Everything you always wanted to know about non-cytolytic enterovirus but were too afraid to ask

[chrisb]

No, I would be interested to see that. Was there a particular aspect of what is said that is relevant to the enterovirus idea?

"..they reported an outbreak of supposed poliomyelitis in LA which was not only more extensive but also milder than any previous outbreak of poliomyelitis in California. Peculiar features of the epidemic which Wilson and Walker noted were the unusual communicability of the disease, the early peak, in June, and the large number of adults affected. In children the disease followed the usual clinical course of poliomyelitis, but, amongst the adults, an atypical form of the disease was observed, in which sensory, vasomotor and arthritic symptoms predominated , and recurrent exacerbation of acute symptoms occurred over the following 12 months. out of 100 cases, 35 still had muscle weakness after a year.

Wilson and Walker suggested that the mildness of the attack, the marked degree of muscle weakness without proportionate atrophy and without loss of tendon reflexes, and the recurrences, all suggested that this was, at the very least, atypical poliomyelitis, rather than the usual form of acute anterior poliomyelitis..."

they considered and dismissed the idea that the symptoms in adults may have been functional.

I shall have to see ifI can sort myself out to copy more as she describes other cases which I don't think are generally referred to.

 

[Jonathan Edwards]

Wilson and Walker suggested that the mildness of the attack, the marked degree of muscle weakness without proportionate atrophy and without loss of tendon reflexes, and the recurrences, all suggested that this was, at the very least, atypical poliomyelitis, rather than the usual form of acute anterior poliomyelitis..."

This is interesting. They record 'marked degree of weakness without proportionate atrophy and without loss of tendon reflexes'. At least taken at face value that indicates a weakness not due to anterior horn cell damage and therefore not pathologically similar to polio at all. They talk of sensory and arthritis symptoms, which are not feature of polio.

They also mention 'supposed poliomyelitis', which presumably means that the virus was not identified. They say that the children had typical polio but unfortunately do not say what they mean by that.

I think it is hard to interpret what happened. Maybe there was a small outbreak of polio in children and at the same time a virus was producing an illness perhaps like ME (although that is far from clear) in adults. Maybe in this case polio was triggering a post viral fatigue. If EBV and Q fever can do it, why not polio? But polio has gone.

What may be of interest is that in the Royal Free outbreak, as far as I know, and also in other outbreaks from which there are still many survivors (post 1940 shall we say), as far as I know, there is no account of any cases of 'typical polio' in children at the time. Absolutely nobody had an illness suggesting anterior horn cell damage. And the whole idea of 'ME' as invented by Acheson seems to have been based on the mistaken belief that patients had paralysis that was similar to polio - due to anterior horn cell damage.

 

[chrisb]

After abreak for coffee I am sufficiently revived to continue:

These symptoms included muscle weakness, involuntary muscle contractions and twitching, clonic movements and cramps in the affected muscles, and muscular incoordination. However, sensory symptoms predominated, with pain in the back and extremities which was often intractable, excruciating and persistent for weeks or even months, and which only responded to opiates. There was also local tenderness of muscles, hyperaesthesia, parasthaesia and areas of anasthaesia, sometimes following the distribution of a nerve trunk and sometimes involving a whole extremity. Vasomotor and trophic disturbances were almost constant findings. There was excessive sweating or abnormal dryness of the skin, coldness, cyanosis and brittle nails. The authors wrote "It was the impression of many observers that a generalised disturbance of vasomotor control occurred in these patients, which best explained the emotional instability. Inflammation of the joints occurred in a third of adult cases, with a third of these being transitory and the remaining two thirds developing permanent arthritic changes in the joints.

 

[Jonathan Edwards]

These symptoms included muscle weakness, involuntary muscle contractions and twitching, clonic movements and cramps in the affected muscles, and muscular incoordination. ...

OK, that long list is pretty much uninterpretable. Maybe there was an outbreak of parvovirus or some other virus that is long since dead. Or maybe this was some form of post-viral syndrome, of which there are all sorts of varieties. It does not sound in the least like polio.

 

[chrisb]

I think that that was what Jenkins said. She thought that it was considered as a typical polio because it was seen with a polio epidemic and by doctors on an orthopaedic ward, and the Royal Free was seen as atypical glandular fever, but the illnesses were alike.

ETA for some reason I did not spot the reference for that paper which is:

Wilson JC and Walker PJ Acute anterior poliomyelitis. Orthopaedic aspects of the California epidemics of 1934
Archives of Internal Medicine, 57,477-492

 

[Guest 102]

But I do agree that there should be a clear distinction made between hypothesis and incontrovertible biomedical fact.

.

Absolutely. And because of the wilful lack of biomedical research, so much about ME *is* still hypothetical. We hope as more robust research is conducted that some of these hypotheses will eventually pan out and give us answers and desperately needed treatments.

There is no doubt that I got ill after a vicious Coxsackie B4 virus - an enterovirus - but I do not personally think the virus persists, I think my ME was more an aftermath of an abnormal response - or an unlucky response - to the Coxsackie virus. I also had a flu-like illness in France, about two months later, and at that time I did not know I was also infected by Coxsackie (picked up in Scotland, was diagnosed on return).

So I had had one virus on top of another. I was very ill but living abroad, only eighteen and trying to fight all the hellish symptoms, in total ignorance of having Coxsackie. Why did this evolve into ME? No one knows. Many others develop ME but not after an enterovirus. Ramsay explicitly said ME was caused by different viruses, though enteroviruses *seemed* to play a significant role.

My point really is that we need to be very careful on MEPedia, not to over-egg hypotheses as fact. I already expressed, for example, my concern about MCAD being described as a ‘common comorbidity’ - where is the research to back this up?

 

[Graham]

There is no guarantee that even 'experts' are in fat expert

Now come on @Jonathan Edwards , in fact I never accused any of the experts of being overweight! I did though both understand and agree with your point.

The protocol is not to prove someone else's work was wrong or misleading, but simply to replace it by new work that says something more interesting.

But it is this aspect more than anything that has destroyed so much trust. The failure to stand up and say that PACE and other similar studies are rubbish, accepting all the consequences that follow in the way in which patients are treated, is a major criticism of the system. The argument seems to be that current treatment, even if known to be wrong and potentially harmful, must not be removed until there is sufficient evidence to come up with new treatment. We must avoid hurting researchers' sensibilities at all costs?

It may well be that a small collection of specialists and researchers know that PACE is rubbish, but as this is not communicated to the general medical profession, let alone the public, it is highly damaging.

I'm glad you don't stand on the sidelines!

 

[Hip]

but I do not personally think the virus persists

It would not be possible to say in an individual case whether an persistent enterovirus is involved; but generally-speaking, the studies show that that persistent enterovirus exists in ME/CFS. These were studies done in the UK in the late 1980s and 1990s using muscle biopsies. There have been a couple of negative muscle biopsy studies, but these were small scale studies of only around 30 patients. The majority of larger studies have shown the presence of persistent enterovirus.

I also set up a MEpedia page listing all the enterovirus studies I could find, both positive and negative (that page is based on an old PR post on the same subject).

I think my ME was more an aftermath of an abnormal response - or an unlucky response - to the Coxsackie virus.

That is kind of part of the persistent enterovirus picture. The speculation is that it requires a high acute infection in order to seed enough cells with enterovirus, before a persistent infection emerges (via mutations in the viral genome). So presumably if immune response was weak during the initial acute infection, such that it led to a higher than normal acute infection, those may be the conditions for creating a persistent enterovirus.

This is likely why most people who catch coxsackievirus B do not get ME/CFS or dilated cardiomyopathy: because when they first caught the virus, it was met by a robust immune response, so that the virus was not able to seed sufficient cells in the body to create a persistent infection.

 

[Jonathan Edwards]

But it is this aspect more than anything that has destroyed so much trust. The failure to stand up and say that PACE and other similar studies are rubbish, accepting all the consequences that follow in the way in which patients are treated, is a major criticism of the system.

Indeed. I was never happy with not being able to criticise. I got a reputation for being predatory standing up at meetings and posing queries. I was banned from all grant giving committees for not being 'pragmatic' enough. I paid a high price. But I do not regret it. I beat them at their own game.

Now I am retired I can sound off like one of those 'troublesome' ageing backbench MPs of the old days. Whether it does any good or not I don't know.

 

[ScottTriGuy]

@Hip

If EVs mutate, would this mean that a patient would need to take multiple anti-EV meds to be effective, similar to how HIV+ patients need to take 3 (or more) ARVs to keep HIV suppressed?

In other words, would EVs mutate 'around' a single anti-EV med, rendering it ineffective? (like with the first HIV med, AZT, that initially caused many patients to improve...for a few months and then they got sick again as the HIV mutated to evade AZT. That's how the 3 drug 'cocktail' emerged.)

 

[Hip]

When I first started out in research I thought people should publish findings that negate the misleading studies in the field. I thought we should debate in the literature why interpretations were wrong. But I rapidly learned that this is considered bad form in science. The protocol is not to prove someone else's work was wrong or misleading, but simply to replace it by new work that says something more interesting.

Poor science does get torn to pieces in journal clubs in academic units. That is where we learn what is good and what is bad. All clinical trainees are exposed to that sort of critique every week for five years or more. But it is never mentioned in the literature.

Yes, I wish people would publish a summary of why a line of research no longer appeared valid, if just for the historical record.

 

[Hip]

If EVs mutate, would this mean that a patient would need to take multiple anti-EV meds to be effective, similar to how HIV+ patients need to take 3 (or more) ARVs to keep HIV suppressed?

In other words, would EVs mutate 'around' a single anti-EV med, rendering it ineffective? (like with the first HIV med, AZT, that initially caused many patients to improve...for a few months and then they got sick again as the HIV mutated to evade AZT. That's how the 3 drug 'cocktail' emerged.)

Because enteroviruses are RNA-based viruses, which mutate more quickly than DNA-based viruses, enteroviruses do have a higher mutation rate. However, such ongoing mutation and evolution of the virus is not really the crux of the matter when it comes to these persistent enterovirus infections.

The crux of the matter is a very specific type of mutation that occurs right at the beginning of the infection, a mutation manifesting actually during the acute stage of enterovirus infection (which is the first few weeks of infection).

Once that mutation occurs in the very early phase, it then sets the stage for a chronic persistent infection which can remain in the body for years. So it's a one-off particular type of mutation right at the start that causes all the problems.


Normally enterovirus is simply not capable of forming persistent infections. In general, enterovirus is completely eliminated by the adaptive immune response during the acute phase. For example, if you catch a cold from a rhinovirus (which is a type of enterovirus), then after some days your immune system will completely clear that virus from your body. That is the norm with enterovirus.

But when that specific type of mutation occurs during the acute phase of certain enterovirus infections, it changes the capabilities of the virus, and produces a virus that is now able to evade the immune system, and form persistent infections.


Not all enteroviruses can undergo this type of mutation. It's only enteroviruses from the enterovirus B species that can get mutated in a way that ends up creating a persistent virus. Enterovirus B serotypes include coxsackievirus B and echovirus.

That's why chronic coxsackievirus B and echovirus infections are found in ME/CFS.

 

[ScottTriGuy]

Excellent clarification @Hip thank you.

 

[Hip]

But the 2017 paper says the RNA cannot replicate, doesn't it?

The small deletions that occur at the terminal end of the enterovirus genome don't entirely block viral RNA replication, they just greatly reduce the replication rate by a factor of around 100.

With this much reduced replication rate, the virus can no longer produce enough viral particles, so the normal virus lifecycle (ie, making lots of viral particles which then destroy the cell by lysis as they burst out of the cell) is interrupted. This is why these non-cytolytic enteroviruses with deletions do not kill the cells they live in.

If they killed the cells they lived in, they would not be able to create a persistent infection, because they would have nowhere to live.

 

[Hip]

@Hip I was wandering if you could fill in my peasant tier knowledge on the immune system? In your article you say that when the 5th primed from the end of the RNA gets deleted it allows the virus to avoid immune detection, why?

That is the million dollar question.

Non-cytolytic enteroviruses replicate very slowly, so it's not as if these viruses are outgunning the immune system by their speed of replication. It's a very slow infection, so in principle you would think that the immune system should have no problem in clearing it.

But the fact that it doesn't clear it suggests this form of enterovirus must have some way of evading the immune response.

The original idea about how non-cytolytic enterovirus evades the immune response was proposed by Tam and Messner in their 1999 paper. They suggested that like two sides of a zip, the positive and negative single stranded viral RNA in the cell join together to make double stranded RNA (dsRNA), and this dsRNA is thought to be more resistant to immune attack.

So that's one idea about how non-cytolytic enterovirus evades the immune response.



But another more recent speculative idea (which sounds plausible to me) was proposed by Lévêque et al in their 2017 paper: they suggest that the genome deletions themselves might make non-cytolytic enterovirus resistant to the type 1 interferon immune response.

When type 1 interferons (such as interferon alpha) are secreted, they activate various immune activities inside the cell. One of these activities is the release of a protein called Ifit1.

The job of Ifit1 is to bind to the viral RNA in the cell, and disable that RNA. However, it is known that in the case of alphaviruses — which have deletions in exactly the same part of the genome non-cytolytic enterovirus — that these deletions prevent Ifit1 from binding to the viral RNA. This is simply because the part of the RNA that Ifit1 binds to is actually the deleted part.

So because the very spot that Ifit1 binds to on the RNA is deleted and no longer present, Ifit1 can no longer attach to and disable the viral RNA. This is known to occur in alphaviruses, and this is a mechanism that alphaviruses use to evade the immune response. So Lévêque et al speculate it might also explain why non-cytolytic enterovirus RNA appears able to resist the immune response.

 

[Jonathan Edwards]

The small deletions that occur at the terminal end of the enterovirus genome don't entirely block viral RNA replication, they just greatly reduce the replication rate by a factor of around 100.

That didn't seem to be the conclusion in that 2017 paper at least as far asI managed to read. Do we have documented cases of only mutant and not wild type RNA being extractable from clinically relevant human tissue in vivo? Cell culture studies look too artifactual to be much use to me.

 

[Hip]

Do we have documented cases of only mutant and not wild type RNA being extractable from clinically relevant human tissue in vivo?

Yes, in the Chapman and Tracy et al 2005 paper, they isolated persistent coxsackievirus B3 from the hearts of mice that had been infected with this virus one year earlier. They detected mutations in the virus: the small deletions in the 5′ terminal region of the viral genome. That was the first time these deletions were demonstrated.

Then Chapman and Tracy et al 2008, they found the same genome deletions in heart tissues from a fatal human case of coxsackievirus B2 myocarditis.

Lévêque et al 2012 found the these 5′ region deletions in cardiac biopsy samples of 4 of 20 patients suffering from idiopathic dilated cardiomyopathy.

Bouin et al 2016 also found these same deletions in a cardiac biopsy from a patient with dilated cardiomyopathy.



I find all these papers too dense with virological details to understand. Nora Chapman gave an easier to understand presentation about her non-cytolytic enterovirus research at the 2010 Invest in ME London conference. She also gave a very similar 15 minute presentation at the International Symposium on Viruses in CFS, Baltimore 2008, which you can see on YouTube here.

 

[Jonathan Edwards]

That is not a very extensive literature, @Hip. I would discount mouse studies because the issue is actual occurrence of this phenomenon in people. The first case you quote seems to be a fulminating fatal myocarditis. That does not sound like chronic non-cytolytic infection. It looks as if the virus was mutant - but presumably the infection was cytolytic if the heart was destroyed. That does not seem to fit. The others have found the same mutation in a handful of chronic cases.

All this seems to show to me is that sometimes Coxsackie virus exists in a mutant form with a deletion. It also seems to persist, which I think we knew all along. I am not impressed that one should hang a new concept like 'non-cytolytic infection on a handful of case reports that are pretty hard to interpret. This looks to me very much like some enthusiasts who have got stuck studying something that was of interest thirty years ago trying to scrape together a few observations to justify continuing to study it.

 

[Hip]

All this seems to show to me is that sometimes Coxsackie virus exists in a mutant form with a deletion. It also seems to persist, which I think we knew all along. I am not impressed that one should hang a new concept like 'non-cytolytic infection on a handful of case reports that are pretty hard to interpret.

The purpose of these studies which detected the deletions in the 5′ non-translated region of the virus is not to demonstrate that non-cytolytic / non-cytopathic enterovirus exists, which as you say, we already knew. There does not appear to be much doubt that chronic non-cytolytic enterovirus infections exist, as these infections are routinely found in studies and clinical work on chronic myocarditis, dilated cardiomyopathy and ME/CFS (these persistent human tissue infections do not lyse or destroy cells when transferred to a cell line, hence are called non-cytolytic / non-cytopathic).

Rather the purpose of the genome deletion studies is to gather evidence supporting a theory on how enterovirus can persist. As mentioned, enteroviruses are normally only capable of acute infections which get eliminated the the adaptive immune response. So the fact that we see enterovirus causing persistent infections constantly producing viral proteins needs explaining; the mechanism of persistence needs to be brought to light.

Nora Chapman in the 2005 study proposed a mechanism, which she explains in the video, and it is also explained in the MEpedia article. That mechanism details how these deletions in the 5′ non-translated region at the end of the genome alter the virus, and confer the virus with the ability to form persistent infection — an ability it did not have before these mutations occurred. From the literature I have read, her explanation of the mechanism of enteroviral persistence is generally accepted.

What is not generally accepted is the idea that these chronic enterovirus infections cause the diseases they are found in; but that's to be expected, as we all know association does not imply causation.

This understanding of the mechanism of persistence will likely have significance for pharmaceutical companies who wish to develop antiviral drugs which can specifically target persistent enterovirus infections. Dilated cardiomyopathy accounts for around 45% of all heart transplants, so if this condition could be treated instead with a simple antiviral drug, then that would be a big advance. Not to mention what such a drug might do for ME/CFS patients.

 

[Inara]

I think what struck me most was the confidence with which the medical profession in general made assertive statements about what could and could not be the case

That's not restricted to medicine. Think about Maxwell's theory or the quantum theory (Einstein simply wouldn't think about thinking about it; now, the quantum theory still is a theory and will be adjusted I think, but that's how it goes).