Author Archives: Amy Proal

The power of intracellular pathogens: how they “take down” the immune system and dysregulate human pathways

March 12th, 2018 by Amy Proal

A human white blood cell

Hey there! Last podcast I talked about how many “harmless” microbes in the human body are able to evolve to a state where they cause inflammation and illness. I explained that this “shifting” towards virulence (or disease) is much more likely to occur in people whose immune systems are compromised or debilitated. There are many factors that can negatively impact the health of the human immune system. I plan to discuss many of them in greater depth. But today I will focus on one major factor – the ability of key pathogens to slow and dysregulate the human immune system as part of their most basic survival strategies.

The most fundamental way that pathogens “take down” the human immune system is by infecting and living inside human cells – especially human white blood cells. Under conditions of health, these white blood cells traffic the body, where they engulf or digest pathogens that originate from both the internal and external environment. BUT, many key pathogens have evolved to thwart this process. They have evolved to remain alive inside these white blood cells; inside the very cells that are supposed to kill them! They can survive this way for long periods of time in a chronic, persistent state. Doing so clearly gives them a HUGE advantage or “edge” over their human host. If the situation was compared to a human war, it would be as if a soldier were able to inhabit the body of its opponent, and then make the opponent act in its own interest.

There are two main implications of this “intracellular persistence”:

1. The infected cell is unable to perform its basic immune system duties.
2. Any pathogen inside the infected cell can interfere with the way the cell “encodes” its human DNA.

Let me explain this second point in more depth. The center of every human cell has a copy of a person’s DNA. This DNA “codes” for the ~ 21,000 human genes that determine who we are and how we deal with our environment. These genes are eventually “translated” into proteins – proteins that direct our human pathways to correctly regulate the immune system, the nervous system, human metabolism and other critical aspects of human function. It follows that any pathogen able to persist in the center of a human cell can directly “mess with” the cell’s DNA and the vital human processes it regulates. The situation could be compared to a computer hacker. Imagine a hacker that not only breaks into your computer, but gains access to the programming code that directs your operating system and websites. By either directly altering this code, or changing the way the code is translated into programs, the hacker can cause your computer to malfunction in an almost infinite number of ways.

Here’s an example of how a pathogen can dysregulate a human cell in the above fashion. A research team in Louisiana studied what happens when Cytomegalovirus (CMV) infects a human white blood cell. They found that CMV significantly changed how the cell expressed its human genes. In fact, the virus “turned up” the expression of 583 human genes, and “turned down” the expression of 621 human genes. These changes were so profound that the infected white blood cells actually changed into an entirely different type of cell – an “inflammatory” cell that promoted CMV’s ability to infect neighboring cells. No wonder then that, as the paper points out, chronic CMV infection is associated with retinitis, gastrointestinal disease, hepatitis and pneumonia – but also with chronic inflammatory conditions like multiple sclerosis and atherosclerosis (heart disease).

Cytomegalovirus (CMV)

A recent study by researchers at Stanford confirms how broadly CMV can “re-program” the human immune response. The team studied identical twins and isolated cases in which one twin was infected with chronic CMV and the other was not. They tested the activity and function of 204 different parts of the human immune system in each twin. These 204 immune system “parameters” included the amount/frequency of certain immune cells, the ability of such cells to signal, and the concentrations of immune system proteins called cytokines. They found that, in general, 119 of these immune system “parameters” were altered in the twins with CMV. In fact they calculated that, in people infected with CMV, “the lifelong need” to control the virus causes approximately 10% of all T cells in the body to become directed against it. This led the team to conclude that the immune response is “very much shaped by the environment and most likely by the many different microbes an individual encounters in their lifetime.”

OK – but how exactly does this major pathogen interference happen? All human pathways that control the immune response, metabolism, cognition etc are regulated by receptors – protein molecules that control pathway signaling. In very simple terms, receptors function in a “lock and key” fashion. Imagine that you want to get into a very secure house. You might have to first open locks on an outside door. Then you might open or adjust a series of other locks on inner doors and/or other checkpoints before finally entering. Now, picture these locks as receptors associated with a human pathway. Under conditions of health, very specific human proteins bind into each receptor. These proteins are analogous to the keys in my house example. If the correct human protein or “key” fits into the correct human receptor or “lock,” the receptor/protein “complex” changes shape in a way that allows that part of the pathway to correctly move forward. This process happens over and over at each receptor “checkpoint” until the pathway fulfills its final, correct role in the body.

Now consider this: many pathogens have evolved to create proteins that are very similar in size/shape to human proteins (this redundancy is sometimes called molecular mimicry). If a pathogen with this ability survives inside a human cell, its OWN proteins may begin to “bind” into human receptors. Gradually, the human proteins meant to control a particular receptor may be increasingly replaced by these similar “enemy” or pathogen proteins. What happens? The slightly different shape of the pathogen proteins cause the receptor to act/signal differently than it would under conditions of health.

An excellent example of the above (that I’ve discussed in several published papers) is the ability of pathogens to dysregulate activity of the human Vitamin D Receptor (VDR). The VDR is an extremely important receptor located at the heart of every human cell. It controls the expression of almost 1000 human genes. It also controls the activity of very important parts of the human immune response. These include TLR2 (a protein the body uses to identify and target pathogens), and several kinds of antimicrobial peptides (natural antibiotic molecules the body uses against infectious threats).

It comes as little surprise then that the fungus Aspergillus fumigates has evolved to create a protein or gliotoxin that can bind into the VDR and directly slow its activity. This results in tremendous survival gains for Aspergillus – the altered VDR can no longer correctly express many of the human genes it regulates. The expression of TLR2 and the antimicrobial peptides under VDR control also diminishes. Now, Aspergillus can flourish in the face of the host’s weakened immune defenses.

In fact slowing VDR activity (and subsequently parts of the human immune system) is such a logical survival mechanism that many other key pathogens have also evolved ways to dysregulate the receptor. When Epstein Barr virus infects human B cells it dramatically lowers VDR activity. Mycobacterium tuberculosis slows VDR activity by a factor of ~3.3. Borrelia burgdorferi, Cytomegalovirus, and Mycobacterium leprae also slow VDR activity to varying degrees. HIV almost completely “shuts down” the VDR. I should be clear that some of these pathogens don’t create proteins that directly bind into the VDR. They interfere with the VDR receptor in ways a little too hard to explain here:) But you get the overall trend right!?

Now factor this in: the human body’s pathways are tightly interconnected. So any change in human signaling has “flow-on” effects that impact yet other pathways and systems. In the case of the VDR, the receptor’s activity is tied to that of related receptors that control hormone signaling (especially the estrogen-beta receptor). It follows that the “original” intracellular infection may also prevent these related receptors/pathways from functioning correctly.

But let’s go back to the immune system, and continue to use the VDR pathway as an example. Let’s say CMV infects a person’s B cells and lowers VDR activity. The person’s immune response suffers as a result. Now, it becomes easier for that person to pick up OTHER pathogens from the external environment. Or, pathogens already in that person’s microbiome find it easier to act in a more virulent fashion. These “new” pathogens may also infect the person’s cells, where the interference they cause further slows the immune response. A “snowball effect” may ensue, where each “new” pathogen begins to facilitate the survival of the next.

A successive infection “snowball” could drive many forms of disease

In many of my papers I’ve referred to this snowball effect as “successive infection.” When I think of successive infection I literally picture a snowball rolling down a hill. The snowball begins with one intracellular infection, and then grows, as more and more pathogens take advantage of the dysregulated atmosphere in the body. As more pathogens “join” the snowball, the disease state that the snowball represents pushes the human body farther and farther away from a state of health.

It’s important to realize that once the “successive infection” process has started, environmental factors besides infection can “add” to the snowball. These include chemical exposures, stress, immunosuppressant drugs, over-used antibiotics and any other factors that cause either the immune response or microbiome balance to suffer. In fact, any of these other factors can “start” the snowball too.

I realize this is a major claim, but in my mind, a successive infection snowball could drive almost any form of inflammatory disease. That’s because such wide-range of different pathogens and negative environmental exposures can contribute to the process. Pathogens are also able to infect so many parts of the human body (nerves, brain tissue, lung tissue). If the snowball represents disease, the specific illness any one person develops would stem from their UNIQUE infectious history, their unique medication history, their unique chemical exposure history, and the many, many different ways these and other variables can interact with/feed into each other.

If you think that’s a stretch, consider this: almost every analysis of the human microbiome identifies new microbes never before detected by previous studies. For example, just a few months ago, Steven Quake at Stanford identified thousands of “new” microbes in human tissue and blood. Many of them are potential intracellular pathogens we didn’t even realize could survive in humans. This led Quake and team to state that these novel microbes “may prove to be the cause of acute or chronic diseases that, to date, have unknown etiology…”

Also, research teams continue to discover new examples of how pathogens might interfere with human pathways. For example, a team in Indiana just identified viruses that create proteins similar in size/shape to the human protein insulin. They found these viral proteins could bind into human insulin receptors that determine how the body correctly regulates blood sugar levels. The viruses that create these insulin-like proteins are regularly detected in human blood and fecal samples. The team concluded that “since only 2% of (human) viruses have been sequenced, this study raises the potential for discovery of other viral hormones which, along with known virally encoded growth factors, may modify human health and disease.”

So I’ll end on the following note: never underestimate the “power” of intracellular infection. Human microbiome studies that fail to account for possible intracellular pathogens may miss part of a larger, more important picture of disease. For example, the gut microbiome dysbiosis now tied to many inflammatory conditions may be a downstream result of intracellular infection in the blood or other tissues. AKA: the widespread drop in immune system activity driven by intracellular pathogens may “set the stage” for an atmosphere that favors gut microbiome imbalance. This is certainly the case in HIV/AIDS, for example (for more context read this blog post).

Ok phew! That’s it for now. Next podcast I’ll go more into immunosuppressive drugs, antibiotics, and other factors that can also compromise the human immune system and/or the human microbiome. Talk then!

Towards a more nuanced view of the human microbiome: pathogens and evolution

March 1st, 2018 by Amy Proal

Vast microbiome communities persist in human tissue and blood (Image:

Podcast transcript:

People often ask me for a quick description of the human microbiome. I appreciate their interest, although it’s actually a difficult request: the microbiome is extensive and capable of vast range of behavior. Nevertheless I’ll usually say something like this:

“When I refer to human microbiome I’m talking about the trillions of bacteria, viruses, fungi, bacteriophages and other microrganisms now understood to persist in complex communities in all human tissue and blood.”

But beyond that: how do I best summarize microbiome ACTIVITY? Aka: What are these microbes in and on us are doing?? I’ve learned that most friends expect a response along these lines:

“The microbes in and on us are our friends. They work night and day to defend us from disease, improve our metabolism, break down our food, control our digestion, educate our immune systems and beyond.” In such a description, all the microbes we harbor are portrayed as allies or even heroes: organisms that continually devise ways to benefit or support our health and well-being.

This kind of description is often accompanied by a specific mindset: the belief that microbiome pathogens (those that cause disease) are of secondary importance to those that “help” us. And that to strongly focus on the presence of these myriad pathogens is to give microbes in general an overly “bad reputation.”

In my opinion, this portrayal the microbiome is too one-sided: At the very least it should be accompanied by an asterisk stating something like: “this is an attempt capture microbiome behavior under conditions of extreme health.” And it’s true, of course, that people with extensive microbiomes (take Olympians!) can function with incredible vigor and ability.

But the vast majority of us sustain a range of minor to major inflammatory health issues. Some people deal with acne, others have trouble tolerating or digesting certain foods. Insomnia, anxiety, metabolic problems…are all extremely common in today’s human population. Beyond that, we are living in the midst of an epidemic of chronic disease, in which the incidence of nearly every well-characterized inflammatory condition is on the rise (diabetes, obesity, heart disease, the “autoimmune diseases” to name just a few). In fact, an estimated 60% of Americans suffer from at least one chronic condition (and that number only includes people who have been officially diagnosed).

A growing number of these inflammatory conditions/symptoms are tied to microbiome dysbiosis: cases in which a microbiome community shifts towards a state of imbalance. This dysbiosis can be severe: pathogens tend to outcompete their more symbiotic neighbors, leading to drops in community diversity that promote an inflammatory environment.

This means that in the vast majority of people, the microbiome is acting in a nuanced fashion. Some of the microbes we harbor may perform health-related functions, BUT a significant number of others may be acting as pathogens. In fact, certain pathogens are so effective at promoting chronic symptoms that they can “push” the microbes around them towards a dysfunctional state. These pathogens are sometimes called “keystone pathogens.” A good way to remember that is that the activity and gene expression of keystone pathogens are KEY to promoting disease.

In my opinion then, it is vital that we continually recognize and encourage the general public to understand that persistent, chronic, pathogens can also play a central role in any microbiome community. To do so is not giving microbes a “bad reputation.” It is simply accurate. In fact, the current, alarming epidemic of chronic disease begs that we study these persistent pathogens with more urgency than ever before.

It also helps to interpret microbiome activity via the lens of evolutionary biology. When microbes are described as “friendly” people often assume that there is actual intent on their part to help us. In reality, any symbiotic or “commensal” microbe/human relationship is still driven by the most basic tenants of evolution: “survival of the fittest” and “reproductive fitness.” The goal of any microbe in/on our bodies is to stay alive and pass its genes (encoding its particular traits and characteristics) to the next generation. So if a microbe is, for example, breaking down our food, it’s because doing so also benefits its own ability to survive and reproduce. The fact that such behavior also helps its human host is simply an evolutionary coincidence.

In most ecosystems on earth (like the rainforest), commensal relationships are actually rare. This is partly because one species in a commensal relationship has only to slightly change its gene expression and/or metabolism for the relationship to shift, and begin to favor one species over the other. This “shifting” happens very easily with microbes because they can quickly adjust their gene expression patterns: they turn certain genes “on” or “off” in order to functional optimally under different environmental conditions. Microbes can also – through a process known as “gene transfer” – acquire completely new genes: genes that allow them to function in totally new ways. And, as with most organisms, a chance mutation in a microbe’s genetic code (their DNA or RNA) can endow it with yet other novel traits and abilities.

Take E.coli. Strains of E.coli are detected in almost every analysis of the human microbiome, even in studies conducted on healthy people. This makes sense, since people digest seemingly innocuous E.coli strains on a regular basis. But most of us are also familiar with E.coli’s ability to cause severe food poisoning and other forms of illness. This range of E.coli behavior is due to the fact that it can evolve to persist in many, many different states – some WAY more pathogenic (or able to cause disease) than others. Take a study done by researchers in Portugal. The team found that when “commensal” E.coli were exposed to human white blood cells, they rapidly mutated and changed their gene expression: the commensal E. coli evolved, in less than 500 generations, into virulent, or disease-causing, clones. When these virulent clones were injected into mice, the mice suffered an increase in inflammatory symptoms.

This “virulence”, or ability of a microbe to cause disease, is dictated in large part by the activity of genes that express “virulence factors”: molecules that help bacteria, viruses and other microbes better persist inside the cells of the immune system, better penetrate host tissue, better obtain key nutrients from the host, better produce toxins…and better engage in many other behaviors that cause human health to suffer.

E.coli as seen under a microscope.

Every microbiome community (even in healthy individuals) harbors microbes capable of expressing these virulence factors. These include potential pathogens present in the earliest microbiome communities that an infant inherits from its mother (microbes generally acquired via the placenta, vaginal canal, and/or breast milk.) In other words, every human is “seeded’ with potential pathogens from the beginning of life.

It follows that pathogens capable of expressing virulence factors can evolve and “emerge” from microbes already present in a “healthy” microbiome community. Let me give you an example. Researchers in England recently published a paper called “Severe infections emerge from commensal bacteria by adaptive evolution.” The team studied the bacterial species S.aureus. Like E.coli, S.aureus is frequently detected in the nasal (nose) microbiome of healthy individuals. However, S.aureus can mutate and change its gene expression so profoundly that it can also cause life-threatening infections.The English research team found that these life-threatening strains of S.aureus could easily descend from commensal S.aureus bacteria already colonizing a person’s nose. In other words, depending on conditions in the body, S.aureus is able to mutate from a harmless microbe to a major pathogen within the same person. This progression towards virulence involved an increase in the activity of genes that promote toxin production, abscess formation, immune evasion and many other processes that harm the human host.

Also key to the study’s findings was that, in the majority of cases, this within-person evolution towards S.aureus virulence occurred in subjects that had “risk factors for infection”: these included a weakened immune response, co-morbid conditions (the subject was sick with one or more inflammatory diseases), and/or medical interventions (subjects had undergone surgery or a procedure).

That brings me to my main point. To best understand microbe behavior we must not ignore the elephant in the room: the human immune system. The human immune system, and its extensive army of immune cells and antimicrobial molecules, plays the greatest role in determining how the microbes in any human ecosystem act, survive, and behave. Lately I’ve been describing this relationship with the following analogy:

Imagine a school. The children in the classroom are different microbes in a human microbiome community. The teacher and her support staff (like the principal) are the human immune system. Some of the children in the classroom are trying hard to get work done. Others are not so studious and hope to cause trouble. But these potential “bad students” don’t act out when the teacher is in the room and they can be disciplined by the principal and other staff. HOWEVER – if the teacher leaves the room, and her support system dwindles, these “bad influence” students can easily change their behavior to cause trouble. Without supervision they stop doing work. Further, they begin to encourage the better students around them to join them in promoting disorder. Little by little, even the best students find it hard to behave in the unsupervised classroom environment.”

For example, take the bacterial species Lactobacillus (which is added to many common probiotics). It is often regarded as a purely “good” microbe. However, as this article on the vaginal microbiome points out, some strains of Lactobacillus help maintain a healthy, acidic vaginal environment. BUT other strains can put women at a higher risk of acquiring a sexually transmitted infection. Indeed, Willa Huston, a professor at the University of Technology Sydney states the following about Lactobacillus and vaginal health: “It depends on the type of Lactobacillus, it might depend on the function of that Lactobacillus…and perhaps how it’s interplaying with the genome type of the woman.”

If you’re not convinced, let me give you a great example of how a “good” microbiome community can go bad (evolve in a direction of virulence). We’re all familiar with the fact that our teeth can develop cavities. The process that drives this cavity formation is called periodontitis. Periodontitis is now understood to be an infectious condition in which different oral microbes work together to damage tooth tissue and bone.

But aside from teeth brushing, what factors determine if a tooth develops a cavity or remains healthy? A team in Boston addressed this question by studying how the entire oral microbiome changes its gene expression during cavity progression. They found that community-wide gene expression did not change in teeth that remained cavity-free. But, the oral community dramatically altered its gene expression in sites that progressed to periodontitis. These changes involved genes that control oxidative stress, iron transport, sulfur metabolism, intracellular survival, and bacterial movement among many others. Cavity formation was also correlated with increased expression of virulence factors and higher levels of viral activity.

The team concluded that the entire oral microbial community, and not just a few select pathogens, drives the increase in virulence that leads to periodontitis. In effect, under conditions of increasing immune suppression, imbalance and inflammation, the whole oral microbiome community appeared to act together as a pathogen. This was supported by the fact that, in cavity sites, groups of microbes not usually considered to be pathogens expressed a large number of the disease-promoting virulence factors. Two of these microbes were S.mitis and S.intermedius –  bacterial species usually associated with dental health that are commonly described as “good.”

Single microbes can form into communities called biofilms.

Many cavity forming microbes also team up to persist in communities called biofilms. These biofilm communities are surrounded by a matrix that protects the different microbes inside from the human immune system. Participating microbes use electrical and chemical signals to communicate. This allows them to split up jobs important for their own survival – for example, virulent bacteria may protect the biofilm from outside intrusion, while other bacteria on the inside focus on obtaining nutrients for the community.

Few people would argue that microbes in a biofilm are “helping” their human host. They have instead joined forces in a manner that promotes their own survival over ours. It’s not surprising then that biofilms are connected to an increasing number of inflammatory conditions. In fact, some researchers estimate that the majority of bacteria in the body persist in a biofilm state.

So the way I see it, the human immune system and the human microbiome are actually in a continual state of competition and evolution. In healthy people the immune system has gained “an edge” over the microbiome. It is successfully able to keep microbiome pathogens in check. In inflammatory disease however, the opposite happens. The microbiome – or pathogens (or groups of pathogens) – begin to gain “an edge” over the human immune system…and its various defense strategies. This last situation is especially true in our modern world where an increasing number of environmental exposures and drugs are wearing down the human immune system.

And that’s what I’ll be talking about in one of my next podcasts: What factors most knock down or debilitate the human immune system in our modern society? And how can we help bring our immune systems back to a state of better activity and health? – a state that will help keep our microbiomes in a state of balance.

I will also do another podcast that explains more about how pathogens, and the metabolites they produce, are able to interfere with the human immune system and human pathways that control metabolism.

Interview with Ry Young: Phages and Phage Therapy – Pros/Cons and History

February 17th, 2018 by Amy Proal

Ry Young is a Professor in the Department of Biochemistry and Biophysics at Texas A&M University. He is also director of the Center for Phage Technology. Since the early 1970s Ry has studied bacteriophages: viruses that infect bacteria. He is now one of the world’s leading bacteriophage experts. His lab investigates bacteriophages as models for a new class of antibacterials. He has also participated in several “phage therapy” cases – in which bacteriophages are used as therapeutics to treat pathogenic bacterial infections.

Background information:

Tom Patterson phage therapy case: A recent medical case in which phage therapy was successfully used to treat an antibiotic resistant infection. Please watch this TED Talk for more context.

The interview:

Amy: Hi Ry. Thanks for taking the time to speak with me. You’ve researched phages for decades. How did you get interested in the topic? 

Ry: That’s a tough question. I don’t remember a time when I wasn’t interested in phages. I was a graduate student at MIT in 1968. At the time, one of the founders of the Phage Church was named Salvador Luria. He won the Nobel Prize in 1969, along with Max Delbrück and Alfred Hershey, for work on phage genetics. Luria was my microbiology instructor at MIT and turned me on to phages. So I learned about phages from what they call the horse’s mouth – Sal was a very charismatic person.

Phages attach to the outer membrane of bacteria with a thin “tail,” and inject their DNA into the bacterial cell.

Soon after however, I got my Vietnam War draft notice. I left graduate school to spend three years in the Navy. When I got out I went back to graduate school and started working on molecular biology. But as a postdoc, I got an NIH fellowship and chose to work in the lab of a brand new Harvard faculty member, Mike Syvanen, who was a phage guy. In 1978 I started my own phage lab here at Texas A & M. What I like about phage biology is the rigor of the field. Phages are easy to work with and have powerful genetics. You can posit hypotheses, do experiments, and get quick results.

Amy: So when you started researching phages I assume there wasn’t much of a concept of the human microbiome? 

Ry: No no. We had no concept of that. In fact, when I started working here in 1978, I had little idea that I would be one of the last people to start a phage lab for about 15 years. The entire phage movement, or Phage Church, was disassembling itself during the late 1970s and 1980s. This was based on an attitude in which phages were valued strictly as model systems for genetic experiments; as organisms that could help us work out the fundamental rules of molecular biology. But even then, many phage researchers decided it was time to move away from phage genetics to study other model systems. So the people who went on to found important fields of genetics research – like C. elegans for neurobiology and zebrafish for developmental biology – were mostly all phage people who moved away from the field.

This shift was largely started by a researcher named Max Delbruck, who founded modern phage biology. He was a physicist and reductionist: and if you’re a physicist you study the hydrogen atom and then, when you’ve got a predictive understanding of that…you move up to helium and so on. In fact Delbruck himself moved away from phages in the 1950s. So when I started my lab, I was one of a new wave of researchers who were interested in phages per se, rather than using them as a tool to study something else.

Max Delbruck in the early 1940s.

Amy: I see. I didn’t know that.

Ry: If you’re gonna keep up with phage you should get a copy of a textbook called “The Bacteriophages.” It’s edited by Richard Calendar (he’s retired now in San Francisco). I wrote the book’s one-page foreword in 2006. As part of writing it, I did an analysis of  NIH grants for phage research over the past ~ 30 years. I compared 1972, the height of the phage era, with 2002 (30 years later). I discovered that the number of NIH grants on phages had dropped down enormously. The number of RO1s for real phage research had gone from over 200 in 1972 to less than 10 in 2002. Basically the field had died.

Amy: Yes. But now it seems that the field has recently been reborn. 

Ry: Yes. Which is a real problem because suddenly there’s all these people who want to research phages, but few labs set up for phage research.

Amy: Interesting. Does this situation have anything to do with phage therapy?

Ry: No. The first stirrings of renewed interest in phages (and phage therapy) occurred in the early 2000s. That’s because the terrible antibiotic resistance problem we face was becoming more well-known. Even then, we still didn’t know anything about the human microbiome. So there are two separate current justifications for the renewed interest in phage therapy – One: we’re not getting any good new antibiotics. Two: even if we did we shouldn’t be using them. It’s kind of a double whammy in that regard.

Also, since the time when phage biology mostly died out in the 1980s, all sorts of new technologies have emerged – like next generation sequencing, laser technologies and high resolution fluorescence microscopy. The development of cryo-electron microscopy was especially important, because resolution power became so great that we could finally see the details of the phages. Suddenly, phage-related questions that had previously been beyond anybody’s ability to think about could be studied. Not only that, there’s a huge tide of new genomic information that’s bringing home to everybody what an important role phages play in the biosphere.

Because, back in the 1930s, phage biology was classically so powerful that the phage research community would get ahead of itself. That’s what happened with the original phage therapy. In the 1930s phage therapy was pioneered by a guy named Felix d’Herelle. He was a force of nature and went all over the world promoting phage therapy. So thanks to him, phages became the first widely used biological therapeutic.

But in the 1930s science in general was premature –  we didn’t even know about DNA. And because there were no standards, and no FDA or anything like it, most of these early attempts at phage therapy were kind of hopeless. We’d have been a lot better off if d’Herelle had not discovered phages until the 1940s or 1950s, when the scientific community knew more about molecular biology.

But there are other arguments about why this early phage therapy eventually fell out of favor. Part of what caused the decline was personality driven. And a lot was economics driven, because cheap antibiotics like penicillin were also discovered in the 1930s. No biological treatment like phage therapy is cheap. You can’t mass produce phages like you can penicillin. Also, to treat a patient you need thousands of phages versus one penicillin.

Amy: But in parts of Europe and Russia phage therapy did continue, right?

Ry: The very best person in terms of the history of phage biology and phage therapy is Bill Summers at Yale. He’s a great guy to talk to. He’s gives the world’s greatest seminars. They’re funny but illuminating. He was a phage and virus person way back when he was a junior faculty member, but now he’s become a very prominent scientific historian. He’s written any number of articles and books about phages. He’s the only person who knows the whole story. And unlike some people who mercilessly promote phage biology, he can tell you the underside too. And there are some undersides.

Amy: I want to go into those undersides. But before that let me ask you: Now that the microbiome has been better characterized, how would you describe the role that phages play in the human body?

Ry: Oh I don’t think anybody knows right now. Most of the information we have about phages in the actual body is derived from huge metagenomic experiments. For example, they might sequence your gut microbiome and discover there’s an awful lot of phage sequences in there. But by far the most common phages found by such studies are lysogenic phages – the ones present as prophages inside bacterial DNA. That’s a little misleading because all you can tell from that data is that those prophage sequences are being carried by bacteria. You don’t necessarily know if  the phage in question is even functional, or if it’s actually floating around doing anything.

Bacteriophages seen under an electron microscope.

But I think it’s clear that any microbiome community can’t be understood without understanding the phages too. We know phages are going to play a prominent role in the microbiome – there’s no way it can be avoided. Phages clearly interact, both positively and negatively, with the other species. We also know that the human microbiome changes as people develop. But nobody knows how many of these changes are mediated by phages. One of the things about phages in the gut is that there’s a lot of bacteria in the gut microbiome. This crowded environment allows phages to have a massive and rapid effect. Because phages have to absorb bacteria to kill them, and this rate of absorption is proportional. It’s a two body collision. The more phages and the more bacteria you have, the faster those collisions happen. For example, 200 phages can get made out of one bacterial cell.

So it’s a population density issue: when you have lots of any particular bacteria they become suddenly vulnerable to a single phage particle in the population. Whereas if the bacteria are very dilute, you could have 10^5 bacteria X and 10^5 phage Y (that would be able to grow on X) in the same microbiome community, but they might never encounter each other.

Amy: Right. But phages have been detected outside the gut right?

Ry: Oh yeah. Anywhere where you find bacteria in the human body you’re going to find phages. And for every type of bacteria there’s a different phage profile. Phage biology was really started with the study of laboratory strains of E.coli and closely related bacteria like Salmonella. Probably 99% of phage biology research was done on those microbes in the pre-therapy days (including my own research). And with E. coli, we found that you can go into any sewer or pond and find all kinds of different E.coli phages, in their every possible lifestyle. You’ll find lysogenic phages, and then all kinds of what people call lytic phages. Still, much of the variety of phages for E. coli has still not been tapped – we keep finding new ones.

On the other hand take Staph aureus – a high profile bacterial target (MRSA is surging in hospitals etc). There’s actually very little phage biology for Staph aureus. There’s basically two lytic Staph aureus phages. One is called Phage K (a big myophage). The other is a little podophage (a phage with a short stumpy tail). That’s all there is. No matter where you go in the world, if you look for lytic phages for Staph aureus, you’ll only find Phage K, its cousins, or the little podos.

That makes one ask: Why does Staph have such little phage variety? It’s seems like there was a bottleneck way, way back in evolution and those two phages won. It’s unfortunate in some ways, because we would like to have a lot more different kinds of Staph phages. On the other hand, phage K may have “won” because it evolved in a unique fashion. For a phage to affect a bacterial cell, the bacterial cell wall has to have a receptor that the phage can bind to. The bacterial cell wall receptor phage K and its friends use is a molecule called teichoic acid. And this teichoic acid receptor is essential for Staph: it can’t survive without it. So Staph cannot mutate to lose the receptor.  That’s probably why phage K “won”: it learned to recognize that essential receptor and then all the other phages were out of luck.

Amy: So Phage K outcompeted all the other phages? 

Ry: Yes and because the little podo phage has a very narrow host range, phage K is the only phage that’s useful. So there are a bunch of companies that have made phage cocktails against Staph, and they all turn out to be Phage K (plus friends and family). If fact, phage K has probably been patented 30 times.

Ry:  Do you know about the Bayne-Jones-Eaton report of 1934 that supposedly was the death knell of phage therapy?

Amy: No I don’t.

Ry: In 1934, the American Medical Association commissioned two scientists (Stanhope Bayne-Jones and Monroe Eaton) to survey phage therapy, to see if there was any anything “real” to it. They report they produced was very long – it was published in three separate issues of JAMA. They did a thorough study of all the different reported uses of phage therapy. Their general conclusion was that there was no demonstrated therapeutic benefit. And that’s not saying that phage therapy doesn’t work, just that the science hadn’t been done to show it could. However, the report did have an asterisk which stated something like “maybe we’re being too harsh, it does look like phage therapy works for Staph aureus, but nothing else.”

They came to that conclusion because they didn’t know about molecular biology. And if you don’t know what you’re doing, phage K will still work, because no matter how badly you use it it’ll still work. So there’s plenty of good evidence from thousands of cases showing that phage therapy works for Staph infections – evidence that goes way back to the 1910s – 1920s. But the Bayne-Jones-Eaton report still states that phage therapy has no demonstrated benefit (especially if you just read the top line). Also, that was right about the time when antibiotics started becoming available. So that was it: game over for phage therapy.

Félix d’Hérelle

I recently read the Bayne-Jones-Eaton report in depth while preparing for a two-day FDA workshop on phage therapy in July. I did a historical overview to see what the regulatory landscape was back when it was originally written. My interpretation is that “the fix was in for phage therapy,” because Bayne-Jones had a conflict of interest.

In the report, Bayne-Jones actually refers to the “failure” of phage therapy as the “d’Herelle Phenomenon”. And when you start calling something by somebody’s name, you instantly recognize that Bayne-Jones had something in mind about d’Herelle. At the time of the report, d’Herelle had been in the United States as a professor at Yale. There, he did what he did everywhere: frustrate many of his colleagues – he had a real talent for making enemies. He was self-educated with no college degree, much less a Ph.D. I think he was always very defensive about that. So eventually d’Herelle left Yale and Bayne-Jones took over his position. In modern times that would have instantly disqualified Bayne-Jones as an author of the 1934 report: You take somebody’s job and then write a report on the previous guy’s work!? I don’t think that’s a recipe for fairness. Although the report mattered less because antibiotics were going to wipe out phage therapy anyway.

One consequence of this was that d’Herelle had very sad life. He got nominated for the Nobel Prize about 28 times, but always had enemies high up in the scientific hierarchy that would blackball him. Because by any standard he should have gotten the Nobel Prize. He completely invented the whole area of phage therapy.

Amy: It would be interesting if d’Herelle could see new molecular data on phages.

Ry: Oh gosh yeah. He did a lot of fundamental research with no molecular tools, the lessons of which have turned out to be true a hundred years later. But because he was on the outs with the hierarchy, he ended up gravitating to to the left. The book he wrote about phages was dedicated to Joseph Stalin! Not exactly the way to endure yourself to the academic community. He had a real talent for painting himself into a corner.

That’s how he ended up starting the Phage Institute in the Soviet Union. As a result of that he ended up under house arrest all during World War II, because the fascists took over France. It was almost like a bad airplane novel. So now, in 2018, there’s a new phage therapy era. But to me this era is a rebirth of d’Herelle’s original idea.

Amy: I see. So this current rebirth of phage therapy: What’s the climate here in the U.S.? I know the FDA doesn’t allow it. What are the reasons for that?

Ry: Actually the FDA has been nothing but positive about phage therapy. It’s just that no phage therapy cocktail has been put through FDA tests yet. But the FDA has been very proactive: they’re trying to figure out what to do next. In fact, starting with the Tom Patterson case, they’ve set up a system where if a physician reports an appropriate phage therapy case, the physician can get “expanded access” (what we used to called compassionate use) to use phage therapy. This expanded access can be authorized within a day.

So while the FDA is not going to open the floodgates and say, “OK you can start injecting phage into people, don’t even bother calling us,” they’ve agreed to this expanded access (especially in life and death situations). Most of the time expanded access is used for cancer patients – at the end of life, one might as well give experimental treatments a try. So many potential cancer treatments that aren’t ready yet for clinical trials are also used in the same way.

Amy: But in the USA do you have to exhaust all antibiotic options before being allowed to use phage therapy? 

Ry: I think in general that’s correct. Right now if you want to use something that is not approved by the FDA you have to at least say there’s no other treatment option. That’s not a phage-specific guideline but true for all experimental drugs.

Amy: Might that change in the future because of the current antibiotic resistance problem?

Ry: Yeah I don’t know. I doubt that phage therapy will take the place of antibiotics. In the foreseeable future phages are going to be last resort items, mainly because of FDA approval issues. But there are several other problems with phage therapy. One is that we’re not going to find a broad-acting phage like penicillin. And since you know about the microbiota you could say, “we shouldn’t try to find a phage like penicillin!”

Amy: Yes, that’s a benefit in my mind. Because penicillin acts like an “atomic bomb” to the microbiome. It depletes entire microbiome communities in an effort to target one bacterial species. 

Ry: Right exactly. It would be better if we had antibiotics that targeted the disease bug and not the rest of your microbiota. But one reason we don’t have any small molecule-specific antibiotics is that there was no demand for them in the past. Up until recently Medicine wanted broad spectrum antibiotics.

But now there’s going to be a market for specific antibiotic molecules. I’m confident that pharmaceutical science and the industry are going to try very hard to find mass producible molecules that can target Staph, Psuedomonas etc. Those antibiotic molecules would be “round pegs” that fit fairly well into our established FDA drug testing system. For example, if you can find molecule X that kills Staph aureus, and doesn’t kill your microbiome, that molecule will be of great value. Then, somebody will be able to afford to pay the $100 million to $300 million necessary to put that molecule through the FDA drug approval process. I also think that the lysin technology that’s derivative of phage biology will become relevant, because it’s also a cheaper treatment option than phage therapy itself.

Amy: What is lysin technology?

Ry: Lysin technology is part of the phage story. For 40 years the NIH has been funding me to work on lysis (although I have no stock or conflicts of interest). The last step of the phage infection cycle is called lysis. That’s when the bacterial cell blows up and progeny virons are dispersed. One of the proteins involved is an enzyme called endolysin, or lysin, which means “the enzyme from inside that causes the cell to lyse.” Bacteria are protected by cell walls. But if you degrade the cell wall, which is what the phage does, then the cell blows up and the virus particles come out. So these lysin enzymes have now been developed to work from the outside to target gram positive bacteria like Staph. You take the phage lysin, make a lot of it, and inject it.

Crystal structure of a bacteriophage endolysin.

Lysins are not small molecules, but they’re a lot smaller than a phage. So you can mass produce and patent lysins – two things you can’t do with phages. But I think lysin technology will only be useful for Gram positive bacteria. Gram negative bacteria and mycobacteria have tough outer surfaces that endolysin can’t get at. The mycobacteria have a wax-like coating outside the cell wall that’s very difficult to break open.

Amy: Interesting. Back to phage therapy. Imagine there was a perfect world where we had studied phages in much greater depth. There’s a person with a bacterial infection and you’re trying to find a phage to treat them. You’re allowed to proceed however you choose – how would that work? 

Ry: You have to use multiple phages, otherwise you will probably get resistance very quickly. The word resistance means one thing to you, and a different thing to phage biologists. When phage researchers say “resistance” we are referencing a very specific situation: a phage cannot attach to a bacterium because the receptor it targets on the bacterial cell wall is damaged or missing. To clarify, every phage has a bacterial receptor it targets, and almost all the time those receptors are non-essential for the infected bacterium. So the bacterium can simply delete the receptor by mutation, allowing it to become completely resistant to the phage.

Given that notion, if you want a Pseudomonas phage cocktail to work, you need to have at least three phages that would target every possible Pseudomonas strain they encounter. Each one of those phages would target a different bacterial host receptor. The typical frequency of a knockout mutation for bacteria is one in a million: not a very big number if you’re being treated with one phage (because there are millions of bacteria and phages in the system already, some of which are already resistant). So as soon as you kill off all the bacteria that aren’t, you’re right back where you started within a few hours. But, if you have two different phages attaching to two different receptors, then instead of 10 ^- 6 frequency of a bacterial knockout mutation, it’s 10^-12. If you have three, that’s 10^-18. And nobody has 10^18 bacteria in them, at least no one who’s alive:)

So I’m using your perfect world scenario. In my perfect world there’s a Pseudomonas cocktail that’s ready for strain X of Pseudomonas, and we know its receptor targets. We also know that the bacterial strain we’re trying to target has those three receptors on its surface. Now, we can use that phage cocktail with confidence: we’re probably not going to generate resistant bacteria. That’s my hardliner perspective.

So the use of single phages could create a problem similar to what’s happening with conventional antibiotics? (where treatment can select for mutant bacterial strains) 

Ry: Right. Except resistance to phage happens a lot more frequently and quickly than with conventional antibiotics. If a bacterial cell runs into penicillin, it’s going to die unless it evolves to mutate residues of a particular enzyme in the bacterial cell wall (very difficult). So the mutations that create resistance to antibiotics are actually very rare. The problem is that because we’ve overused antibiotics, and put so much selective pressure in humans, many bacterial disease strains have sucked up genes that were already in the dirt. These are genes that had already evolved to degrade antibiotics.

But bacterial resistance to phages is a major concern. In fact, when I was a graduate student working in the heyday of phage biology, the classical attitude was that the resistance issue rendered phage therapy a joke. My mentor (and other members of the Phage Church) called phage therapy “bizarre” because resistance can occur so quickly. Of course, at the time we were completely unaware of how many more phages existed that we could use for cocktails. This was way before the development of next-generation sequencing.

Amy: To summarize. An upside of phage therapy is the specificity of the bacterial target. But that’s also a downside. And this issue could be addressed by using phage cocktails in lieu of single phages?

Ry: As director of the Center for Phage Technology I get to have official positions which absolutely no one has to listen to. But I always dreamed of being able to make pronouncements:) It’s our considered opinion that no phage should be used in a human unless we know what its receptor is. Period. Full stop. There are plenty of people who disagree with me because they don’t want to go through the trouble of figuring out what the target receptor is. But in the long run I can’t imagine the FDA is going to countenance the use of phages without clear receptor targets. And of course the original history of phage therapy has a bad reputation. There were negative outcomes and bad publicity. So now, when we restart phage therapy 2.0, we need to do it right.

Here’s another problem: right now phage therapy is always used via “expanded access.” When the phone rings in my office it’s always a crisis in which somebody is dying. And I’m not going to tell a dying patient and their family requesting a phage that we need six months to figure out what phage to use. But if we keep operating under these rushed emergency conditions, we’re going to start using phages that all use the same receptor. Then we’re going to get resistance and somebody’s going to die. That could lead to serious legal issues. If someone dies after being given an experimental phage, lawyers could later say, “Doctor yes or no…did you send viruses to this sick man? And were these viruses approved for treatment!?”

Amy: But to use an expanded access treatment the patient has to sign a medical release form right?

Ry: Yeah but that doesn’t help a bit because a patient’s heir’s lawyers are not bound by the patient’s request. So lawyers could still come after you. They wouldn’t come after me because I haven’t got a dime. But they might come after my University. Also, a flipside of the specificity issue is that in order to have any expectation of phage therapy success, we need to know whether the bacteria in a given patient are actually sensitive to the phages we have. That means we need at least one day to screen.

Recently there was a phage therapy case involving a young woman. She had gotten a lung transplant and appeared to be cured from a severe Burkholderia infection. But then the infection came back. So along with 3-4 other labs we tested Burkholderia phages. We tested over 400 phages, but only found three, and all of them were lysogenic. It took us 3 days working 24/7 to do that. Unfortunately she passed away a few days later (for reasons not related to phage therapy, but still).

But if we had already had a refrigerator full of phages, and we had known the receptor for those phages, we could have done some very simple, quick experiments. Or you could use a robot to get the bacterial strain from the patient, and know within 24 hours which of your phages might work against it. And if those phages are already pre-purified they could quickly be in the mail. But right now, no such well-characterized stock of phages for any bacterial pathogen is available. Again, the main reason for that is there’s no market for it. Because as far as I know, with “expanded access” you can’t sell the experimental drug (phage): you have to give it away.

Amy: So again, in a perfect world, it seems like the phage therapy community could use some time to calmly prepare for emergency phage therapy scenarios?

Ry: Yes. We could take highly prioritized pathogens like Baumannii and Pseudomonas and create a well-characterized collection of phages for which we knew the receptor. It wouldn’t even be a multimillion dollar project – more like hundreds of thousands of dollars. Which sounds like a lot of money, but I wonder what Tom Patterson’s bill for eight months in the UCSD ICU was? Maybe something like $10,000 a day? So the money needed to generate phage libraries wouldn’t be that large compared to many related costs. But nevertheless, who’s going to pay for it? And is every reference hospital going to have a walk-in cooler with thousands of phage lysates or purified phages? The other possibility, and the dream of the phage therapy world, is to have phage that works against all targets. For Staph aureus that already mostly exists.

Amy: A superphage?

Electron micrograph images of phage K (American Society for Microbiology)

Ry: Phage K is a “superphage.” You can get a phage K that will eliminate 85% of all the Staph on its own. It would target all Staph, except for the fact that bacteria find ways to fight back against phages that aren’t related to “resistance.” Bacteria also have restriction enzymes and their CRISP-R-associated systems, for example. But I think the dream of a super phage is very close to reality with phage K and its relatives. For other bacteria though, we haven’t found many examples of bacteria with essential cell wall components that phages can be targeted to.

Amy: But we’re discovering new phages every day, right?

Ry: Yes. We’re early in the game. And what’s astounding is that all the technology we’re using to characterize phages (with the exception of molecular sequencing) is really the same that d’Herelle used back in the 1930s. Again, that’s because there’s no current market for phage therapy. But, if people think there could be a market down the road, a lot of progress on how to purify phages could occur. The process could become cheaper than it is now. But currently we make phages for therapy in the same way we do for laboratory experiments. And that’s not something you can do on a large scale.

Amy: It seems to me the FDA will have to “paradigm shift” the way they’re going to evaluate phages and other new biological drugs.

Ry: Right, because it’s not just phages. There’s a lot of other biologicals that are like square pegs in the round holes of a regulatory process geared towards developing organic chemicals. I’m hoping the FDA’s “expanded access” phage decision will increase the climate for public and physician awareness of phage therapy. Also, there’s been some progress.

Amy:  What about pressure from Biotech companies? There is a focus in the Biotech community towards the development of personalized treatment approaches.

Ry: Yes. And phages are the ultimate personalized medicine.

Amy: That makes me think the FDA’s current drug approval process is going to become a problem for a lot researchers and drug companies.

Europe is ahead of us in addressing this issue. Partly because the countries involved are much smaller scale countries. If you’re living next door to the Minister of Health, it’s a lot easier for you to get your ideas across. So Belgium, for example, is pretty far ahead of us in terms of making “expanded access” more regularized. They’re going make it easier for physicians to say “I’m going to use phages.” And there will hopefully be suppliers of good phage. Then, the physician will be able to buy/use phages and have protection against being sued.

I don’t think phage therapy will replace antibiotics though. I want to make sure you understand that. I think at least for the foreseeable future phage therapy is going to be a last resort thing. I would like to see a lot more basic phage biology funded. I think in order for phage therapy to best move forward, we also need a rebirth of basic phage biology research. Because almost everything we currently know about phage biology is based on just phages of E.coli (which is of course what I worked on myself.)

Amy: Yes. A recent study estimated that 30 billion phages may traffic the human body on a daily basis. It does seem like we need more research on all these “new” phages!

Ry: Right. DNA sequencing is nice to have, but it doesn’t substitute for doing experiments that help better understand phage activity and function.

Amy: Did you read the recent Georgia Institute of Technology study which showed that the host innate immune response seems to play a role in helping phage therapy work? Because if that’s the case,  then our widespread use of immunosuppressive drugs and steroids seems like a problem. Especially because such drugs are administered frequently in hospital emergency settings.

Phages interact with the human immune system (Roach et al, Cell Press)

Ry: So I suspect that phage therapy will be mostly useful for reducing initial loads of bacteria, under conditions where a patient harbors enormous amounts of a bacterial pathogen. Because once the bacteria get down to a certain level they don’t collide with the phage anymore. Phages are not small molecules, so they can’t penetrate every body niche to find a bacterial target. So I think that phages will be most useful for somebody who’s got a raging infection. You give them a first round of phages and a knock the bacteria down by two or three logs. Now you’ve got a chance for either the body’s own immune system or antibiotics to take over.

Amy: At least in that scenario antibiotics are being used in a careful situation. As opposed to many other ways in which they are over-prescribed today.

Ry: Yes. There’s a lot of speculation out there that many of our current autoimmune diseases, like Crohn’s, may be due to the fact that our generation (my generation) was the first to be massively dosed with antibiotics all throughout childhood. My generation was convinced that if you didn’t get an antibiotic prescription you hadn’t been treated right. It was the golden age of antibiotics.

Amy: Right. When antibiotics were discovered they were perceived as miracles, which they kind of were, especially during World War II. But that has created a situation where the average person has not been well-educated about how antibiotics work. For example, about the fact that antibiotics can’t kill viruses. 

I think that the American Medical Association is at fault for not educating the public better about antibiotics. I mean they really are. For example, you see all these TV commercials for erectile dysfunction or cancer drugs. There ought to be a way that some of that airtime could be used to educate the public about medical topics like antibiotics. Especially because most drugs advertised in these commercials were developed with NIH funding. So I think big drug companies owe at least that to the public. Also, physicians need to have the courage to say “You have a viral infection. I’m sorry but it can’t be treated with antibiotics.”

Amy: Yes. I know you’re considered a “hardliner” when it comes to the future of phage therapy, but I think the caution you advise moving forward in spot on. We don’t want to create a future situation with phage therapy that could repeat problems we’ve encountered with antibiotics. Understanding and correctly managing phage resistance will be critical.

Thanks so much for taking the time to speak with me. I learned so much from this discussion.   

20 Questions for Gurol Suel: biofilm electrical communication

January 29th, 2018 by Amy Proal

Gürol Süel is a Professor of Molecular Biology and the Associate Director of the San Diego Center for Systems Biology. His research team integrates quantitative biology approaches with mathematical modeling to identify principles of bacterial organization and coordination. Most recently, the team discovered a new form of bacterial communication that arises in biofilms: Ion channel mediated electrical signaling. For more context on this discovery, please read this article in The Atlantic Magazine.

Amy: Thanks so much for speaking with me. Your last few papers on electrical communication in bacterial biofilms blew me away. I think many other people feel similarly! How did you get interested in the topic? 

Gurol: We were very interested in trying to understand higher-order organization and behavior in bacterial communities. That’s because even though bacteria are unicellular organisms they’re not solitary. They actually live in dense communities (biofilms) that have certain features: for example they’re very resilient to stress and antibiotics.

We became interested in biofilm electrical signaling partially because we saw that in biofilms there was some kind of long-range metabolic coordination. We were having a hard time understanding how the bacterial cells could communicate so effectively over such long distances. It seemed that a simple diffusion-based mechanism (like quorum sensing) might not be sufficient to support this communication. My Ph.D. is in molecular biophysics. I also have a background in electrophysiology. For example, I used to do electrophysiology in fruit flies with photoreceptors. So drawing from that experience, I started wondering if there could be a different type of signaling process involved.

We started looking at amino acids critical for slow-range biofilm communication. The most important amino acid appears to be glutamate. Glutamate is very interesting because it’s also an excitatory amino acid in the human brain. In fact, about half of all brain activity is regulated by glutamate. Glutamate is also one of few amino acids that’s charged. So it can’t just get in and out of cells on its own because that’s energetically unfavorable. So you’ve got to have some kind of a proton motor force to actually take up the charged amino acids. And if you’re dealing with a proton motor force then the cell membrane potential is important.

Little by little we pursued this line of thought and realized that we were looking at some kind of ion channel purpose or function in this signaling process. And when you think about ion channels and membrane potentials you think about sodium and potassium – those are the most commonly utilized ions in biological systems. So we looked at both and saw that sodium wasn’t really doing anything. That wasn’t surprising because Bacillus subtilis, the microorganism that we work with, is not known to have any identified sodium channels.

Bacteria form communities called biofilms (Image: The Laboratory of Gurol Suel)

But we saw a lot of beautiful signaling and dynamics going on when we looked at potassium. Then, we took the suggested potassium ion channel and started making mutants. One thing led to another and we realized that these bacteria were communicating using potassium ion channels, in a way reminiscent of what neurons do in the brain with action potentials. Once we uncovered that, we realized that if you have these long-range electrochemical waves propagating through the biofilm, then the waves wouldn’t just stop at the edge but might propagate a distance.

That led us to the Cell paper where we said: What if there’s some kind of motile cells nearby? Would they be sensing or responding? Would they be affected by these types of signals being emitted by the biofilm? Pursuing those questions turned into a fun and intriguing study. Then we said OK – if that’s possible, what if there’s another biofilm further away: could the two biofilms sense each other and interact? That led us to the Science paper about time-sharing of limited nutrients.

Amy: Right. I enjoyed reading those papers because your experiments are so well organized. I’m interested in the redundancy between the way neurons signal and the way you found that bacteria in these biofilms are signaling. Do you think it’s possible that there are any interactions between human nerve signaling and these microbes? That they could tap into each other’s signaling?

Gurol: I mean, we don’t know. That’s the short answer. Because no one’s ever looked at that. Would it be possible? I guess it would be possible. I do know that extracellular potassium signals (the potassium ion channel mediated signals) are not specific to just neurons. We showed it now even in bacteria. But it’s also very important in plants. It’s very important in all kinds of other living organisms – because these ions are very generic, meaning they can affect extracellular charge in any membrane.

So it’s not inconceivable to think that if a bacteria releases some charged particles and there happens to be an excitable cell nearby – maybe a mammalian cell – that the mammalian cell might actually respond to that. I don’t think that’s super crazy. Right now it’s unproven but it’s something we think about.

Amy: Very interesting. What about external electromagnetic fields – do you think that our human electronic devices or any radio frequency networks we’ve set up as humans could be impacting biofilm communities?

Gurol: That’s a very good question. Again I don’t know for sure. But it’s well known that lots of organisms have electro-reception, meaning they can follow electromagnetic fields on our planet. Animals with electro-reception range from whales to birds. So if organisms can sense these fields, what you’re suggesting is probably not super crazy.

There are actually bacteria known to have small particles inside that are sensitive to magnetic fields. We also know there are documented cells that will respond to an electrical field, including bacterial cells. There’s also good work out there clearly showing that just like temperature, pressure and so forth, biological systems are sensitive to magnetic forces and electrical forces. That’s more or less well-established. What we don’t know is whether or not your cell phone is going to have anything to do with your gut microbiome or things like that. Those are the things that, to my knowledge, have not been clearly studied.

Amy: Not really. But interesting future research?

Gurol: Yeah. And as I said, there’s enough strong evidence out there showing that living organisms are sensitive to these physical external properties: pressure, temperature, pH, electrical fields, magnetic fields. That’s not a new concept. So we’ll just have to see. And I don’t know, do birds sit on electric wires because it tingles? Or is it just a good sitting place?

Amy: Haha. There was a study which showed that Interference from human electronics and AM radio signals disrupted bird migration…

Gurol: Yes I wouldn’t be surprised. I think there is enough information out there to at least create a good incentive to look at the topic in greater depth.

Amy: Yes. So I research the human microbiome. If someone asked you to describe the human microbiome knowing what you do now, how would you explain it?

Gurol: Maybe I would say that the human body is an ecological network. We’re existing together with our environment, and our environment is not just about whether it’s cold or rainy outside. Within our bodies we have countless organisms (not countless but lots of organisms) and bacteria are just one of them, in fact we probably have even more viruses than bacteria – nobody’s been able to track down everything. So we harbor a lot of organisms, especially in our gut. And I think it’s a complex interaction that we don’t fully understand. There’s always, of course, a little bit of hype – when we discover something new everyone gets excited, maybe even a little over-excited. But even if 10% of all the stuff out there on the microbiome turns out to be relevant that’s already a lot.

So I would say the microbiome is a little universe of an ecological network within our body. There’s things going on between all the bacteria. But then there’s also things going on between the bacteria and our cells, the human cells. And we have yet to fully appreciate how many and which parts of these processes are super-critical and which are processes that are interesting but maybe not as critical.

The human microbiome is an ecological network

For example, there’s always interesting studies about species composition of the microbiome – studies stating that composition is critical to this process or that process. But we have to remind ourselves that the composition of the microbiome is not a static thing. When I wake up and go to bed my microbiome is probably totally different depending on where I went to dinner and other variables.

So to some degree we can’t be super sensitive to species-specific microbiome changes, because then we would have traumatic, physiological problems throughout the day. That model just wouldn’t work. We also know that the microbiome can recover: you can take antibiotics and really flush out a lot of stuff but then you can very quickly recover your microbiome. So it’s going to be important to discern which processes have an effect but are not supercritical and which processes are actually very important. That’s the kind of stuff that makes it interesting for the future – to see where it leads us.

Amy: Many people research microbes in isolation and outside their natural environment. I know great insight can be gained from that work. But do you think there needs to be a larger push to research how microbes really act in vivo (in their natural environment)?

Gurol: Yeah. So “in vivo” is an interesting term. It’s very important when you’re dealing with something like the study of cell lines in the laboratory. Problems have appeared when people have studied these cell lines and then tried to claim things about cancer: because the cell line was taken out of its natural context. But when you’re dealing with bacteria to say “what is in vivo?” is not even very clear to me. Take the organism we work with – Bacillus subtilis. You have it right now on your eyebrows and on your skin. It’s also in your gut. It’s in the soil. It’s on your tablecloths. If you have fruits and vegetables in your fridge it’s in there. So which of those locations is “in vivo”…right? They are all very different conditions, but are all places where bacteria naturally occur.

So the question of “in vivo” is a blurred, fuzzy, and potentially misleading question: in the sense that we don’t actually know what to call “in vivo.” If you isolate bacteria in the lab it’s more clear what you’re doing. But to say “this is the proper in vivo context” – I’m putting out a little word of caution: these bacteria are capable of surviving under so many different conditions that we have to be careful.

But overall any good scientist would know that if you’re studying a cell you have to understand the context of the cell, meaning the environment of the cell. And if the environment of the cell includes other cells, and those may be the same species, or different species, then obviously that is something that needs to be taken into consideration. But one can still execute very important studies without being shackled to this vague notion of “what is in vivo?”

It’s especially important to remind people of this now, because as a scientific community are very concerned in trying to make sure that our data is reproducible, and that we have reliable results: that if I do an experiment somebody else can reproduce it. But the moment we get into these types of in vivo conditions the problem is very poorly defined. Even the eyebrow skin environment differs between two people based on fat content, pH and other variables. So we’re dealing with a lot of unknowns, and although we can pat ourselves on the back and say “hey we’re studying something in a more natural context,” we have to realize that it comes at a cost – the cost of having less defined conditions and a lack of understanding of certain things. So it’s a balance – there’s no right or wrong – but we have to understand the pros and cons of each of these approaches very carefully.

Amy: OK I see. Still, in any environment, the microbiome community should focus on studying microbe activity in addition to just species, right?

Gurol: Sure. Sure.

Amy: Ok, what about other microbes like viruses, phages, fungi. Do you think that they’re involved in this electrical of signaling as well?

Gurol: We already showed in at least one paper that two very evolutionary distant species of bacteria can communicate electrically – Pseudomonas and Bacillus. One’s gram positive and one’s gram negative, so they’re very different types of bacteria. That is an indication that this signaling is not just limited to a single species of bacteria. Although that would have been a great result too. Because then you would have to ask “how can it be that only one species evolved this thing!?” That would be kind of  interesting on it’s own also. But maybe not surprisingly that does not appear to be the case.

Bacteria use action potentials to generate electrical signals

So we are still at the ground level if you will. These discoveries are still quite recent in terms of scientific years – they’re the opposite of dog years that go extra slow. So time will tell how many other species signal electrically – which ones do, which ones don’t and what that all means. But again it goes back to the earlier question that you asked: how widespread could this be? It has something to do with the fact that the membrane potential is a very fundamental property of any cell. Any living organism that’s a cell has a potential across its membrane. It’s a very ancient set of physiological processes, so it wouldn’t be shocking in hindsight if we found that different species, and even organisms across many different kingdoms, can engage in some form of this communication.

Amy: Agreed. Can you tell me a little more about how you integrate mathematical models along with the study of actual biofilms to get your results?

Gurol: Sure, the mathematical models are quite important. We use them for two main purposes. One is to come up with very specific hypotheses. The nice thing about mathematical modeling is that you can make predictions that are hypothesis driven…more than hypothesis driven: that are mathematically supported. That is very important because not only does it mean that you can define the question you’re asking really well, but once you have experimental results you can also interpret them using the framework of a mathematical model in a way that is much more powerful.

The other reason we’re doing it is that mathematical models allow us to identify concepts, and concepts are very important because a lot of people are interested in “what is this gene?, or “what is this protein?” Those are tangible objects, and so people really like working on those things. Concepts are by definition intellectual things. That means they are not tangible, in the sense that you can’t touch it, you can’t say “here’s the protein.” But concepts are very important because they allow us to bridge across very different types of systems. So if we can use the same mathematical description to describe processes happening in very different organisms that is meaningful. You can say, “There seems to be something that is shared between two systems, and then you can say “Are they trying to solve a similar problem?” Maybe there’s some kind of evolutionary information there.

Hodgkin and Huxley

So identifying concepts is very relevant in finding connections between things that appear on the surface to be different. Like bacteria and neurons: if I you told ten years ago that they might have similarities, then people would chase you out of the room and say you’re crazy. But now we can use the mathematical formulas that were originally developed in the 1950s by the brilliant Hodgkin-Huxley team. They got a Nobel Prize for the work. They identified a mathematical framework and figured out that basic action potentials in neurons are driven by potassium and sodium flux across the membrane. We showed that you can use that same mathematical framework to explain and make predictions regarding electrical signaling in bacterial communities.

Amy: Yes absolutely. How have your results been received? It seems like pretty enthusiastically, but have you had trouble communicating this to people who have only studied microbes in isolation?

Gurol: No, I mean communicating work is a very important part of being a scientist, because knowledge not shared is like knowledge that almost doesn’t exist. If only one person knows about it, that doesn’t help anybody. So communication is critical. And when you’re communicating you cannot blame the audience for not getting something – you have to figure out how to communicate your results so that anybody can appreciate them. So we take great pride in trying to write papers that are accessible, even though obviously we have to deal with the fact that we’re communicating technical scientific work. I think this pays off in that if you’re able to explain WHY something is important, people can make their own judgement on whether it’s critical or not. The dangerous thing is if people don’t understand it and they’re just confused. Confusion leads to frustration. At that point it’s very difficult to decide if something is noteworthy or not.

Amy: I think you did a great job with communication. I like how you put the more complex math towards the back of the paper.

Gurol: We try to make sure that you can read the paper, and then if you are interested there’s more meat you can sink your teeth into. I’m sure there’s still much room for improvement but we try.

Amy: No, you did a really good job. Can you tell me little more about ion potentials and about how biofilms are able to coordinate behavior to time-share nutrients?

Gurol: Sure. So you mentioned our three latest papers that you liked. The first paper describes how potassium ion channels are utilized by bacteria in biofilm communities to send long-range electrical signals. These signals allow them to coordinate their metabolism. That was an exciting finding that set many other things into motion. In the next paper we described how these electrical signals could extend beyond a single biofilm community. If there’s planktonic motile cells swimming around (because biofilms exist in aqueous environments) it’s not crazy to think that they could be recruited into the biofilm community (even different species). We knew that bacterial motility is driven by proton motor force – the cells utilize an electrochemical gradient across the membrane to generate the energy required to churn flagella: swirly appendages that allow bacteria to swim. We said, “Well if you have electrical signals maybe they would have an effect on this process of motility.” And indeed, we found that that electrical signals can direct and attract motile cells from a distance to the community. That’s interesting because now you can see how electrical signaling can turn a single species community into one that recruits other species too.

We followed that finding up by saying, “OK so if there’s two biofilm communities – two physically separated communities at some distance – not super far away but, with respect to the size of a bacteria some significant distance – what would happen?”

We saw that if these communities share the same environment they become coupled – they can sense each other so to speak. This happens in two main ways. One is this electrical signaling which syncs communities under metabolic stress. But there’s also competition between two communities, where resources are limited and there’s only enough to sustain one community. We found that under such conditions, communities started modulating the coupling between them and transitioned to different types of dynamics: they began time-sharing the limited resources. Time-sharing is a well-known concept. Take vacation homes or cloud computing (where it’s called resource sharing). Another example would be if you have one toy and two kids that want to play with it. You have the kids take turns playing with the toy. It turns out this same strategy is being employed by these bacteria. It’s an exciting discovery because time-sharing is a very dynamic process. With vacation homes or cloud computing time-sharing is not trivial – somebody has to set up a calendar saying, “You take August I’ll take June.” So it requires some kind of higher organization or somebody who coordinates things.

Bacteria use flagella to “swim”

But we found that bacteria naturally time-share through their dynamic system behavior. They can actually transition from being in phase where they’re competing a lot (and that’s OK if there’s enough nutrients) to then switching to time-sharing behavior when nutrients are low. In that case, there’s nobody from the outside telling which colony to do what. The process naturally emerges through the interaction between the communities. It’s very interesting then that bacteria are capable of executing what appear to be nontrivial strategies for coping with limited resources.

Amy: It’s extremely interesting. Do you think we’ve underestimated bacteria? With each new study it seems that bacteria are able to do much more than we previously believed.

Gurol: I’m always amazed by any living organism. If you realize how long these guys have been around (especially bacteria) they’ve survived mass extinction, one after the other. Not just the extinction that killed dinosaurs, but also the earlier extinction where even more species disappeared.

So they’re extremely resilient. They’ve been able to colonize from the Arctic to the depths of the ocean. It’s fascinating. You know, sometimes I go to the east coast and I have trouble adjusting to the new environment! So I’m impressed by these bacterias capabilities. Also, I’m sure there’s still a lot we don’t know. It’ll be a fun future to figure out what else they can do.

Amy: I agree. I know this is jumping ahead but do you see any possible therapeutic interventions based on these findings? Maybe you could interrupt an electrical signal in a biofilm-driven illness?

Gurol: Sure. We really like doing basic research and then letting the findings lead us in those directions. But we are certainly not blind to the possibility that our discoveries may also have biomedical implications. We’re trying to be careful – we don’t want to put blinders on and just focus on one thing. But we do obviously realize that biofilms are a major public health threat. Biofilms are responsible for the majority of infections in clinics. There are people dying and billions of dollars at stake. So yes. We’re aware of these problems and are slowly trying to explore those avenues as well.

Amy: What are you going to study next?

Gurol: We’re still trying to understand some of the basics. We’re still very intrigued by this new form of communication between bacteria – what else can it do? At the other side, at the smaller scale, we’re also trying to understand if in addition to ion channels, there are other genes and proteins playing regulatory roles. So we’re diving both in the direction of more detailed depth and more rigorous research. But also going in the other direction to see “How widespread is this and what other interesting behaviors can come out of this?” I suspect this research will lead to more unexpected findings.

Amy: Is there a growing community of researchers who are getting interested in this? Or are you guys working mostly alone?

Gurol: No, there’s a growing community. In fact we’re going to have a first session on biofilm electrophysiology at the upcoming American Biophysical Society Meeting this February. There will be speakers from all kinds of different institutions and from around the world. So there’s definitely a community that’s building and I think this field has a lot of potential. It’s obviously a very small field only in its infancy, but we’re trying to draw in more people and create a connected community.

Amy: Yes. Some of papers you cite are from the 1970s. How did that earlier research play into your findings?

Gurol: As they say, we’re standing on the shoulders of giants. Smart people existed before we came along. There’s been beautiful studies done – going all the way back to Hodgkin and Huxley. In fact, I would go back as far as Luigi Galvani in the 18th century who did this beautiful experiment where he cut a frog in half and used electrodes to make the legs twitch. It really freaked people out and led to books like Frankenstein where the monster is brought to life with electrical currents. So this field goes back hundreds of years. Obviously a lot more people work on electrophysiology in the human brain because that’s an important organ and it’s easy to see the relevance. But people have looked from plants to fungi to bacteria because the ability of cells to use electrochemical potential across the membrane is so ancient.

Amy: Yes. So let’s say you were talking to a student who wanted to study biofilms or microbiology. What kind of skillset would you recommend this student get to best succeed in the field? 

Computers play a central role in the study of microbes

Gurol: First you have to learn how to think like a scientist, because once you understand how that process works you can work on anything. In terms of a more concrete skillset, the more diverse the better. If you have a unique background and have done things that others have not done then, in the future, maybe you can see things from a different perspective. You might discover something new. I’m sure my background in electrophysiology allowed me to think about biofilm signaling a little bit differently.

I’ll also say this, the future is as clear: we’re heading in a direction where being able to communicate in terms of a mathematical language is key. Also understanding large data sets and analysis – that’s not going away. So having no fear of computers and software and mathematics and physics: that will be important, because again these boundaries are blurring. I think being interdisciplinary is very important.

Amy. Yes. I wish I had a physics background. I’m jealous. Before we go would you like to leave people with any other message?

Gurol: We covered a lot of stuff. Hopefully we’ll have more stuff to talk about in the future. But I’m happy that people like you and others are taking note and finding our work interesting. In this day and age it’s very important for people in science and people in science communication to make work accessible and reach out to the public. Because the more we educate the general public and try to help them understand what science is and how it works, the better it is for our society.

Amy: I do. Thank you again for this fascinating conversation.

Immunostimulation: embracing a new treatment paradigm for chronic disease

January 12th, 2018 by Amy Proal

Think back to the last time you got the flu (virus). The fever, the runny nose, the aches, the sore throat – what causes these and related symptoms? Most flu symptoms are not driven by the virus alone. Instead, they result from a “battle” between the virus and the human immune system. Symptoms begin when the immune system recognizes the flu virus and creates inflammatory proteins called cytokines in an effort to target infected cells. If infected cells are successfully killed, more inflammation is generated as toxins and cellular debris enter the bloodstream. In addition, antibodies may be created in response to these cell and viral byproducts, again leading to a rise in inflammation.

Most cold medicines lower symptoms by suppressing parts of the human immune system

We don’t have antivirals capable of killing the flu virus. So we “treat” the illness by letting this immune system “battle” run its course. In most cases, the human immune system “wins” over time, and the inflammation caused by cytokines, toxins and antibodies drops. We begin to feel better and life goes on.

In some cases, patients manage the flu with over-the-counter medicines. These include Motrin, NyQuil, and antihistamines. In other cases a doctor may prescribe steroids or immunosuppressive medicines. These medicines lower symptoms but do nothing to target the virus driving the illness. In fact, these medicines “work” by shutting down various parts of the human immune response towards the virus. They “tone down” the battle between the immune system and the virus so that less inflammation is generated.

The above medications make patients FEEL better. But they may actually impede recovery from the flu by allowing the virus to survive with greater ease. For example, Canadian researchers found that anti-fever medications suppressed fever in patients with the flu, but also allowed flu viral particles to spread more easily from person to person. Indeed, the team estimates that the use of anti-fever medicines by flu patients contributes to a 5% increase in general flu cases and deaths.

Despite these negative outcomes, our entire medical system centers on immunosuppressive treatments that “knock down” parts of the immune system to suppress symptoms. Patients with autoimmune disease are regularly prescribed immunosuppressive medicines like prednisone, rituximab or TNF-alpha inhibitors. Humira – a TNF-alpha inhibitor used to treat arthritis and other “autoimmune” conditions – is the “best selling prescription drug in the world,” with a $38,000/year price tag per patient.

Why this focus on immunosuppressive therapies? Most immunosuppressive treatments were developed before ~2004, during a time when the human body was believed to be largely sterile. Under these conditions the “theory of autoimmunity” gained hold. If inflammation was detected in patients with a range of conditions it was assumed to be a result of the immune system “going crazy” and attacking human tissue.

The discovery of the human microbiome greatly challenges this “autoimmune” model of disease. We now understand that vast microbiome populations persist in every human body site – from the gut, to the brain, to the placenta, to the liver and beyond. An increasing number of “autoimmune”/inflammatory conditions are now tied to dysbiosis or imbalance of these microbiome communities. This means that in “autoimmune disease,” the human immune system may be attempting to target pathogens in the microbiome in lieu of attacking human tissue. Indeed, an increasing number of studies demonstrate that the “autoantibodies” used to diagnose autoimmune disease are often created in response to a range of bacterial, viral and parasitic infections.

This growing association between infection, “autoimmunity”, and inflammation helps explain the poor long-term outcomes associated with immunosuppressive therapies. Patients administered prednisone or TNF-alpha inhibitors tend to feel better in the short-term, but relapse is common, and often expected. Each relapse can require higher doses of immunosuppressive medication to get symptoms “under control.” Meanwhile, patients are at greater risk for developing a second or third inflammatory disease, and are more likely to suffer from acute infectious conditions like tuberculosis. Long-term health outcomes associated with prednisone are so poor that the slogan “pred ’til dead” is commonly invoked (patients who start prednisone often require higher and higher doses of the medicine until they die from the underlying disease).

This begs the question: what if we treated autoimmune disease in the exact opposite fashion? If microbiome dysbiosis contributes to autoimmune disease then treatments that SUPPORT the immune system could target pathogens driving inflammation. By addressing this infectious root cause of inflammatory symptoms, such treatments might induce actual improvement or even recovery.

How might patients with “autoimmune disease” or related inflammatory conditions respond to “immunostimulative” or immune-supporting treatments? Case histories from the turn of the century offer clues. In the early 1900s, mercury was used to treat syphillis: a sexually transmitted infection caused by the bacteria Treponema pallium. Mercury “deliberately stimulated the immune response” in patients with the disease. This resulted in a phenomenon known as the Jarisch-Herxheimer reaction (named after the researchers who characterized it). As the activated immune system targeted Tremponema pallium a “battle” not that different from that associated with targeting the flu virus ensued. Patients suffered a temporary increase in symptoms including fever, chills, myalgia, and headache as cytokines were released and debris from dying bacterial cells entered the bloodstream. However, if patients endured these symptoms they generally “turned a corner,” where symptoms subsided as Tremponema pallium was gradually eradicated.

In the 100 years since the Jarisch-Herxheimer reaction was described in syphilis, it has been further documented in patients administered immunostimulative therapies for a broad range of infectious conditions. These include Lyme disease, leptospirosis, brucellosis and tuberculosis. More recently, the term “immunopathology” has been used in place of Jarisch-Herxhimer to refer to “a systemic inflammatory response consistent with elevated immune activation.”

Immune activation and immunopathology in the treatment of HIV/AIDS

Over the past decade, immunostimulative treatments have been developed to treat HIV/AIDS and cancer. These therapies are also characterized by temporary symptom increases as the activated immune system attempts to target root causes of inflammation. Current HIV/AIDS treatment centers on highly active antiretroviral therapy (HAART). Patients administered HAART receive a cocktail of anti-retroviral drugs, each of which impedes the ability of the HIV virus to replicate and spread. Prior to HAART, the HIV virus survives by dramatically slowing key parts of the human immune response, including the CD4 cells that normally target infectious agents. This means that when the virus is “contained” by HAART, the immune system “wakes up” and identifies pathogens acquired during previous periods of immunosuppression.

IRIS leads to increased symptoms as the immune system “re-activates” (Source: Khan Academy)

What happens next is a form of immunopathology. The activated immune system starts to target pathogens it could not recognize before HAART was initiated. The patient begins to experience temporary increases in symptoms ranging from fever, to malaise, to neurological dysfunction. Symptoms wax and wane with time, but generally decrease as the immune system better targets a range of previously unrecognized pathogens. The HIV/AIDS community has named this process “Immune Reconstitution Inflammatory Syndrome”, or IRIS.

A number of well-known pathogens are linked to IRIS symptoms: the herpes viruses, cytomegalovirus, hepatitis B and C, M. tuberculosis and Mycobacterium avium among others. Often however, symptoms increase despite the fact that no pathogen can be identified on routine blood tests. This suggests that newly identified microbes, like many of the thousands recently detected in tissue/blood by Stanford researcher Stephen Quake, are also being targeted by the activated immune system.

Several key patterns have been observed in patients experiencing IRIS. One is the “unmaking” of infections that can range back to childhood. Let’s say a patient suffered from bacterial meningitis at age eight. The meningitis microbe may re-appear on IRIS-related blood tests thanks to the renewed immune “attack” against its presence. This supports the fact that pathogens acquired throughout life can persist in our microbiome communities, where they may contribute to chronic symptoms.

Second, patients experiencing IRIS often ‘develop’ autoimmune conditions as the immune system reactivates. These include sarcoidosis, diabetes mellitus, rheumatoid arthritis, lupus and Graves disease. This strongly suggests that pathogens targeted by the IRIS immune response also drive symptoms associated with these related inflammatory disease states.

Cancer therapies target tumors by activating the immune system

The latest cancer therapies also seek to activate the immune system. According to the American Cancer Society, these novel immunotherapies “stimulate your own immune system to work harder or smarter to attack cancer cells.” Cancer immunotherapy treatments include CAR-T therapies: treatments that remove disease-fighting T cells from a patient, genetically modify them to better recognize and attack tumor cells, and then add the activated cells back into a patient’s blood.

Cancer immunotherapy activates human T cells

Response to CAR-T immunotherapy results in serious immunopathology. Nearly all patients administered CAR-T therapy experience a rise in symptoms due to what has been named Cytokine Storm Syndrome or CSS. As implied by the name, CSS results when an immune system “battle” between activated T cells and cancer cells causes massive amounts of inflammatory cytokines to be released into the bloodstream. Resulting symptoms are characterized by fever and in more severe cases, renal insufficiency, pulmonary insufficiency and altered mental status. Sometimes CSS is so strong that patients die from the reaction. Again however, if patients endure/survive the treatment they often enter a state of remission or recovery.

Factors driving CSS are debated by the cancer community. Researchers more familiar with the concept of immunopathology regard CSS as an “on-target” effect of CAR T-cell therapy—that is, its presence demonstrates that active T cells are at work in the body.” In other cases however, CSS is described as a poorly understood “side effect” of immunotherapy. For example, the Washington Post recently published an article on CSS titled: “New cancer therapies have perplexing side effects.”

This “side effect” viewpoint fails to consider a growing body of research linking cancer to infection. For example, “dramatic, continual alterations in the microbiome” were directly responsible for tumor development in a model of colon cancer. Another study found significantly altered microbiome populations in human breast tumor tissue. This imbalance was correlated with decreased expression of key antibacterial response genes. Even signaling peptides created by bacteria have been shown to directly induce tumor formation.

It follows that CSS may result, at least in part, from an immune response towards infected tumor cells. Immunotherapy may also target pathogens that control tumor development by altering the activity of human metabolic pathways. If this is the case, CSS may be the cancer equivalent of IRIS in HIV/AIDS. This is supported by case histories showing that some cancer patients undergoing immunotherapy also develop “new” autoimmune/inflammatory conditions.

Immunotherapy results in Cytokine Release Syndrome (Breslin, 2007)

For example, the Washington post describes a patient named Diane Legg’s response to cancer immunotherapy, stating: ”Her therapy knocked back her cancer, and she’s glad she got it. But the drug also gave her “almost every ‘itis’ you can get: arthritis-like joint pain, lung inflammation called pneumonitis and liver inflammation that bordered on hepatitis, in addition to the uveitis.”

Did immunotherapy really “cause” Legg to develop these new illnesses? Or, as with IRIS, did the conditions arise due to the “unmasking” of pathogens acquired during earlier periods of illness? It’s also worth noting that patients undergoing immunotherapy often present with “new” bacterial, fungal and viral infections. One study identified 43 infections in 30 immunotherapy patients in the first month of treatment, with infections causing the deaths of two patients.

More research is needed to clarify how these infections correlate with CSS. To move forward, researchers developing immunotherapy treatments must be trained to understand the complexity and extent of the human microbiome capable of driving inflammation. This poses a challenge, since at the moment the immunotherapy and microbiome research communities are not well connected.

Managing CSS in cancer and IRIS in HIV/AIDS is also a great challenge for doctors administering the immunostimulative therapies. Most physicians are not trained to consider the microbiome beyond the gut. They are also not taught to understand the general concept of immunopathology. Indeed, palliative medicine has gained such traction that early trials of cancer immunotherapy did not even anticipate a CSS response. This New York Times article titled “When drug trials go horribly wrong” describes testing of an early immunotherapy treatment (TGN1412). After infusion of TGN1412, all six human trial volunteers faced CSS leading to “life-threatening conditions involving multi-organ failure.” According to the Times, “the outpouring of toxic molecules when T-cells are activated…could not have been predicted from prior animal studies using the drug.”

Physicians also suffer from a lack of solid immunopathology treatment guidelines. According to the Washington Post “Many doctors are not up to speed on how to spot and handle an immune system revved up by immunotherapy.” In severe cases of both CSS and IRIS, physicians are forced to prescribe antibiotics and antivirals to manage symptoms associated with infection. However these drugs kill only a fraction of bacteria and viruses capable of driving symptoms. This forces many physicians to “dampen down” symptoms with corticosteroids or other immunosuppressants, the use of which counters the point of treatment in the first place.

Immunostimulation can target root causes of inflammatory disease (Source: Ty Bollinger)

For immunostimulative therapies to truly succeed then, medicine would need to embrace an entirely new paradigm. Drug companies would turn their energy towards developing new antivirals, new antibiotics or antibiotic alternatives. They would attempt development of palliative medicines that help symptoms without destroying the immune response. Tests that better detect and characterize pathogens in the microbiome would be prioritized. Physicians and drug developers would incorporate knowledge of the microbiome into all treatment practices.

The success of immunostimulative therapies also hinges on the willingness of institutional review boards (IRBs) to accept immunopathology. IRBs decide whether patients are allowed to enroll in a particular drug trial. At the moment, many IRBs are unwilling to allow patients with “autoimmune disease” or non-fatal inflammatory conditions to test immunostimulative therapies. This is based on a “do no harm” mentality that does not support increasing symptoms in an effort to improve long-term health.

We must prioritize human microbiome health (Image: BioCote)

What these IRB boards may not realize is that most patients with “autoimmune disease” or non-fatal inflammatory conditions are more than willing to feel temporarily worse (even for years) if offered hope of actual long-term improvement. Serious “autoimmune/inflammatory disease” can feel like a living death. For example, a patient named Anne Ortegren recently committed suicide after decades of suffering from the neuroimmune disease ME/CFS. In a letter written before her death she stated, “A very important factor [in choosing to die] is the lack of realistic hope for relief in the future. It is possible for a person to bear a lot of suffering, as long as it’s time-limited. But the combination of massive suffering and a lack of rational hope for remission or recovery is devastating.”

This leads to a final consideration: the greatest hope for immunostimulation in ANY disease hinges on the ability of treatment to be initiated early, or in a preventative fashion. Medicine must learn to support the immune system BEFORE pathogens push the microbiome far out of balance. “Bringing back” the immune system after years of neglect inevitably leads to severe symptoms and complications. For example, the severity of CSS in cancer is directly related to the tumor burden of the patients (patients with fewer, smaller tumors experience less CSS). In contrast, immunopathology in early-stage disease can be easy to treat and tolerate.

That is why I have a vision: In this vision, medicine respects and supports the immune system from the earliest days of life (even in the womb). Microbiome health forms the cornerstone of new therapies, with treatment administered at the first sign of symptoms. New drugs that target pathogenic bacteria, viruses and fungi are central to therapy. The need for immunosuppressive medicines in chronic inflammatory disease drops…to the point where maybe, one day, they are largely regarded as a failed relic of Medicine’s past.






Interview with Robert Moir: Infection in Alzheimer’s/brain microbiome

December 18th, 2017 by Amy Proal

Robert Moir is an assistant professor of neurology at Harvard Medical School and Massachusetts General Hospital (Boston). He studies Alzheimer’s disease and other inflammatory conditions characterized by neurodegeneration. His research team has shown that the amyloid beta protein associated with Alzheimer’s “plaque” is a potent antimicrobial peptide. Please read this blog post for more context on this important discovery. 

Background information:

Antimicrobial peptides: “Natural antibiotics” created by the human immune system. They are able to kill a range of bacteria, viruses, fungi, and other pathogens.

Innate immune system: The branch of the immune system that creates antimicrobial peptides. These peptides and other innate immune system cells form the body’s “first line of defense” against infectious agents.


Robert hey! Thanks for taking the time to speak with me. First question: your discovery that amyloid beta is an antimicrobial peptide is HUGE. What made you decide to investigate its possible antimicrobial properties in the first place?

The idea came from a somewhat chance discovery. On Fridays I go through my walkabout time on PubMed (a website that catalogs scientific journal articles). I rove wherever impulse takes me. I read a paper about LL37, a well-described human antimicrobial peptide. It was obvious from this paper that LL37 and amyloid beta share clear similarities: both structural similarities and the ability to form amyloid (protein-like deposits that can accumulate in tissues in certain disease states known as amyloidopathies).

At the same time, my mentor Rudolph Tanzi (he’s in the office next door) had just gotten back results from a screen of Alzheimer’s linked genes. He got the results back literally the same day I read the LL37 study. Well, most of the Alzheimer’s linked genes he characterized were also innate immune system genes. We both looked at one another and thought: “Antimicrobial peptides and the innate immune system are the way to go.” Then we did a bunch of experiments that went nowhere. It turns out that several classical methods used to study antimicrobial peptides don’t work for amyloid beta.

In some cases amyloid beta’s ability to target certain microbes is more than 100-fold stronger than penicillin.

Then I went on vacation with my six year-old son to the White Mountains. James Kirby of Beth Israel Deaconess Medical Center had invited us, and the trip gave me the opportunity to get his feedback. He helped us develop assays to correctly test amyloid-beta’s antimicrobial activity. Key to this correct assay preparation was the fact that amyloid beta had to be in its more oligomeric forms (the biologically active form of the peptide). Once we got these assays right, we immediately found that amyloid-beta had very potent antimicrobial activity. In some cases its ability to target certain microbes is more than 100-fold stronger than penicillin.

Me: Wow very interesting. Can you explain a bit more about what it means for amyloid beta to have “oligomeric forms”?

Amyloid-beta is the chief name for a peptide that self-associates to build multiple differently shaped molecules, each with its own molecular structure and activities. Small proteins that can self-assemble and generate diverse structures like this are sometimes called “lego peptides.” Amyloid-beta assemblies are called “oligomers.” Each particular oligomer structure has unique antimicrobial activity. In this way, amyloid-beta spontaneously generates a population of diverse oligomers able to target a broad spectrum of pathogens, and even microbial toxins released during infection. This turns out to be key for amyloid-beta’s effectiveness as an antimicrobial peptide in the brain. Oligomers can target many different microbes while non-oligomeric amyloid-beta is only effective against a limited range of pathogens. This strategy is so effective that we have not been able to find a pathogen that oligomeric amyloid-beta can’t inhibit to some degree.

Me: So these oligomers are a little like antibodies? In the sense that the immune system creates a range of antibodies in response to an infectious threat, each with the ability to target different species/strains of microbe(s)? 

Yes, they are like a cheap man’s antibodies. But antibodies need whole cells in order to be produced and are metabolically complex and expensive to make. Amyloid beta’s ability to form into a wide range of different oligomers is a simpler, metabolically cheaper, and far more ancient immune strategy. Even very ancient animals like jellyfish (which are 500 million year old primitive multicellular organisms) have this ability to create antimicrobial peptides that recombine to increase molecular diversity.

Moir (right) with colleague Rudy Tanzi (left) (Image: Jon Chase/Harvard Staff Photographer).

What’s hard to believe is that many researchers still regard amyloid-beta’s generation and activities as an ‘accident’ and continue to develop therapeutic strategies around this idea. This assumption was reasonable back when amyloid-beta was discovered in 1984. The peptide was thought unique to the human brain and believed to be only generated under the disease conditions found in Alzheimer’s. In addition, amyloid-beta is made by cutting the peptide out of a larger precursor protein that is embedded in cell membranes. No other peptide was known to be generated this way, so it was considered something that must only occur under disease conditions.

However, it’s been known for over two decades that amyloid-beta is continually made in the normal brain throughout our lives – and not just the human brain. All vertebrates make amyloid-beta in their brain, most of them the exact same peptide as we make. Furthermore, the mechanism that generates amyloid-beta is now known to be a common cellular pathway that is involved in making many different important normal peptides. But, the assumption that amyloid-beta is junk persists, even though the key assumptions underpinning this idea have been disproved. I still frequently hear the ‘amyloid-beta is junk’ idea justified by an old argument that is demonstrably wrong. It goes something like this: “Since Alzheimer’s occurs in individuals past reproductive age (that is post-menopause) there is no evolutionary pressure to remove it from the genome.”

Sounds plausible, until you start looking beyond the narrow precepts of the Alzheimer’s field and consider amyloid-beta in a broader biological context. Only three species of animals undergo menopause: us, dolphins, and pilot whales. Most other animals reproduce to within 1-2% of when they drop dead – and many social animals live well into old age. And yet, all vertebrates make amyloid-beta, with over 60% making the exact same form of the peptide we do. Moreover, they have been doing this for over hundreds of million of years! (The human amyloid-beta gene is expressed in coelacanths, a family of ‘living fossil’ fish that date back over 400 million years). In fact this data would suggest the exact opposite of the ‘amyloid-beta is junk’ argument – it supports the idea that amyloid-beta contributes to survival fitness throughout life in all vertebrates.

Also, anyone with grandparents knows that they typically play an important part in helping successfully raise children. In biology it’s known as the “grandmother hypothesis.”  Simply put, the grandmother hypothesis says that menopause allows aging mothers with an increasingly high risk of death from childbirth to stop direct reproduction and pass on their genetic material to future generations by helping raise their grandchildren. Grandparents are not reproductively irrelevant!! Any gene that intrinsically causes dementia and negatively impacts grandparent survival and their valuable store of accumulated experience, is going to be selected against. The gene is going to change or be eliminated. But amyloid-beta is 100% conserved from 400 million years ago.

The human amyloid-beta gene is expressed in coelacanths (Image: American Museum of Natural History).

What all this is telling us is that amyloid beta must actually be very important. It must be at the heart of important biological processes.  Indeed, human amyloid-beta is one of the top most conserved proteins in all of biology (as far as I have been able to ascertain, second only to the protein ubiquitin).

Me: Yes it seems clear that amyloid beta must have an important role in human health and disease. How has your research moved forward based on that possibility?

We set out to test whether amyloid beta has antimicrobial activity. We found that if you add amyloid beta to a microbial broth it will inhibit or straight out kill a range of microbes. We published a paper with those and related results in 2010. It was greeted with mixed reception. Some researchers (mostly young researchers) were enthusiastic about it. Older researchers not so much. From these detractors we got feedback like, “That’s fine, but battery acid can kill organisms in a test tube too.” In other words, they were critical that the experiments were done in a test tube and not in a living organism. For most antimicrobial peptides, demonstrating potent microbial killing activity in a test tube is enough to establish identity (it’s a very difficult activity for a protein to pull off), but it’s not an unreasonable critique. Showing protective activity in a living animal is the gold standard for confirming a protein is an antimicrobial peptide.

So we set out to show that amyloid-beta could have antimicrobial activity in living organisms. We tested its activity in genetically modified mice, fruit flies, nematode worms, and cultured cells. The work took us four years to complete. Our keystone finding was that the ‘plaque’ amyloid-beta generates is as important as the peptide itself. These amyloid-beta “plaques” directly entrap and neutralize microbes. Then, by chemically generating a burst of oxygen radicals – bleach in lay terms – they destroy the trapped pathogen. Just to be on the safe side, the plaque remains intact in the brain, entombing forever any microbe that may have managed to survive. This “entrapment” mechanism is not unique to amyloid beta. For example, alpha-defensin 5 and other human antimicrobial peptides create amyloid structures that function similarly.

So to summarize, expression of amyloid-beta protects cultured cells and nematode worms from lethal infections. In unpublished work we have also confirmed that amyloid-beta also protects against infection in fruit flies. In mice, the most important animal infection model, over-expression of amyloid-beta is protective against bacterial and viral encephalitis (brain inflammation driven by infection). Finally, if you “knock out” mouse (murine) amyloid-beta the mice develop increased susceptibility to encephalitis. So what we now have is convincing data from animal models that that amyloid-beta functions in humans as an antimicrobial peptide.

Me: Has it been hard to advance these findings?

One side of the story has gotten the most attention. If amyloid-beta is an antimicrobial peptide, one plausible inference is that Alzheimer’s is caused by an infection or infectious processes. I’m kind of agnostic about that implication, in that Alzheimer’s could also be a disease where immune pathways have gone wrong. There are a number of examples of ‘sterile inflammatory’ disease in which an immune pathway has become dysregulated and pathological. No pathogen involved.

Alois Alzheimer

But having said that, there’s mounting circumstantial evidence suggesting that infection plays a role in the disease’s etiology. The first guy to support that was Alois Alzheimer himself. In the 1970s-80s many Alzheimer’s researchers thought infection played a central role. But the discovery of amyloid-beta in 1984 (ironically) shifted the focus away from infection. Amyloid-beta was assumed to be all bad, bad, bad, and its accumulation blamed for the disease. It’s a simple explanation to a complex problem, which made it attractive. But it’s increasingly at odds with emerging data. The drug company Merck’s last Alzheimer’s drug lowers amyloid beta levels but did not slow the disease.

Amyloid-beta alone does not give you Alzheimer’s disease. You also need inflammation and tauopathy, another pathology in brain. It may still be effective to control amyloid-beta if you get it early enough in the disease process. Controlling amyloid-beta may help slow the cascade of events that promotes the neuroinflammation that ultimately kills neurons in Alzheimer’s. But, the question remains: what is driving the deposition of amyloid plaques in the first place? Old models hold that amyloid-beta does it because it’s catabolic junk with an unfortunate propensity to form functionless plaques that induce inflammation. But…could plaques and neuroinflammation in Alzheimer’s actually be an immune response to a genuine immune challenge from microbes in the brain? Perhaps. If it is, then targeting microbes in the brain may be a better way to go. What is clear is that more data on the role of microbes in Alzheimer’s etiology is needed. At the moment the “amyloid is bad” idea continues to dominate and most academic efforts are still focused on this model.

Interestingly, we may be seeing the beginnings of a shift in the way “Big Pharma” is looking at Alzheimer’s. Their costly drug failures seem to be making them open to exploring alternative models, including the “antimicrobial protection hypothesis of Alzheimer’s” – which is what we are calling this new emerging model of the disease. For example, “Big Pharma” is exploring if neuroinflammation in Alzheimer’s can be dampened down independent of amyloid-beta production or plaque deposition. My colleague Rudy Tanzi has discovered that the gene CD33 is a big on/off switch for immune cells in the brain. Maybe switching the gene off could help patients manage early Alzheimer’s symptoms. Most current first-line anti-inflammatories wouldn’t have the same effect since they tend to target the adaptive immune system. Antimicrobial peptides and amyloid-beta are part of our much more ancient and primitive innate immune system (the “front-line troops” of the immune response). Companies are now pursing CD33 as a possible drug target.

Dr Doo Kim’s lab in research Unit at Massachusetts General Hospital has created a special three-dimentional human neuronal cell culture system that’s dubbed “Alzheimer’s in a dish” by the media. The system helps us scan innate immune system drug candidates. The technology accelerates drug screening and reduces cost more than 10-fold compared to conventional approaches. It should allow new potentially useful drugs to be identified much faster.

Me: Interesting. But I’m confused. Do you think amyloid beta should be removed in patients with Alzheimer’s (despite its antimicrobial activity)?

Maybe not removed, but it’s certainly a good idea to control it. Amyloid beta may be a little like cholesterol – heart attacks are exacerbated if cholesterol is in the wrong place at the wrong time. But if you remove cholesterol completely, other serious health problems arise. So as was done with cholesterol targeting therapies, I think the first goal should be to better understand what amyloid-beta is actually doing in the brain and develop strategies accordingly. That would mean reducing bad effects while preserving amyloid-beta’s role in immunity.

What may be happening is that initially, amyloid-beta rises to do battle in cases of brain microbiome dysbiosis (imbalance). Part of this response is amyloid-beta induced inflammation. But, prolonged activation of innate immune inflammation by amyloid-beta leads to tissue damage and neurodegeneration.

One thing is that any treatment aimed at managing amyloid beta would work best if administered in a preventative fashion. Again it’s a little like cholesterol and statins. After a third heart attack it rarely helps to give a patient statins. You would want to begin treating 10-20 years ahead of time to prevent this. But with amyloid-beta in Alzheimer’s we currently have no effective assays to predict who will get the disease and when to intervene. You can’t give a drug to everyone over 65.

Me: Wouldn’t it make the most sense to just target whatever infection(s) are causing amyloid beta to be produced in the first place?

Well yes, a longterm solution may be vaccination against the microbe giving you trouble. But here’s the thing about that: there are research groups that have been pushing the role of infection in Alzheimer’s for a long time, but different pathogens are identified in their studies. Herpes simplex virus 1 is a common candidate, but also chlamydia pneumonia. This suggests there is no single pathogen driving the illness. Instead, many different bugs may be involved in Alzheimer’s. For example, our studies have found that amyloid-beta has strong antimicrobial activity against the herpes viruses and these viruses are linked to increased plaque deposition, However, herpes virus are only detected in about 60% of Alzheimer’s cases. What about the other 40%?

So it’s clear that we’re not dealing with a “classical” infection.” The findings don’t support Koch’s postulates: the idea that only ONE microbe can cause ONE disease. But we’re moving away from this “classical” infection model. We’ve been taking a close look at what microbes are found in the brain and started what we are calling the ‘Brain Microbiome Project’.

Me: Your lab is also studying the brain microbiome!?

Yes. We’ve been scanning brains as part of collaborative work with Mt. Sinai. And we have found that even “non-sick” humans harbor over 200 organisms in the brain. Those numbers don’t even include the virome (viruses). And we know that a bunch of herpes viruses can also survive in the brain. There’s even vertical transmission of certain viruses in the human genome.

So infection in Alzheimer’s may be similar to what we’re seeing in conditions like Crohn’s disease. In Crohn’s the bowel is disrupted but the entire gut microbiome is involved. It’s not a single pathogen but disruption of a whole microbial community. Within that framework certain key pathogens may “push” the community out of balance and contribute to disease more than others. In Crohn’s it’s where the microbiome meets the innate immune system that things go wrong and host pathologies arise. That’s where the problem in Alzheimer’s may also lie: at the interface between microbes and a foot soldier of innate immunity – amyloid-beta.

Going back to the brain microbiome: the microbes don’t just sit there. They cooperate, they’re competitive, they interact with the host and each other: it’s a true microbiome. What this means for Alzheimer’s is there could be general dysbiosis (imbalance) of the brain microbiome. There’s a normal brain microbiome, but in Alzheimer’s something may go “out of whack” and some of the bugs go bad (they become bad players). It could be compared to ulcers. Ulcer formation is linked to the bacterium H. pylori. But H. pylori is actually also a member of the normal gut microbiome. That means H. pylori contributes to ulcers under certain negative conditions. This is tentative, but in Alzheimer’s maybe the herpes viruses also go out of control. That is the model we are exploring at the moment. 

The brain microbiome and gut microbiome communicate via the Vagus Nerve

Linked in with that model is the gut microbiome because the gut microbiome and the brain microbiome communicate a lot via the vagus nerve. There’s lots of traffic, with bacteria in the brain/gut talking to one another via this highway all the time. Some products of gut fermentation like Short Chain Fatty Acids (SCFAs) literally travel the Vagus Nerve (physical translocation). Immune cells in the brain need these gut microbial SCFAs to mature correctly. Conversely, certain bacteria in the gut live exclusively off chemicals generated in the brain that are transported to the gut. Vagus Nerve traffic may include bacterial signaling molecules called quorum sensing molecules. In this sense microbes in the gut and microbes in the brain may be “talking,” and possibly reaching decisions about what to do next.

So, what’s going on in the brain can have dramatic effects on the gut. But, the gut microbiome can also affect neurological functioning. For example, the disruption of the gut microbiome is now linked to depression – it’s a two-way axis.

And remember, because amyloid-beta can form a vast number of oligomers, it’s able to react against an large range of pathogens. So right now amyloid beta’s activity leaves open a big question mark as to the exact nature of the infection it may be targeting in Alzheimer’s or related conditions. Which means that vaccination against one pathogen in Alzheimer’s (that I mentioned as a possibility before) might prove too simple an approach. Instead, we may want to ask “Can we modulate disease progression by manipulating the microbiome and/or the gut/brain axis.”

Me: You’ve been studying amyloid beta’s antimicrobial activity against HHV6. Where is that study?

Yes, we’ve tested amyloid beta’s activity against herpes simplex virus 6 (HHV6). There are no good animal models of chronic HHV6 infection, so we are using the “Alzheimer’s in a dish” system to look at this at the moment. Each chip is like a little human brain, that is basically thinking and shooting signals back and forth. This makes the experimental model more human-like and allows for better testing. We found that HHV6 induced a large amyloid burden in the “Alzheimer’s in a dish” system within a day. Key to our findings is that these herpes viruses are “low and slow” microbes whose pathogenic activity ramps up with opportunity. This makes HHV6 extremely effective at seeding amyloid beta deposition over time, even over the course of decades. We hope to get funding from the Cure Alzheimer’s Fund to develop a Alzheimer’s disease mouse that can be infected with human HHV6 (the mice need to be ‘humanized’ for a receptor critical for HHV6 infectivity).

But you can’t formally read about the results of that study at the moment. We submitted a paper with the findings to the journal “Cell.” The journal insisted on publishing an online pre-print before the paper was formally accepted. That pre-print was critiqued by researchers who reject a role for infection in Alzheimer’s etiology – not that our study actually claimed that. It showed strong data that amyloid-beta protects against herpes in the lab. Then, Cell rejected it after 6-weeks. The situation has hurt that study tremendously. While I understand that scientific journals are motivated by a genuine belief that expediting dissemination of new and exciting data is a good thing, I would caution other research teams not to allow a pre-print to go up before the study is accepted for publication. It can go disastrously wrong and it’s not worth it. Not worth it at all. We are submitting the findings to another journal.

Having vented a little on this… I have to say that healthy skepticism is key for advancing science. My problem is not criticism – throughout my career legitimate concerns and criticisms have been invaluable for refining our ideas and showing the way forward. My problem is when strong data with no obvious flaws are rejected out of hand because they do not fit current dogma and are dismissed for perfunctory reasons.

Phew! It’s sure hard to get new findings published. With that in mind, how do you get funding for your research?

This past October, Moir and Tanzi spoke about infection/Alzheimer’s at this “Cure Alzheimer’s Fund” Symposium.

NIH funding is hard for us to get – they are somewhat risk adverse and typically fund studies that explore prevailing ideas: “Evolution not revolution.” Fortunately, we are supported by some foundations that are willing to take risks. One is the Cure Alzheimers Fund in Boston, whose founders include successful Venture Capitalists. They are used to high risk ventures. For them, if only 50% of their funded projects have a successful outcome that’s a good result – it’s much better than the success rate among new businesses! I also get funds from Good Ventures, which is part of the Open Philanthropic Project out of California.

Me: Have you read some of the recent studies that have detected a range of fungi in Alzheimer’s brains?

Yes. And amyloid-beta is strongly anti-fungal. But of course there are fungal communities in healthy brains too. The organisms we’re finding in the human brain are incredible. For example there are amoeboid worms! Up to 20% of humans harbor toxoplasmosis (the Toxoplasma gondii parasite) in the brain. We’ve even detected another worm in some of our brain samples that was previously only thought to infect dogs. Remember that the nasal bulb is a primary source of entry for these and other microbes. Interestingly, efferent nerves from the nasal bulb trace straight back to brain areas where amyloid-beta formation starts.

Have you seen the Lund University study showing that PrP (prion) protein is also a potent antimicrobial peptide?

Yes. And we’ve tested PrP’s activity in our own lab. It’s a strong antimicrobial peptide. But it goes beyond that – amyloid creation happens in a range of human inflammatory conditions. Diabetes is an amyloid disease (in both type 1 and type 2 diabetes amylin is created in the pancreas). At high levels, this amylin is toxic to pancreas islet cells and highly pro-inflammatory. But it’s also one of the most potent antimicrobials we have ever tested – it’s able to target most pancreatic pathogens, including E, faecalis, a common cause of pancreatitis. There’s also an amyloid generated in the heart that is linked to heart disease. So the potential importance of recognizing amyloid can play a normal and protective role in immunity has legs well beyond just Alzheimer’s.

Me: Do you talk to other research teams studying these other forms of amyloid?

Yes. We have multiple collaborations going on around the world. In fact I collaborate so often with other labs that I spend half my time on the phone. There’s both a collaborative and competitive atmosphere that helps drive things along. I am delighted we are no longer alone in pursuing this line of investigation.

That being said, most research teams aren’t thinking this way. And for many of these teams, accepting amyloid-beta’s role as an antimicrobial peptide is like trying to turn around an 800 pound gorilla on a dime. It’s a paradigm shift, so it will likely take years for the focus to shift.

For example, when we tried to publish our 2010 paper on amyloid beta’s antimicrobial activity, the peer review process was extremely frustrating. We submitted the paper to the journal “Nature.” From the start the editors seemed hesitant about publishing the findings. Then they consulted with an Alzheimer’s “expert”, who it seems from comments didn’t even bother to read the paper, but rejected it anyway. The most recent paper in 2016 was rejected six times without review. The reviews we did get from one top-tier journal were some of the most appalling I’ve ever read in my life. One reviewer kept asking why we did not see amyloid in our control worms – the ones that DO NOT express amyloid-beta.

After decades of rejection, Barry Marshall and Robin Warren won the Nobel Prize (Picture:

Me: That is extremely frustrating. I find it interesting that the scientific community frequently references the story of Barry Marshall and Robin Warren. The two researchers discovered that h.pylori bacteria plays a key role in driving ulcer formation (during a time when ulcers were believed to be caused only by stress and spicy foods). When they first presented these results, they were ignored and in some cases even mocked by other research teams. It took decades for their new finding to be taken seriously. Then, in 2005 they were awarded the Nobel Prize. The story is often referenced as though the scientific community has learned from this experience and is now more open minded. But when I look at how your work has been received thus far I’m not sure I see much progress.  

Yes. Also, Barry Marshall was my old microbiology instructor when I studied at the University of Western Australia. He did struggle with recognition for a long time. There are all sorts of urban legends about how he kept the research going in the face of the wall they kept butting up against.

Me: True. And final question: What studies are you planning to do next?

In a next study, we hope to look at the actual microbiome in the brain. Then remove amyloid plaque from this tissue and see if we can identify the exact microbes the plaques have trapped. We’ll do this by seeing if we can recover the genetic material of the seed microbe.

Me: That’s amazing. I can’t wait to see what you find. I’m going to let you go because I want you to get back to work immediately:)


Cholesterol, fat, and human metabolism: a microbiome-based paradigm shift

December 15th, 2017 by Amy Proal

At the age of 64, after a morning playing golf, president Dwight D Eisenhower had his first heart attack. As Pulitzer Prize winning author Gary Taubes describes in his book “Good Calories, Bad Calories” Eisenhower’s heart attack “constituted a learning experience on coronary artery disease (CAD).” After the event, his doctors, considered the top experts in the field, gave the public a “lucid and authoritative description of the disease itself”, followed by twice-daily press conferences held on the president’s condition. Soon, most of America, particularly middle-aged men, became intently aware of dogma promoted by these “experts”: the notion that CAD is caused by eating foods high in fat and cholesterol.

Like much of the rest of the nation, Eisenhower began to avidly lower his fat and cholesterol intake. Yet this plan of attack was counterintuitive for Eisenhower who, before his heart attack, had none of these supposed risk factors connected with CAD. His blood pressure was only seldom elevated, his weight throughout his life remained around 172 pounds (considered optimal for his height). His total cholesterol was below normal – his last measurement before the attack was 165 mg/dl, a level that heart disease specialists today consider safe. He had even quit smoking six years earlier in 1949.

President Eisenhower after his first heart attack.

After gaining four pounds, the ex-president reduced the amount of food he ate for breakfast, then eventually sacrificed lunch completely. His doctor was mystified at “how a man could eat so little, exercise regularly, and not lose weight.” Eisenhower then renounced butter, lard, and cream. But despite these additional dietary changes his cholesterol levels began to rise. “He’s fussing like the devil about cholesterol,” wrote his doctor. “He has eaten in the last week only one egg, one piece of cheese.” It got to the point where Eisenhower’s doctor started to lie about his cholesterol levels in order to keep the president calm. At one reading, Eisenhower was told his level was 217 when it was actually 223. On his final day in office, he was made to believe his cholesterol was 209 when in reality it had soared to 259. Finally, in 1969 at the age of 78, Eisenhower died of heart disease. By that time he’d had six other heart attacks.

We now know that the medical experts advising Eisenhower and the American public were wrong. In CAD, lipids, or fatty molecules, do accumulate in the arteries of patients with the disease. Indeed, CAD results from atherosclerosis: a gradual hardening and clogging of the arteries due to lipid accumulation. However, a growing body of research demonstrates that these lipids are not sourced from dietary fat and cholesterol. As Taubes describes it, the long held dogma about heart disease, in which cholesterol clogging the arteries and excess body fat are viewed as culprits, “as though the fat of a greasy hamburger were transported directly from the stomach to the artery lining” is no longer support by scientific evidence.

“Consensus thinking” led to incorrect CAD guidelines

It’s worth reading Taubes’ full book to better understand the scientific climate that, over the past decades, incorrectly linked to heart disease to dietary fat and cholesterol. For one thing, regulatory bodies were attracted to the simplicity of the “dietary fat/cholesterol clogs arteries” disease model. It was easy to communicate and implied that basic nutritional guidelines could prevent the illness. The model was also promoted at numerous “consensus” conferences: large meetings at which evidence to the contrary was often blatantly excluded.

For example, the largest diet-heart trial ever carried out in the United States was not included in medical or political debates about the best diet for the America public. Because the results opposed what was becoming the consensus view on diet and CAD, they went unpublished, (they were later published in a small cardiology journal that very few people read). The trial, which included 9,000 residents of various mental hospitals found that men on a low-fat diet had a slightly lower risk of heart attacks, although women did not. Overall, patients who had eaten a low-cholesterol diet were associated with a greater risk of heart disease.

Ancel Keys on the 1961 cover of Time Magazine. There, he advanced the idea that dietary fat “clogs the arteries”

A scientist named Ancel Keys played a central role in perpetuating the belief that dietary fat/cholesterol clog the arteries. He famously linked dietary fat to heart disease after studying seven distinct populations around the world who ate diets relatively low in fat and also seemed to have a lower incidence of CAD. However, researchers at the University of California, Berkeley later found that Keys had chosen only six countries for his comparison though data was available from 22 countries. When all twenty-two were included in the analysis, the link between fat and heart disease vanished.

In fact, Keys theory implicating diet as the cause of heart disease appeared on the cover of Time Magazine in 1961 despite the fact that at the time, only two studies had directly tested the connection. One of these studies actually proved Keys wrong. It was a British trial, in which the fat content of the meals of a group of men who had previously suffered from heart attacks was reduced to 1/3 of its previous level. A control group continued to eat a normal diet. After three years the average cholesterol levels dropped from 260 to 235, but the recurrence of heart disease in the control and experimental groups was essentially identical. “A low-fat diet has no place in the treatment of myocardial infarction,” the authors concluded in 1965 in the Lancet medical journal.

Recent studies continue to disprove the “dietary fat/cholesterol” model  

Lipids are “fatty molecules”

Despite studies like that described above, the “dietary fat/cholesterol clog arteries” model for CAD became so popular that debate on the topic continues in 2017. Many physicians and policy makers still issue “heart-healthy guidelines” urging Americans to lower dietary cholesterol/fat despite ever-increasing evidence to the contrary.

For example, a 2016 study by researchers in Finland found that a relatively high intake of dietary cholesterol was not associated with an elevated risk of coronary heart disease. Another meta-analysis found no association between consumption of saturated fats and either coronary heart disease, ischemic stroke, type 2 diabetes, death from heart disease or early death in healthy adults. A different study even demonstrated an inverse association between saturated fat and stroke (i.e. those who ate more saturated fat had a lower risk of stroke).

The authors of the first meta-analysis conclude their paper by stating: “Coronary artery disease pathogenesis and treatment urgently requires a paradigm shift. Despite popular belief among doctors and the public, the conceptual model of dietary saturated fat clogging a pipe is just plain wrong.”

In 2014 Time Magazine reversed its position on dietary fat and CAD

In 2014, even Time Magazine published a cover story reversing its position on Ancel Key’s earlier claims. The article was titled: “Eat butter. Scientists labelled fat the enemy. Why they were wrong.”

So where do the lipids (fatty molecules) driving CAD come from?

This begs the million dollar question: if lipid accumulation in CAD is not sourced from dietary fat and cholesterol, then where do the lipids in arterial plaque come from!? The results of a seminal study by researchers at the University of Connecticut provide a novel “answer” to at least part of this question. Indeed, the team’s findings are so important that they are positioned to change the future of heart disease research.

The team, led by Frank Nichols, found that two important forms of lipid detected in diseased artery walls are created by bacterial members of the human microbiome. More specifically, both lipids (Lipid 430 and Lipid 654) are made by bacteria from the phylum Bacteriodetes: a family of bacteria so common to the human body that they represent 1/3 of bacteria currently able to be cultured from the human intestine.

The team arrived at their results by analyzing lipids in atheroma collected from patients at Hartford Hospital in Connecticut. They identified bacterial Lipids 430 and 654 in the samples, and distinguished them from human lipids by analyzing their chemical structure. Both Lipid 430 and Lipid 654 contain fatty acids with branched chains and odd numbers of carbons (a structure not typically associated with human/mammalian lipids).

In a press release about the study, Nichols stated the following about Bacteriodes bacteria: ”I always call them greasy bugs because they make so much lipid. They are constantly shedding tiny blebs of lipids. Looks like bunches of grapes.”

Recovery of Lipid 430 and Lipid 654 in common Bacteroidetes bacteria.

Interestingly, both Lipids 430/654 were shown to activate TLR2 – a protein at the heart of the immune system’s response towards foreign/infectious threats. This suggests that the human immune system recognizes Lipids 430/654 as foreign, and mounts a sustained inflammatory response to their presence. These results “jive” with the fact that atherosclerosis is now understood to be a serious chronic inflammatory disease.

Nichols and team contend that Lipids 430/654 are created by Bacterioides bacteria in the gut/mouth. Under such conditions, lipids produced by these microbes would travel to the arteries via the bloodstream. However, additional research is needed to confirm that bacteria capable of living directly in the arteries don’t additionally contribute to lipid creation. For example, one study found that periodontal bacteria in the mouth can directly invade human arterial skin cells in culture.

In addition, Stanford researcher Stephen Quake recently reported the presence of thousands of previously undetected microbes in human tissue/blood. Many of these “new” microbes may also produce lipids capable of collecting in the arteries. It’s also worth noting that dead white blood cells (macrophages) are often found near lipids in arterial plaque. Since many intracellular pathogens directly infect macrophages, these dead cells may represent those parasitized by infectious agents.

Bacterial lipids may disrupt human metabolic feedback pathways

The accumulation of bacterial lipids in human arteries helps account for the “plaque” and chronic inflammation characteristic of atherosclerosis. However, bacterial lipids may also promote various forms of disease by interfering with human metabolism.

An extremely important paper by Tony Lam and team at the University of Toronto details feedback pathways that control hormonal signaling between the human gut and the human brain. These pathways help the body regulate appetite control, energy expenditure, and other important mediators of human metabolism. Lam’s paper shows that, in conditions like diabetes, glucose levels are regulated by a complex network of these pathways – many of which sense hormone/nutrient production in the gut and relay this information to the brain via the nervous system.

Gut peptide hormones and regulatory signals are released along the length of the gastrointestinal tract. (Lam et al)

Most of the pathways described in Lam’s paper are too complex to detail here, but the CCK pathway provides a good example of their general function. CCK is a gut hormone released following a meal. It acts on local gut receptors to help the body regulate glucose levels. CCK is secreted in the small intestine in response to fatty acids or lipids. It conveys information about these lipids (amount, composition, structure) to the brain. The brain integrates this information with signals arriving from related pathways and signals back to the gut to adjust glucose levels accordingly.

Under conditions of health, the CCK pathway is tightly controlled by the human body. It is governed by a series of “checks and balances” that help keep glucose and other hormones/nutrients in a normal range. This begs yet another important question: what might happen to the CCK pathway (and related pathways) if foreign, bacterial lipids accumulate in the body?

While there are differences between human and bacterial lipids, the fatty molecules created by both species also share many common chemical structures (molecular mimicry). For example, the Nichols study found that Lipids 430/654 are similar enough in structure to human lipids that they can be “broken down” by the same human enzyme (PLA2).

This means that human signaling molecules may be able to “sense” the presence of bacterial lipids. In the case of the CCK pathway (in which CCK is created in response to lipid concentration) this could significantly change the information the hormone conveys to the brain: in simple terms, the brain would begin to adjust glucose levels based off a sum total of human lipids AND bacterial lipids.

Many of the Bacteriodes bacteria that create lipids are major human pathogens – eg. members of the genera Prevotella, Tannerella, Capnocytophaga and Porphyromonas (a driver of to tooth decay). If the human brain integrates lipids created by these pathogens into the input it uses to adjust glucose levels and other aspects of human metabolism, serious illness may result.

In summary, there are three main ways bacterial lipids can drive human heart/metabolic disease:

  1. Bacterial lipids are created (or travel) to human arteries where they may contribute to formation of arterial “plaque.”
  2. The human immune system reacts to these foreign bacterial lipids, which results in sustained, chronic inflammation.
  3. Human feedback pathways may incorrectly “sense” foreign bacterial lipids, and factor them into signals controlling human glucose levels and other aspects of human metabolism. This would “throw off” or “mess with” these human pathways in a manner that promotes disease.

Imagine the satisfaction of going back in time to tell President Eisenhower about the Nichols and team findings. While preliminary, they suggest that he could probably relax, eat a second egg, and feel a little less crazy. The same goes for millions of patients around the globe, who must “re-learn” basic heart disease guidelines – but in the context of research that may better address the “root cause” of their symptoms.

Interview with neuroscientist Michael VanElzakker: Vagus Nerve, ME/CFS, latent infection and more

December 7th, 2017 by Amy Proal

Michael VanElzakker Phd, is a neuroscientist affiliated at Massachusetts General Hospital, Harvard Medical School, and Tufts University. He has two primary research interests: PostTraumatic Stress Disorder, or PTSD, and Myalgic encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS). He recently published a paper describing a novel hypothesis for ME/CFS that centers on infection of the Vagus Nerve.

Note: This interview was transcribed from written notes and not an audio recording.

Background information:

Hypothesis: a proposed explanation made on the basis of limited evidence as a starting point for further investigation.

The UK PACE Trial on ME/CFS: A highly criticized trial on ME/CFS. The Trial’s results don’t fit with what most ME/CFS patients experience in real life. Please read journalist David Tuller’s detailed critique of the trial.

Unrest: A 2017 Documentary about ME/CFS. Summary: “Jennifer Brea is about to marry the love of her life when she’s struck down by a fever that leaves her bedridden. When doctors tell her “it’s all in her head,” she turns her camera on herself and her community as she looks for answers and fights for a cure.”


Me: Hi Mike, thanks for taking the time to talk! How are you?

Mike: No problem. I’m good, actually working on setting up a second screening of “Unrest” here at the Martinos Center. We’ll show the documentary on a big screen in one of the conference rooms. Have you seen it?

Me: Yes, it’s very good. My story has a lot of parallels with Jen’s. It accurately portrays a lot of what I’ve dealt with for over 12 years. It’s very painful for me to watch. How do you feel about it?

Mike: It’s great. I’m glad they showed the story of Karina Hanson being taken away from her parents – that’s something only the ME/CFS community knew about for many years. It was sort of surreal to see the story presented on a big screen. It really adds to the film.

Me: Definitely. Also Mike, before I forget, can you clarify your different jobs? You work at both Harvard and Tufts?

Mike: The office I’m sitting in now is at the Martinos Center for Biomedical Imaging, which is part of Massachusetts General Hospital. Mass General is the teaching hospital for Harvard Medical School, so I get a semi-automatic Harvard Med affiliation with my position. But I’m almost never on the main Harvard campus.

Me: Cool, and you teach at Tufts?

Mike: Yes, I’m an adjunct instructor. This semester, I’m teaching a class on research methods that’s required for undergrad clinical psychology majors – the idea is that it will help them to better think critically about the scientific literature by being able to understand methods. I also teach “abnormal psychology” to clinical psychology majors. And next semester I’ll teach a seminar that focuses on the intersection of stress and memory.

Me: Interesting. I’m glad you’re teaching research methods so that your students might learn how to not set up a trial like the UK PACE Trial on ME/CFS.

Mike: I know. In these classes, I work in ME/CFS as a subject as often as I can. And I have used the PACE trial as an example in the research methods class. First to show the importance of choosing reliable/realistic study outcomes. Also though, I’ve pointed out how the PACE trial team went on a press tour with claims that went well beyond their data. I feel that the impact of PACE on patients would not have been so negative if the team had stuck to talking about what they actually found (in terms of results). But they decided to tell the popular press that they discovered how to make people with ME/CFS recover. Again, a claim that’s just way beyond their actual data, even setting aside critiques of how they arrived at those data. Also, in the abnormal psych class I talk about how in some cases psychosocial models for disease aren’t as strong as many people assume and that many conditions that were once thought to be psychosomatic are now understood to not be.

Me: Yeah, the metrics they set for “recovery” in the trial are pretty ridiculous.

Mike: Yes, not to mention that they sent out a biasing flyer in the middle of the trial to all participants, a trial based in subjective measures. Quotes from selected patients saying things like “I feel good” or “This is helping me.”

Me: That’s crazy, I didn’t know that. What an incredible form of bias (once subjects read what other subjects say it can influence how they might rate their own experience).

Mike: Yes it sometimes looks like the PACE team so strongly believe their hypothesis that they can’t see perfectly plausible alternate interpretations of their data. And the ME/CFS UK atmosphere seems to be particularly politically charged, more so than here.

Me: The whole thing is very frustrating. I get particularly annoyed because I’ve now found doctors, friends etc. who take my ME/CFS really seriously. So in my own case, I’m not forced to deal with the “psychosomatic” mindset anymore. Which has freed me up to spend more time learning/reading about actual biological dysfunction in ME/CFS. I wish it could be that way for everyone.

Mike: Yes, it’s frustrating for me in that sense too. When joining the ME/CFS research community I had to make a decision: to put my head down and just do science, or to also really make an effort to interact with and advocate for patients. I decided to advocate for patients, and in order to do that I’ve had to spend hours reading about PACE and digging into related critiques (because that’s what patients are dealing with day-to-day). But that’s time I could spend researching the biology. Also the 5 million that went to PACE: I could do a lot of really relevant/important ME/CFS research with that money.

In terms of doctors, ME/CFS patients have to do a lot of work to find a doctor. Few doctors are familiar with the condition. Sometimes I’m asked “Who’s a good ME/CFS doctor in Boston”, and I don’t know someone to recommend. I know well-intentioned physicians, but they haven’t necessarily kept up with the ME/CFS literature. Also, I think doctors find it harder to accept ME/CFS patients because it takes more time per patient to figure out what might be going on. As we know, its certainly not a situation where they can just prescribe an ME/CFS pill. So right now there isn’t much of a place for the condition to “sit” in the medical world – should it be in infections disease? or neurology? – and we’ll likely see ME/CFS shift and change between specialists for a while.

For example, right now I’m collaborating with a pulmonologist to run exercise tests on ME/CFS patients. It’s a collaborative study where we’ll do patient brain scans before and then after exercise testing (with a focus on tracking changes in the autonomic nervous system). But even this great pulmonologist doesn’t exactly specialize in treating ME/CFS patients, specifically.

Also medical textbooks need to be updated to describe ME/CFS more accurately. In my abnormal psychology class I’ve made a point of showing my students how our own textbook poorly describes the illness. The textbook description isn’t terrible, but it could be way better. There’s mention of ME/CFS in the section of the book that gets into somatoform disorder (a mental disorder that manifests as physical symptoms). Somatoform disorders are a real phenomenon, but is probably vastly over-diagnosed. So I’ll tell students, “Somatoform disorders are real but they ought not to be the first place doctors look!…and should not form the default thinking on any disease. Also it’s not OK to resort to assuming someone has a somatoform disorder because the blood tests you’re running don’t show clear results.”  The influential psychiatrist Allen Frances, the primary author of the previous DSM (the manual of psychiatric diagnoses), recently stated that somatoform disorder should not be the default diagnosis for unexplained physical symptoms, and I agree.

Returning to the textbook we use in class, I actually met the textbook author at a conference. We got a drink. In a very collegial way I pointed out how the book’s description of ME/CFS could be updated. For example the XMRV controversy that the book mentions is behind us in my opinion. I’m not sure how seriously the textbook author took my feedback but he sounded sincere and said thank you.

You know what has also been frustrating lately? It seems like each new press release about ME/CFS research states, “Our findings put to rest the idea that ME/CFS is a psychosomatic illness” But IMO, research has existed for decades showing ME/CFS is a biological disease. So by mentioning the psychosomatic theory for ME/CFS we are actually giving it credit. I think we should act like it’s not even worth referencing.  

Yes. I’ve realized I need to change my own approach. In the past I’ve been using the standard ME/CFS intro sentence when writing about it: Something like “Chronic Fatigue Syndrome is a poorly understood, crazy complicated disorder/mystery that has everyone completely baffled.” But over the last months I’ve realized that description is just untrue. Of course not everyone agrees on what triggers the illness, but in some cases we kinda do. Many patients (if you ask them!) can tell you how their symptoms started (I got Mono, I got sick when traveling in Colombia and never got better, etc.). It’s hardly like we’re dealing with a complete black box of information. Also, there’s “mystery” surrounding almost every human chronic disease. For example in cancer, we don’t understand why cancer cells can’t stop replicating. But we don’t describe cancer as a hopeless mystery. So I’m going to stop using that kind of language. I feel like it intimidates clinicians/researchers and adds to an aura of debate (“is this psychosomatic or not?”) that’s not justified. It also gives clinicians an excuse to throw up their hands up and say, “It’s just too hard to treat patients with ME/CFS.”

Me: That’s great. I hope other research teams follow your lead. Also, considering this climate, what made you willing to research ME/CFS?

Mike: I have a friend with serious ME/CFS. I was a late user of the internet but when I got on Facebook I reconnected with people I hadn’t talked to for years. I messaged a high school friend of mine, “What have you been up to?” It turned out she had been forced to drop out of law school because of ME/CFS. She was sick as hell. And before having to drop out she was so talented that she was set to graduate early. She’s an engaging person and I could feel her frustration about being sick. Also anger at dealing with being told she had the “yuppie flu” and other misconceptions. This is a bright, motivated person and I could see her devastated by the illness.

At the time I was a University of Colorado undergraduate student. My research focused on stress/memory in PTSD (still my second life!). But I had friends doing neuroimmunology research. I would go to those friends’ presentations and talk to them about neuroimmunology. I also found myself brainstorming about my sick friend. I formed the basis of my Vagus Nerve hypothesis for ME/CFS. Then I chatted with Linda Watkins, a professor in the Department of Psychology and Neuroscience. She’s an esteemed researcher who’s done great work on the intersection of the immune system and the nervous system. She even won the Spanish version of the Nobel Prize. I spoke with her and asked, “What would happen if a neurotrophic virus got right into the Vagus Nerve? Could that look like ME/CFS?” She’s not a hyperbolic person but she seemed really stunned and told me, “Oh my God, all it took was for someone to put it together.” That moment has carried me through a lot of frustration and disappointment as I tried to get people to listen to the idea and follow up.

Me: I love how you listen to your friend and to other patients and use their feedback to help inform your research

Mike: Yes, there’s an obnoxious strain of elitism that often impacts ME/CFS patients. It centers on the idea that patients are fooled by their own silly brains. I feel the opposite way. For example, I have great appreciation for my friend with ME/CFS. I constantly ask her very personal questions about her body/symptoms, and she’s totally willing to give me feedback. To act this way your have to be willing to step outside typical institutional roles in which people like me are the real experts. You have to be willing to think, “This patient is more than likely just as smart as I am. I’m going to interact with this person as a thoughtful, critical human who has real insight into their condition.”

Me: Yes! Before we go on, could you explain the ME/CFS hypothesis you’ve published? It centers on infection of the Vagus Nerve…

Mike: Sure. So when an otherwise healthy person gets sick with pretty much anything, many of the symptoms they experience will be similar no matter what specifically is making them sick. For example, people with both Strep and the flu feel extra tired, achey, and have problems with lack of concentration, loss of appetite, reduced sex drive, etc. This is true despite the fact that the flu is  a virus and Strep is a bacterial infection. These similar symptoms occur as part of the body’s general “sickness response:” a response driven by the innate immune system (the more ancient branch of the immune system that first recognizes and combats infection).

These “sickness response” symptoms overlap with important primary symptoms of diseases like ME/CFS. This suggests that a sustained and exaggerated, “sickness response” may be a core component of ME/CFS. How might this come to pass? When the innate immune system detects pathogens (microbes causing disease), it generates cytokines (signaling proteins) in an effort to target and draw attention to the infection. These cytokines are created locally at the site of infection (they don’t always make it into the general bloodstream). Still, the brain is able to perceive and respond to this cytokine activation. So how does this work?

Cytokines are relatively large lipophobic polypeptide molecules, so they don’t easily cross the blood/brain barrier. Instead the brain senses changes in cytokine activation via a nerve that conducts signals from the body to the brain, the Vagus Nerve. The Vagus Nerve is an extremely important conduit between the nervous system and the immune system (especially because it densely innervates parts of body in contact with outside world like the lungs and stomach).

A number of key pathogens such as the herpes viruses and the bacterial species Borrelia burgdorferi (Lyme disease) particularly like infecting nerve tissue (they are neurotropic). The hypothesis I put forward in my paper posits that, in ME/CFS, a number of different possible pathogen(s) may directly infect the Vagus Nerve. Resulting cytokine signals relayed to the brain then cause a sharp rise in “sickness response” symptoms. Because the signaling is directly on this sensitive nerve, the brain “overreacts” to these signals and sort of perceives that the entire body (and not just the Vagus Nerve) is infected with pathogens. If this is the case, these “sickness response” symptoms will be exaggerated. In addition, the signaling may not stop correctly, and could induce a snowball effect “feed forward” loop that sustains chronic symptoms. This would be similar to the way a war veteran might still sense pain in an amputated toe (inflammation has triggered a cascade reaction that can sustain itself).

If I were to update the hypothesis today, I would clarify that the Vagus Nerve also plays a large role in relaying information about the human microbiome to the brain, and adapts signaling based on microbiome composition/activity. I would also add rat/mouse studies showing that a pathogen doesn’t have to be fully “active” for nerve and glial tissue to sense its presence. Many viruses persist in latent, chronic forms that aren’t detected well on blood tests but could still be contributing to symptoms.

Me: I love the emphasis on infection of nerve tissue. I’m not sure many people realize how easily nerves can become infected. But question: is it possible that, in ME/CFS the Vagus Nerve is infected, but other chronic pathogens are also infecting other areas of the body (immune cells in the blood, etc.). In that case, the inflammation that triggers “sickness response” by the brain via the Vagus Nerve might not be exaggerated, but actually correct?

Mike: That’s totally possible and something we should also be considering. In order to test that possibility we could look for glial cell activation in the periphery (areas farther away from the central nervous system). Maybe we could add LPS (molecules that trigger a response towards infection) into the periphery and see how Vagus Nerve signaling is impacted. But those are early ideas, and we’d have to sit down more seriously to really tease things apart.

Me: Aren’t most microbes in the body technically “latent” in that microbiome populations are acquired in the womb and often persist with us throughout our lives?

Mike: I was talking more about acquired pathogens. But I agree that the microbiota count as organisms that survive for long periods in the body. And if components of the microbiota begin causing inflammation then the nervous system will likely detect it, relay it to brain, and trigger “sickness behavior” and other related responses.

Me: I see. What factors made you decide to formally publish your hypothesis?

I was a grad student at Tufts working studying PTSD when my advisor went to Korea for a few weeks with her family, so I had a little extra time. I thought, “Why don’t I just write this up?” – I had put a lot of text together from my previous life when I was working in rat labs. But at the time I didn’t intend to get into the ME/CFS field. I thought that if I wrote up my hypothesis, and better explained some info on how cytokines work etc., another research team might take the idea and run with it.

Me: I take it that didn’t happen?

Mike: No. A lesson I learned is that if you want something done you kinda have to do it yourself. Other people might become organically interested in your hypothesis but are unlikely to pursue it as doggedly as someone personally invested. Now I find ME/CFS very interesting to work on, but it’s very different than studying PTSD. In the PTSD research community there exist differences in opinion but most of us are pulling on the same rope. Of course there are controversies, but the atmosphere is not as politically charged as with ME/CFS, where there is broad, controversial argument about what the disease even is. And with ME/CFS I spend a lot of my energy doing patient advocacy work, communicating with patients, etc.

I actually think that part of what I’ve tried to contribute to ME/CFS is being able to talk about this condition while having fancy institutions (Mass General Hospital etc.) listed next to my name, which seems to help ME/CFS legitimacy. It’s a little crazy that thanks to the building I sit in, I can get people to listen to some of the things patients have been screaming for decades.

Me: I think it’s great you were willing to form a solid hypothesis about what causes ME/CFS. When I publish on ME/CFS I also tie my insight to a specific hypothesis. I think doing so makes what I put forward easier to follow, test and even discredit if necessary. But many research teams seem nervous about publishing hypotheses. Why do you think that is and should that mindset change?

Mike: Scientists generally ought to be at least trying to tie their data to a hypothesis. I do wish the field would put more effort into looking at how their data relate to existing hypotheses. Most papers are currently published as phenomenology (aka “this is happening in people’s bodies”), but many lack a framework that attempts to explain why things are happening. That makes it much harder to decide what direction to move in next. In ME/CFS we’ve known for years that metabolism, the immune response, the neurological system are off. It’s time to get more into the specifics of why this might be happening.

Me: Your specific hypothesis aside, I also like the way you wrote your recent ME/CFS paper. You use “simple” language that patients can understand, and went out of your way to define difficult terminology. 

Mike: Yes, in the original draft I was writing largely for patients. I actually included way too much background and it got too long, so I dialed it back. But my writing was very much intended for people who didn’t already know the stuff. When I look at the ME/CFS research community almost everyone involved with the disease is personally impacted. So they didn’t set out to formally study ME/CFS from the get-go. This leads to situations where some orthopedic surgeon with a sick cousin decides to research ME/CFS. Such people may need more context for areas outside their training.

That being said, a lot of the simplified language in the paper is a result of the peer-review process. It was the most contentious review process I’ve ever had – some of it was really condescending and smug. The responses had “ou” spellings so I wondered if one of my reviewers was a UK-based psychiatrist. I had to lay stuff out so this reviewer would understand. For example, I created the “terms table” in the paper as a response to the reviewers seeming to not know some of the terms.

I also got a comment from a reviewer commenting on a perceived misuse of an anatomical term stating, “The author seems to have a lack of medical schooling.” And I had to respond, “The reviewer may be unfamiliar with the spinal cord literature” because I was using the term correctly. The reviewer even said they ended up reading an enormous amount of extra literature to follow what I was saying. I appreciate that they did that, but the whole thing took a lot of time.

Also, my original draft didn’t mention the PACE trial. But I got some feedback from a psychiatrist who convinced me to write a section about it. At the time I didn’t really appreciate the depths of the problems in that study, most of which are not visible from only reading the actual publication. I saw it was published in the Lancet (a very highly regarded journal) and assumed it had undergone a rigorous peer review. Now I regret that.

Me: In your paper you mention possible treatment options that fit your hypothesis. How has that been received?

I wanted to be careful talking about treatments because I know that desperate patients sometimes run out and try new things based on very scant evidence. I mentioned antivirals and I have seen some people improve by taking them. But treatment is probably going to require critical thinking and a personalized approach. I don’t think there will ever be an ME/CFS pill – instead we’ll have to think about mechanisms and what’s going on with individual patients. Looking at a patient’s history is important (for example one patient might have had bad chicken pox as a teen, another Mono, another may have gotten sick after traveling). It will probably have to be systems that we target. That can make clinical trials super messy, with some treatments hard to test in a standard randomized placebo controlled trial in which all patients receive the same exact intervention.

We may also need separate treatment interventions (or treatment cocktails): treatments that attempt to get rid of possible infectious agents and others that could help patients manage the putative excess inflammation itself. When it comes to therapies that target the Vagus Nerve, Transcutaneous Vagal Nerve Stimulation is promising. The treatment targets the branch of the Vagus Nerve that comes close to the outer ear and allows action potentials to be sent down the nerve. This stimulates the normal anti-inflammatory reflex of the Vagus. I know a handful of ME/CFS patients who’ve been playing around with this Vagal stimulation. Some have felt better, some have noticed no effect, and others have suffered from side effects. So there’s a spectrum of responses but it’s one arrow in the quiver to think about.

Me: I’m a big fan of how you share information on Twitter. In fact, you recently asked for brain scan donations in a Tweet. I think that’s awesome. It helps people donate money towards a specific goal with an outcome that’s easy to follow

Yes, so far my salary as a post doc has been paid by donors. These donations are sent to Mass General and are enormously helpful. I’m really appreciative. But it’s also so hard to start a research position from scratch. Normally as a post doc I would enter an already-established lab that’s already doing the kind of work I want to do. But I’m starting something totally new with my ME/CFS research. It’s a stupid amount of work:) For that particular project I want to run a handful of brain scans to get good pilot data. Just five scans would be enough, so that’s what I am trying to raise. And even if donations trickle in slowly, when I reach $5900 I can do the scan immediately. That’s opposed to applying a formal grant for millions of dollars and waiting long periods for a decision.

It’s a bit of a Catch-22 because it’s hard to apply for big money without data, and hard to get data without money. If I do the scans with private funding I can use my pilot data to hopefully get a larger government grant. I want to call the NIH’s bluff and see if they really are willing to fund research here. Francis Collins and the NIH seem earnestly invested in helping ME/CFS. So I’m gonna get good pilot data and say, “Here you go!” Now really fund this:)

Me: I’m changing the subject a little, but since you work with PTSD, what do you think about the use of psilocybin as a treatment? I’ve read several fascinating articles on early trials of psilocybin in cancer (it dramatically improved patients’ emotional health)

In the PTSD field, psilocybin research has not advanced as far as MDMA research (MDMA is also called ecstasy). For patients with PSTD, MDMA can certainly help them feel good and help them talk/open up. But it seems as though that’s not the only mechanism at play. It’s also been shown to enhance fear extinction learning in rodents. It’s very worth studying but it’s important the drug is administered in a safe context. Both psilocybin and MDMA are fascinating areas of research. It’s too bad the USA still has a strong drug war mentality that makes conducting studies with these drugs much harder to do.

Me: Can you test for prions in patients with ME/CFS? In my last blog post I wrote about their antimicrobial activity (a topic I find fascinating).

Mike: No unfortunately my lab isn’t set up to detect prions. But I’ve been chatting with some folks about prions, here and at MIT. The antimicrobial activity of both prions and amyloid beta is kinda a million dollar idea. I’ll also send you a recent study showing that bacteria themselves can create prions.

Me: Aw too bad you can’t test for prions but send me that study!

Mike: OK

Me: I should probably let you go…you have lots of work to do! Thanks so much for taking the time to talk.

Mike: No problem. Take care:)

Antimicrobial activity of amyloid beta and PrP in neurological disease: a paradigm shift

November 17th, 2017 by Amy Proal
Amyloid beta (brown) forming near a nerve cell (pink) of a patient with Alzheimer's (Graeber et al)

Amyloid beta (brown) forming near a nerve cell (pink) of a patient with Alzheimer’s (Graeber et al)

Many neurological conditions are characterized by the formation of proteins or “plaque” in brain tissue. Two such proteins are amyloid beta and prion protein (PrP). Amyloid beta forms the “plaque” associated with Alzheimer’s disease. PrP has been detected in the brain/nervous system of patients with Parkinson’s Disease, schizophrenia, bipolar disorder, and even major depression.

Most of the scientific community currently regards both amyloid beta and PrP as drivers of neurological inflammation and disease. In Alzheimer’s, amyloid beta “plaque” is believed to be a useless substance that promotes symptoms and degeneration. PrP is best known for its association with Mad Cow Disease. In Mad Cow Disease and related “prion disorders” PrP protein is believed to fold incorrectly. This “misfolding” allows PrP to better access brain tissue, where it is also believed to cause symptoms and dysfunction.

However, several recent studies by researchers at Massachusetts General Hospital (Harvard) and Lund University Sweden challenge the above assumptions. In fact, their groundbreaking research calls for a complete re-evaluation of the role amyloid beta and PrP play in neurological disease. The Harvard/Lund studies demonstrate that both amyloid beta and PrP have a previously undiscovered function: they are potent antimicrobial peptides. Antimicrobial peptides are natural, broad spectrum antibiotics created by the body that destroy bacteria, enveloped viruses, fungi and even transformed or cancerous cells.

How does this change the game? If amyloid beta and PrP are antimicrobial peptides, they serve a protective role in patients with Alzheimer’s, Parkinson’s, and other conditions. Both proteins almost certainly form as part of the immune system’s response to infection in patients with these conditions.

Amyloid beta

The antimicrobial activity of synthetic forms of amyloid beta and LL-37 were determined as minimal inhibitory concentrations against 12 microorganisms (Moir et al)

The antimicrobial activity of synthetic forms of amyloid beta and LL-37 were determined as minimal inhibitory concentrations against 12 microorganisms (Moir et al)

Amyloid beta’s potent antimicrobial activity was first characterized by Moir and team at Massachusetts General Hospital. In 2009 they reported that, in the laboratory, amyloid beta inhibited the growth of eight important pathogens screened by the study (see chart to the right). These included the bacterium S. pneumoniae: the leading cause of bacterial meningitis.

The pathogen that Moir and team identified as being most sensitive to amyloid beta is the fungus Candida albicans. Indeed, Moir and team found that the antimicrobial activity of amyloid beta was so strong that in some cases, its activity exceeded that of LL-37 – one of the body’s most potent and broad-spectrum antimicrobial peptides.

Infection-induced amyloid beta deposits co-localize with invading S. Typhimurium cells in 5XFAD mouse brain. (Moir et al)

Infection-induced amyloid beta deposits co-localize with invading S. Typhimurium cells in 5XFAD mouse brain. (Moir et al)

In a second 2016 study, Moir and team showed that amyloid beta protects against fungal and bacterial infections in mouse, nematode, and cell culture models of Alzheimer’s disease. In fact, when mouse brains were infected with the bacteria Salmonella Typhimurium, amyloid beta formed rapidly in response to the infection (with amyloid beta deposits closely associated with invading bacteria)

The team concluded:

..our finding that amyloid beta is an antimicrobial peptide is the first evidence that the species responsible for amyloidosis (accumulation of amyloid beta) may have a normal function. This stands in stark contrast to current models, which assume amyloid beta deposition to be an accidental process resulting from the abnormal behavior of an incidental product of catabolism (breakdown). Our data suggest increased amyloid beta generation, and resulting Alzheimer’s pathology, may be a mediated response of the innate immune system to a perceived infection.

PrP protein

Around same time that Moir and team characterized amyloid beta’s antimicrobial activity, a Swedish team at Lund University reported that PrP protein is also an antimicrobial peptide. The team found that synthetic PrP peptides killed the bacterial species Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. PrP also displayed potent antimicrobial activity against the fungus Candida parapsilosis.

Expression of PrP as an antimicrobial peptide in wounded skin (Schmidtchen et al)

Expression of PrP as an antimicrobial peptide in wounded skin (Schmidtchen et al)

The team further reported that PrP “breaks up” microbial membranes in a fashion similar to “classical” human antimicrobial peptide LL-37. They also showed that PrP is created as a response to wound healing in human skin. Indeed, in human skin cells, PrP protein induced production of TGF-α, a cytokine the body creates to target infectious agents.

Other independent studies support PrP’s role as an antimicrobial peptide. A team in Germany found that PrP expression was increased in patients with H pylori-infection, and not in non-infected controls. PrP levels decreased to normal after successful eradication of the H pylori bacteria. The team concluded that, “H pylori creates a milieu for enhanced propagation of prions in the gastrointestinal tract.”

Another study demonstrated PrP formation in skin cells removed from patients with psoriasis, contact dermatitis, squamous cell carcinomas, and viral warts (conditions tied to infection/skin microbiome imbalance). In these skins cells, cytokines associated with a response towards infection increased PrP production. A different team found that sheep with scrapie (an infectious condition) and lentiviral mastitis secrete prions into their milk (these prions were not created in sheep without the viral infections).

Alpha-SYN production is also promoted by toxic insult, local inflammation, and oxidative stress: conditions all associated with the presence of chronic infection.

Factors contributing alpha-SYN creation in Parkinson's (Melki et al0

Factors contributing alpha-SYN creation in Parkinson’s (Melki et al0

PrP’s ability to act as an antimicrobial peptide is particularly relevant in light of research connecting Parkinson’s Disease to prion creation. This October, the Journal of Neuroscience published two articles implicating alpha-SYN (a form of PrP) in Parkinson’s Disease. While this association between alpha-SYN and Parkinson’s has been reported for decades, the papers clarify the latest thinking on the topic. The authors of each paper debate details of how alpha-SYN accumulates in the Parkinson’s brain, but both conclude with a common belief: alpha-SYN proteins CAUSE or drive Parkinson’s disease progression.

Key to the above assumption is that both research teams do not appear to have read the Lund University study showing that PrP is an antimicrobial peptide. Or if they have, they didn’t cite the paper in their references, or refer to the findings anywhere in their writing.

The Journal of Neuroscience published two articles on prions in Parkinson's this October

The Journal of Neuroscience published two articles on prions in Parkinson’s this October

This is a major problem, because just like in Alzheimer’s, alpha-SYN is very likely not causing PD, but is instead accumulating in response to an as yet uncharacterized infection, or communities of infectious agents. In fact, the discovery that amyloid beta and PrP are antimicrobial peptides IS A GIANT CLUE suggesting that infectious agents in the brain are contributing to both disease states.

What infectious agents might these be? There are many possibilities. Researchers at the University of Alberta recently detected an ecosystem of chronic microbes in the autopsied brains of patients with HIV and a range of other medical conditions associated with severe neurological dysfunction. They detected 173 bacteria and bacteriophage-derived samples. Many of these microbes were identified inside macrophages, astrocytes, microglia, and other cells of the immune system.

Several microbes identified in the brains were pathogens associated with specific human diseases. They included Delfia acidovorans, a pathogen implicated in endocarditis, bacteremia, and urinary tract infections. Delfia acidovorans has even been identified as part of the bacterial community in the arterial wall of patients who have aortic aneurysms. Viruses (including various herpes viruses) were also identified in many of the brain samples.

Another study by researchers a the Universidad Autónoma de Madrid studied the brains of patients with Alzheimer’s. All eleven Alzheimer’s brains studied were infected with a range of fungal organisms.

These preliminary findings suggest that we should greatly prioritize new studies that search for microbes in brain tissue and the central nervous system. If more microbes are identified, the data may transform current models of Alzheimer’s, Parkinson’s and other neurological conditions.

Alzheimer's disease may cost $1 trillion dollars by 2050 (Image: Benny De Grove, Getty Images0

Alzheimer’s disease may cost $1 trillion dollars by 2050 (Image: Benny De Grove, Getty Images0

Progress in this area is stymied by the fact that most members of the scientific/medical communities are not aware of the findings discussed in this post (amyloid beta and PrP’s antimicrobial activity and/or the latest studies on microbes in the brain).

This may explain why, since 2002, 400 Alzheimer’s drug trials have been run and all 400 have failed. Many of these drugs have attempted to remove amyloid beta from the Alzheimer’s brain. But if amyloid beta is an antimicrobial peptide such treatments are removing a result of the disease process rather than the cause. No wonder they don’t work!

Meanwhile, Alzheimer’s disease is the sixth leading cause of death in the USA, with a new case diagnosed every 66 seconds. Without treatment, these numbers are predicted to explode to 16 million Americans with the disease, at a cost of over $1 trillion dollars by 2050. Then, factor in patients with Parkinson’s, schizophrenia, and other conditions tied to PrP and the impact of the discoveries described in this post is TREMENDOUS.

For example, Bill Gates just announced a new mission: He is investing $50 million of his own money into the Dementia Discovery Fund, a private-public research partnership focused on Alzheimer’s research. However, the research teams he is funding do not seem aware of Moir and team’s findings on amyloid beta and/or the paradigm shift in Alzheimer’s research that the discovery warrants.

We cannot pour this kind of money into research going in the wrong direction. There is too much suffering, too much debilitation, and far too great a cost to society.  Please, if this post speaks to you, share it with as many members of the medical community as possible.


Note: It’s possible that amyloid beta and PrP play a “dual role” in chronic disease. They may combat infection initially, but cause problems if their levels become very high. For example, Moir and team point out that LL-37 (the most well-studied human antimicrobial peptide) can become cytotoxic to host cells at very high concentrations.


A final consideration: could prions (PrP) play a role in ME/CFS?

In 2008 I watched prion researcher Adrianno Aguzzi speak at the “Days of Molecular Medicine Conference” in Karolinska Sweden. His team reported that activated B lymphocytes play a critical role in facilitating prion formation.

Norwegian researchers have reported that the immunosuppressive medication ritauximab can alleviate ME/CFS symptoms. Rituximab “knocks out” B lymphocytes. If prions play a protective role in ME/CFS, their inability to form in the presence of Ritauximab would decrease patients’ ability to fight infectious agents. This would result in decreased immunopathology (less of a battle between immune cells and microbes). Inflammation and symptoms would subsequently drop, although infectious agents driving the disease would be able to spread with greater ease.

Interview with evolutionary biologist Paul Ewald: infection and chronic disease

November 11th, 2017 by Amy Proal

Paul W. Ewald is an evolutionary biologist, specializing in the evolution of infectious disease. He received his Ph.D. from the University of Washington, in Zoology, with specialization in Ecology and Evolution. He is currently director of the program in Evolutionary Medicine at the Biology Department of the University of Louisville.

Below is an interview I conducted with Ewald in 2008. What he describes is as relevant today as the day we spoke:

Screen Shot 2017-11-11 at 11.31.26 AMHow do the concepts of evolutionary biology support the idea that pathogens are to blame for most diseases?

When we consider the possible causes of disease, it’s important to make sure that at our starting point, we put all categories on the table. I believe the most useful way to do this is to think in terms of three main categories:

  • inherited genes
  • parasitic agents (this includes bacteria, viruses, fungi, protozoa)
  • non-living environmental factors (too much or too little of a particular substance, radiation, exposure to a chemical etc.)

Once we have this spectrum of categories in mind we ask, “Have all three areas been investigated?

At this point scientists tend to make an error. They decide that if they have found enough evidence for categories 1 or 3, that category 2 is not playing a role. This is a fundamental problem that has led the medical community to misunderstand the cause of most debilitating chronic diseases.

So, which of the three categories is overlooked? Category 1 certainly isn’t – once scientists figured out the structure of DNA and the nature of mutations they were extremely eager to show their relationship to disease. Category 3 hasn’t been overlooked, largely because of the fact that we can sense environmental causes of disease. We suffer from a stomach ache after eating contaminated food or feel the pain from a sunburn.

But, if we look back at every decade, there has been a lack of research on category 2 relative to its actual importance in causing disease. Our track record shows that we have consistently failed to fully understand the role that pathogens play in causing disease and this trend has continued up until 2008.

There are many examples of how we have continually overlooked the category of infectious disease. I’m not talking about acute infection – researchers were essentially able to work out the mechanisms of acute infection from 1880 to about 1920. I’m talking about chronic infection, and thus the role of pathogens in causing chronic disease.

Take the case of peptic ulcers. The idea that bacteria cause peptic ulcers was first solidified in the 1940s, then independently investigated and solidified again by Marshall and Warren in the 1970s. It took over 20 more years before the relationship was finally accepted by mainstream medicine. Now, when people look back on previous theories about the cause of peptic ulcers they think, “Oh, isn’t it surprising that we didn’t understand the cause for so long!” or “We should have known better!” But when they proceed to hypothesize about the cause of other diseases, they go right back to the dogma. They haven’t learned the lessons from the peptic ulcer story.

Instead, they should think in an opposite fashion. If we find that one disease has an infectious cause, we should learn from that information and seriously consider the same possibility in other diseases.

Think about syphilis. In 1913, it was discovered that the disease resulted from infection with the bacterium Treponema pallidum. Soon, the disease was dubbed the “Great Imitator” because its symptoms often resembled those of other diseases, particularly in the later stages. I think syphilis should be called the “Great Illustrator,” because it’s a disease that imitates a whole spectrum of other diseases. This suggests that we should be actively looking for a pathogenic cause in these other diseases as well – especially since so many illnesses are still considered to be of unknown cause. Back in the day, the psychoses associated with syphilis and schizophrenia were grouped together into a single category of illness. But as soon as syphilis was found to have a bacterial cause, we separated syphilitic insanity from what is now called schizophrenia, and assumed that schizophrenia was not caused by infection. Rather than just separating the two diseases we should have actively pursued the hypothesis that schizophrenia also has an infectious cause. The information we can gain from these kinds of relationships is far more enlightening than any genetic data.

That’s one of the realities of medicine – researchers tend to deny associations. Denial plays a major role as scientists love to hold on to the current dogmatic explanation. This suggests that in order for pathogens to be fully tied to chronic disease we will have to wait until the current powerful people pass away and a sufficient number of young people entering the arena without these vested interests mature into positions of influence, to tip the balance of expert opinion. This is something that Charles Darwin, Max Planck, and Thomas Kuhn all agreed with.

That’s because powerful people tend to hang on to the opinions that made them powerful even if there is no longer sufficient evidence to support their views. It’s a social problem that relates to the weakness of the mind. Human beings didn’t evolve to be scientists. Instead they evolved to be competitive – to grab and hold onto what is theirs. Hence the name calling often observed among the medical community and the resistance among scientists to fund or support ideas other than their own, ideas that question the validity of current dogma.

From an evolutionary perspective is it possible that current diseases of unknown cause could all be genetic diseases?

No. Take schizophrenia again. Evolutionary biologists understand that if an allele (a sequence that codes for a gene) were to code for a disease it would slowly get weeded out of the population, particularly since people who are sick are much less likely to reproduce (especially people with a severe disease like schizophrenia). Yet a person’s chances of getting schizophrenia are 1 in 100. The reality is that faulty genes cannot maintain this frequency. If schizophrenia was a genetic disease, then according to the rules of mathematics, it would only occur in about 1 in every 10,000 people. The current frequency of the disease is just far too high.

Some might try to rationalize the 1 in 100 number by saying that schizophrenia is influenced by environmental factors, but if this were the case the environmental factors would have to be widespread and consistent across much of the world which is highly unlikely. Yes, some populations do have a higher incidence of schizophrenia than others, but that variability is much better explained by the idea that some populations harbor more of the pathogens that cause schizophrenia then others.

This highlights another issue. The fact that illnesses tend to run in families does not mean that only faulty genes are at work. Family members could just be passing each other pathogens. If one member of a twin pair has schizophrenia, there is a 35-60% chance that the other member of the twin pair will have it. However, this may be just a reflection of the fact that both twins were exposed to the same pathogens in the womb.

Pathogens also possess the ability to evolve and adapt at rapid rates, meaning that even if the host acquires a defense against them they can often find away around it. As previously mentioned, genetic disease would gradually be weeded out of the population. But as soon as you hypothesize that a disease has an infectious origin, and that the pathogens causing it can adapt and evolve, it is possible to explain how diseases can be perpetuated indefinitely in quite severe forms.

So you support the idea that the genetic mutations picked up on by many scientists may be induced by pathogens?

Yes, it’s possible. We know that some viruses and bacteria mutate and damage DNA. Similarly, the compounds created by the body in order to continually combat pathogens are often potent molecules that can also cause genetic mutations.

Give me some examples of diseases in which an infectious agent is certainly to blame.

Cancer is really a special case of the problems we have discussed. The same dogma has been driving how the disease is viewed for so long. But if people are able to recognize the dogma for what it is, they can take a better look at definitive evidence about the disease. Taking a look at the track record of cancer researchers is a good way to decide whether the consensus view is right or wrong.

Back in 1975, mainstream medicine agreed that about 0.1% of human cancer cases were caused by pathogens. When it came to the rest of cases, their view was that they were probably caused by a combination of inherited predispositions and mutagens. Then in 1985, the percentage of cancer cases they tied to pathogens was 3%, and they continued to make the same argument about the remaining cases. In 1995 the percent of pathogen-induced cancer cases was accepted to be around 10%. Now, we’re at 20%. Still, mainstream medicine contends that the other 80% of cases do not have an infectious cause, but the questions is – do you believe them anymore? In this sense, the clarity of hindsight can help a lot. Between evolutionary instinct and plain common sense we can view the issues of pathogens and cancer much more effectively.

Or, take a disease like atherosclerosis in which noting patterns of infection is unavoidable. There are bacteria in the lesions of people with the disease and all kinds of inflammatory markers. What we need to do is take a step back, divorce ourselves from our predispositions, and look at these ideas together.

Modern medicine has done a poor job looking for clues of continued infection. This may be partly explained by the fact that in many cases, it’s hard to link a pathogen to a disease because the pathogen grows and spreads so gradually. So the time at which a person becomes symptomatic may be years after the onset of infection.

Recognizing these patterns requires thinking broadly and deeply, but medical professionals and researchers have been trained to think narrowly. They’ve tried to follow a model that resembles a NASA undertaking for a great moon mission in which every person brings his or her own particular specialty to the table. But that model doesn’t work for medicine. Instead medical professionals need to work together with a unified theory in mind. But at the moment, they don’t have a unified theory, and without a conceptual model to guide them, researchers are only able to determine risk factors for disease rather than come to an understanding of the overall cause.

Evolutionary biology is the most synthetic area of biology. It asks why things are the way they are, and integrates knowledge of how things work mechanistically. Evolutionary biology promises to be the most synthetic area of medicine for the same reason.

While we’re on the subject of cancer, it and heart disease are now considered to be inflammatory diseases. Wouldn’t the presence of inflammation be a red flag that pathogens are to blame?

Yes. And the immediate questions researchers should be asking is “What causes inflammation?” One thing that we clearly know causes inflammation is the presence of an infection. So, as soon as I hear the word inflammation I think, “What infectious agents are at play?”

That brings us to the concept of autoimmune disease – the idea that the immune system just “goes crazy.” I think the fact that the concept of autoimmunity was developed in the first place is largely related to the fact that our brains have not evolved to think scientifically. People who have studied disease from their own point of view have recognized that the immune system is extremely important. But as we’ve learned more about the immune system, we’ve realized that it is an extremely complicated system – as complicated as the brain. Just like we can’t look at one type of neuron and infer information about the entire brain, we can’t try to understand the characteristics of only some immune cells and think we understand immune function.

So, over the years, as researchers have been daunted by the complexity of the immune system, it has seemed logical that such a complex entity has the potential to go wrong. Because they are limited by the power of their brains, they tend to simplify the issue and view the immune system in the same way they would view a truck that could break down. There are two problems with this type of thinking. For starters, we can’t trust our intuition that something complex is likely to malfunction. The fact is, the immune system functions just fine in a large proportion of the population. The only logical way to explain the immune activation seen in patients with “autoimmune disease” is to suggest that there is some sort of agent pushing the immune system off balance. This argument is only strengthened by the fact that the same evolutionary forces that would cause a serious disease to be weeded from the population would also cause those people whose immune systems are prone to self-destruction to be eliminated from the population.

The concept of autoimmune disease has progressed to the point that now even researchers who previously dismissed the possibility of infection are accepting the possibility that “autoimmune” disease could be triggered by infection. This is some progress, but it’s not enough. Especially since the concept of autoimmunity encourages doctors to prescribe immunosuppressive steroids to patients. But if persistent infection is involved these steroids may exacerbate the fire by allowing pathogens to spread.

Do you believe that pathogens could be involved in the aging process?

Aging is a super-category. We’ve gradually lumped together more and more symptoms under the category of natural aging. Many of these symptoms are the same as those caused by diseases that surely have an infectious cause. In that sense, you could view much of what we now call aging as an incapacitating illness that leads to a decrease in function. We know that inflammation and the interaction of the immune system with pathogens can destroy tissue. So it’s not surprising that the tissues of a person who harbors a lot of pathogens would age earlier and alter their biological structure earlier in life. I do believe it is inevitable that people will eventually die of old age, but I suspect that this should generally happen when they are 80-100 years old. But we are increasingly seeing signs of aging-related diseases in people who are much younger.

What does evolutionary biology have to say about psychosomatic illness?

Personally, I believe that we label an illness as psychosomatic when we don’t really know what’s going on with the patient. It’s a last resort diagnosis – a black box. If we knew more about what was causing their symptoms we could address the issue more clearly.

Looking at psychosomatic illness from an evolutionary viewpoint, you could say that those people who might exaggerate how sick they feel in order to gain attention and resources could have an evolutionary advantage. But if that’s the truth, it only accounts for an extremely small percentage of cases. It’s also true that often an illness will have both a psychological and physical component. But just because a psychological component is identified doesn’t mean the physical component should be overlooked. Plus, most mental illnesses are probably the result of infection too. Chronic Fatigue Syndrome is a good example of a disease that up until recently has been dismissed as psychosomatic just because researchers couldn’t figure out the cause. On the contrary, it’s quite a serious illness.

What role do you feel the Internet will play in facilitating acceptance of an understanding of pathogens in disease?

I think the Internet plays an incredibly beneficial role as it can provide information to anybody who is willing to put in the time to learn terminology and information presented in the literature available on the Internet. I believe it will, and already is, changing the patient/doctor relationship and also the relationship of the general public with the government mainly because we can now check up on things and check up on them quickly. I can find information in half an hour rather than spending an entire day at the library – and think about the fact that this is happening all around the country.

Of course, now there is so much information on the Internet that it’s too much for an individual mind to keep up with. Sometimes you have to read quite a bit of literature in order to extract the relevant information. That’s why we need people to team up and share information. What we need is small groups of people poring over information together. In this way they can develop a more thoughtful, broad outlook. This is in contrast to the current medical model in which doctors and researchers are trained to specialize in such a narrow area of knowledge. They know very little about issues outside their area of expertise and have trouble seeing the big picture. Thus, what we need is for the NIH to put money into grants that foster interdisciplinary insights.

Do you think that the peer review system and pharmaceutical industry are standing in the way of understanding chronic infectious disease?

I think the peer review system is becoming less important because there are so many other outlets where people can put up information. So I don’t see it as too big of a barrier.

When it comes to pharmaceutical companies, it’s important to recognize that they are very good at some things and very bad at others. What they are good at is product promotion and marketing, and working in innovative ways when the resulting product can bring in lots of money. The problem is the products that make the most money are not necessarily the products that actually help people the most.

Basic economic principles first put forth by Adam Smith show that the free enterprise system does not work well under certain situations. Writing over two hundred years ago, he argued that free enterprise cannot be expected to generate an effective national defense. For modern society, pollution control would be another example. If we want to move in that direction there needs to be a profit driven motive, or we have to get the government to do the things that do not generate sufficient profit. Otherwise, it just won’t happen.

If you think about it, there isn’t very much money to be made off a vaccine because a person uses it once or twice in his or her life and that’s it. Instead, think of the amount of money to be made off a statin when a person is going to take it every day of their life. There’s just not much motive for drug companies to invest in products that are cures or very good preventatives. We don’t have to condemn drug companies, just recognize this role that they are playing in drug development. If we want to develop a drug under a high priority situation that may result in a curative solution we can’t count on the pharmaceutical industry. In cases where the free enterprise system doesn’t result in a situation that may benefit the population the government has to step in and provide funding for the possibility at hand.

What kind of approach to research would best expedite the process of better understanding the role of infectious agents in disease?

Most experts in the health sciences advocate a building-block approach to the problem of causation. They try to understand the workings of disease at the cellular and biochemical levels, in hopes that solutions will eventually emerge. Even among infectious diseases, however, the fundamental achievements have occurred more through the testing of deductive leaps than by building-block induction. What it boils down to is that we need both types of development but we can’t have one without the other. Working incrementally can be great as long as scientists understand the big issues and the larger concepts that need to be guiding their research. But right now we have way too many scientists working in building block mode, missing what’s going on outside the box.

Why do doctors often have such a problem accepting the idea that pathogens are to blame for more diseases then commonly accepted?

Because it’s not in the textbooks. They are trained to look at a patient and try to match them with something in a textbook. These medical texts don’t consider the preponderence of evidence across the entire spectrum of possible causes of most chronic diseases. The evidence implicating infectious causation tends to be a casualty of this restricted perspective, leading to the result that consideration of infectious causation in medical texts is minimal for chronic diseases of uncertain cause.

So until infectious pathogenesis is accepted to the point that it is in the medical texts taught in medical school, they will continue to consult only the standard operating procedure. If they can’t put a label on the patient’s illness it falls into a bin of “unexplained phenomena” which goes back to what I was saying earlier about psychosomatic illness, since they have a tendency to speak dismissively about what’s in the bin.

The problem is that considering a new pathogenesis or a cause that isn’t in a textbook requires thinking hard about unknown problems. They just don’t have the time or training to think logically and deeply about such issues. This is evidenced by the fact that you can go to five different doctors, get five different explanations for your problem and be recommended five different treatment options.

What got you interested in this area of research?

My interest in evolutionary medicine began in grad school around 1977. I came down with a bad case of diarrhea and was thinking about whether I should treat the symptoms or let the illness run its course. At first it seemed like it was most logical to let the illness run its course because that seemed to be my body’s way of eliminating the pathogen. But then I thought about the fact that the pathogens might be manipulating me. If I expelled them, they might be endowed with an evolutionary advantage that would allow them to persist and infect others. I realized that my intuition couldn’t provide me with the answer. That led me on a long web-like series of connections. The more I started to consider medical problems in the light of evolution the more I realized that some diseases simply cannot be caused in a way they are explained by current dogma. So I’ve tried to look at disease in a balanced way, to put all possibilities on the table, and from there to figure out what’s feasible and what’s not.