. This was apparently from unpublished work by Stephen Straus and H. Robart. At 15:02 he talks about the difference between how Robart did it and how he did it.

I believe all of these issues are what led him to using stomach biopsies as a much more reliable indicator of active infection.

 

[Hip]

That's a very interesting part of the video, @halcyon.

Just for the benefit of those here who don't have access to this video:

At timecode 14:30 in the 2010 Invest in ME video, Dr Chia says that sometimes in his blood tests on ME/CFS patients he would find enterovirus RNA, but on other occasions at a later date, even in the same patient, his tests would not find any virus. And then at a later date still, the test might once again find the virus in the blood of the same patient.

Dr Chia said this made him realize that there was actually very little enterovirus in the blood. And in fact he would later determine that there were, on average, just around 200 copies of the enterovirus RNA per ml of blood in his ME/CFS patients, which is a very low amount.

Dr Chia said this low amount of enterovirus in the blood probably explains the discrepancies between the studies which found enterovirus in ME/CFS patients, and the studies which did not — with such low amounts in the blood, it is quite easy for studies to miss these viruses during testing.

This understanding also made Dr Chia realize that he needed a better way of detecting enteroviruses in ME/CFS patients, and so this is what spurred him on to develop his pioneering technique of testing for enteroviruses not in the blood, but via a biopsy of the stomach tissues (his immunohistochemistry test).

 

[Gingergrrl]

@Hip & @halcyon in your opinion, is the ARUP testing for coxsackie and echo viruses still pretty accurate even though not a stomach biopsy? I am curious to hear your thoughts since you are both much more familiar with enteroviruses than I am.

 

[Hip]

@Hip & @halcyon in your opinion, is the ARUP testing for coxsackie and echo viruses still pretty accurate even though not a stomach biopsy? I am curious to hear your thoughts since you are both much more familiar with enteroviruses than I am.

Dr Chia considers the stomach biopsy to be the gold standard in terms of detection sensitivity.

However, the stomach biopsy cannot determine which specific serotypes of coxsackievirus B or echovirus are present in patients.

The ARUP Lab antibody blood tests do provide this specific serotype information: the two ARUP Lab tests encompass all 6 of the known coxsackievirus B serotypes (CVB1 to CVB6) and 5 out of the 32 known echoviruses. But the ARUP Lab tests cannot detect any enteroviruses (known or unknown) outside the range they test for.

Dr Chia says the stomach biopsy test detects most enteroviruses, known or as yet unknown (if I remember rightly, Dr Chia has pointed out that there may conceivably be unknown enteroviruses present in ME/CFS patients).



I don't know for sure, but I think this specific serotype information from ARUP Lab might be useful in terms of tailoring the antiviral treatment.

For example, Dr Chia found that the combination of interferon and ribavirin was effective for coxsackievirus B3 infections, but not effective for coxsackievirus B4 infections (he mentioned this in the Invest in ME 2009 ME/CFS Conference). However, I understand he rarely uses interferon these days, so I am not sure whether this specific serotype information is useful or not for tailoring the antiviral treatment.



So as you can see, there are pros and cons for the stomach biopsy test, and the ARUP Lab antibody tests.

The stomach biopsy test is relatively cheap. In this stomach biopsy requisition form from Chia's lab, the cost is stated as $250 (although that does not include the cost of the endoscopy procedure performed by a gastroenterologist to obtain a tissue sample from the stomach).

Whereas I believe the two Coxsackievirus B and echovirus antibody tests are something like $500 each.

 

[halcyon]

I don't know for sure, but I think this specific serotype information from ARUP Lab might be useful in terms of tailoring the antiviral treatment.

I believe this is true. For example, Dr. Chia said that echovirus 7 is completely unresponsive to oxymatrine. He's verified it in vitro.

 

[halcyon]

@Hip & @halcyon in your opinion, is the ARUP testing for coxsackie and echo viruses still pretty accurate even though not a stomach biopsy? I am curious to hear your thoughts since you are both much more familiar with enteroviruses than I am.

I believe the antibody tests are accurate. They are far from perfect, but they're a good place to start. They have a few problems:

1) The presence of elevated antibodies on a single test doesn't prove that the virus is present right now. It can only prove, indirectly, that it was present at some point recently. With that said, Dr. Chia has shown that antibody titer seems to correspond with the amount of virus and symptoms present. When treating the infection, symptoms improve and titer falls. If relapse occurs, the titer rises again.

2) There are over 100 known enteroviruses, but the antibody tests can only detect 11 of the more common ones. Dr. Chia has numerous clinical examples of patients with enterovirus positive stomach biopsies that test negative on the antibody panel. This means a negative on the panel can't rule out an enterovirus infection.

Elevated antibodies were the first piece of evidence Dr. Chia found but he knew it wouldn't convince anybody so he moved on to trying to find the virus in the blood with PCR. This isn't very reliable for the reasons listed above in the thread, so he moved on to looking in stomach tissue. As @Hip mentioned, this is pretty much the gold standard. It's quite easy to find virus there with at least 4 biopsy samples. The only downside is that it's a somewhat invasive procedure to obtain the samples.

From what I've seen, Dr. Chia is confident using only clinical symptoms and the antibody panel to diagnose and prescribe treatment.

 

[Gingergrrl]

Thanks again to both of you for all the additional info. I have only one high titer and will be repeating the test to see which direction it goes. I will not be doing a stomach biopsy or anything invasive but am curious to learn as much about this as I can. I really hope that Dr. Chia gets the support and funding to continue his research.

 

[Hip]

I believe this is true. For example, Dr. Chia said that echovirus 7 is completely unresponsive to oxymatrine. He's verified it in vitro.

That's very interesting. @halcyon, you always have these interesting nuggets of information! Where did you come across that one, may I ask?

Though is it possible to test in vitro (in cell lines) for the Th1/Th2 immunomodulatory effects of oxymatrine, because presumably you haven't got the full immune system present in cell lines? The Th1/Th2 immunomodulatory effects of oxymatrine may be the main way oxymatrine fights off enteroviral infections.