Emergence and Evolution of Avian H5N1

This episode discusses the emergence and evolution of avian H5N1.

Our episode guest is Kevin Legge, Pathology Endowed Professor of Pulmonary Immunology Research, Vice Chair for Research in the Department of Pathology, Professor of Microbiology and Immunology, Director, Department of Pathology Research Flow Cytometry Facility, Director, Interdisciplinary Program in Immunology at the University of Iowa, and Associate Director at the Nanovaccine Institute.

This episode was recorded on September 30, 2024. Respiratory disease is an ever-changing topic in research and the status of infections across the US and around the world. Please see the Center for Disease Control and the US Department of Agriculture for the most up-to-date information on the status of infections and recommendations.

The information in this podcast is for educational purposes only and does not constitute medical advice or diagnosis.

The Nanovaccine Institute was founded at Iowa State University building upon core partnerships with the University of Iowa and the University of Nebraska Medical Center. Our purpose is to use nano-based technologies to tackle emergent diseases that have a devastating impact on human and animal health.

Iowa State University does not discriminate on the basis of race, color, age, ethnicity, religion, national origin, pregnancy, sexual orientation, gender identity, genetic information, sex, marital status, disability, or status as a U.S. Veteran. Inquiries regarding non-discrimination policies may be directed to Office of Equal Opportunity, 3410 Beardshear Hall, 515 Morrill Road, Ames, Iowa 50011, Tel. 515-294-7612, Hotline 515-294-1222, email eooffice@iastate.edu.

Transcript

Balaji Narasimhan (00:06.904)

Welcome to the Nanovax Podcast, a Nanovaccine Institute curated podcast sharing the latest research and advancements in Nanovaccines and nanotherapeutics for human and animal health. The Nanovaccine Institute is a consortium of researchers, institutes, national laboratories, and companies founded and coordinated by Iowa State University. We are developing nanovaccines and nanotherapeutics for respiratory infections, neural disorders, antimicrobial resistance, cancer, aging, and animal health. Our vision is that nano-vaccines and nanotherapeutics will revolutionize how we prevent and treat disease. Hello, I'm Kathleen Ross. I am Balaji Narasimhan. I'm Mike Roof. This season, you will hear from experts across a range of disciplines who study respiratory diseases and antimicrobial resistance.

The first half of the season will focus on respiratory disease and antimicrobial research within the Nanovaccine Institute. The Nanovaccine Institute is pioneering the development of novel, nano-based countermeasures, such as a universal influenza vaccine. We will also talk about other emergent and impactful diseases that have radically changed how we consider and develop policy around global public health. The second half will focus on the challenges and countermeasures to antimicrobial resistance.

Nanovaccine Institute researchers have developed novel nanomedicine therapies to treat chronic disease caused by bacterial pathogens that have developed antimicrobial resistance. Episodes will explore the different corners of these research topics through presentations on the latest findings as well as conversations among the experts.

Balaji Narasimhan (01:59.086)

Hello everyone and welcome to this episode. I am Balaji Narasimhan, Director of the Nanovaccine Institute and Anson Marston Distinguished Professor and Vlasta Klima Balloon Faculty Chair in the Department of Chemical and Biological Engineering at Iowa State University. In this episode, we will be talking about the emergence and evolution of avian influenza. Since 2003, avian influenza viruses, including H5N1, have circulated in 23 countries primarily affecting wild birds and poultry, resulting in nearly 900 reported human cases, primarily among people who have had close contact with infected birds and animals. Very recently, a highly pathogenic avian influenza virus called HPAI H5N1 has spread to infect more than 50 animal species. As of June 2024, so just a couple of months ago, the virus has not demonstrated any genetic evidence of the ability to easily spread from person to person. But public health officials are closely monitoring this as part of overreaching pandemic preparedness efforts. Here to talk with us about this today is an expert in this field, Dr. Kevin Legge. Dr. Legge is a professor of pathology, vice chair for research, professor of microbiology and immunology, director of the department of pathology research flow cytometry facility, director of the Immunobiology Graduate Program, Interdisciplinary Program in Immunology, and associate director of the Nanovaccine Institute. Kevin's primary faculty appointment is at the University of Iowa, and his lab focuses on dendritic cell immunobiology and pulmonary immunity. He has been studying immunity to influenza virus for 25 years, and so he's an expert in this area, and we're very happy to have him here. Kevin also has an active project on avian influenza that we collaborate with him on. And this project is focused on developing a nanoparticle-based vaccine against avian influenza. This research is currently in progress and supported by the National Institute of Health, specifically the National Institute of Allergy and Infectious Disease. So before we dig into the research, perhaps the best place to start is an overview of avian influenza. So this season, we have heard about livestock management regarding avian influenza.

Balaji Narasimhan (04:23.318)

for logistics to mitigate spread and economic dimensions of the disease. But let's maybe begin with a conversation about the epidemiology of the disease. So Kevin, welcome. And perhaps we could start with maybe you giving a brief overview on sort of what's the source of avian influenza, particularly H5N1. What's some of the history behind this virus? Yeah, great. I'm glad to be here. So I guess let me start with an understanding of how we have flu viruses in general and how they move through the community. We really deal with two things with flu. One is what we call antigenic drift. This is the slow movement of the virus through mutations that occur every year. This is why we get vaccinated every year. That's why your immunity to this year's virus may not be protected against next year virus. And this is small amino acids changes within the virus that escape our neutralizing antibody responses.

But flu is a negative sense segmented virus, RNA virus. And so it has eight segments. And if two flu viruses were to infect one cell, new viruses can appear. So segments one, three, and five could come from virus A, and the remaining segments come from virus B to create a brand new virus. We call this antigenic shift. And around 1997, what we noticed, at least in poultry, as well as some migratory birds, was a brand new flu virus, an H5N1, what we'll call high path avian flu. And we'll talk about what that is in a minute. But really it acquired one segment from a goose virus. Seven of the other segments came from either duck or quail viruses. And we had one of these antigenic shift events creating this brand new avian H5N1 virus. So how is this different from seasonal influenza?

Yeah, so seasonal influenza are low path strains of even H5N1 behave more like what we think of in a seasonal way, right? So this virus has a, its entry protein is called HA or hemagglutinin. It binds to sialic acids on the surface of cells to inner cells. For most of the time, what happens is that HA has to be cleaved

Balaji Narasimhan (06:46.624)

by enzymes that are found in our respiratory tract. This is why this is a virus that's respiratory tropic. You don't see it, say, in the intestines, or you don't see it in the kidney, you don't see it outside of really the respiratory tract. High path avian influenza virus has a polybasic amino acid residue that has occurred naturally in the HA that allows it to be cleaved by enzymes throughout the body. So you see systemic movement of this virus throughout the body, as opposed to just local immunity.

or local tropism because of the cleavage only within the respiratory tract. There's other some other unique aspects of that. And so we know that this polybasic amino acid residues allow systemic movement of the virus, but avian viruses in general tend to use a different sialic acid than human viruses do. And in particular, they like to bind to alpha two three link sialic acid residues where human viruses like to bind to alpha two six link sialic acid residues. And we know over time there are mutations that concur in these avian viruses that will become more human adapted, if you will, to bind the human respiratory tract. In fact, we have both alpha 2, 3, and 2, linkage sialic acids in our respiratory trees. But the avian binding sialic acids are really lower in the airways. And so it's thought that maybe you need 10 times as much of the avian viruses to actually infects a human than what you would need with, I say, a seasonal human-based influenza virus that likes that sort of tropism. And then there's another unique aspect of that. At that HA's cleave, we have fusion of the virus with the membrane of the cell to allow infection. And that occurs in different pHs for avian viruses versus human viruses. So there's a really big burden that has to be overcome for an avian virus to really replicate well in humans. Changes in the HA cleavage sites, changes in what it can bind to, and then changes in the pH susceptibility of the virus. So if the virus is not going to behave well with the pH, it would be degraded by acids. But if it changes to something more human-like, it would be able to survive that and you would actually get infections. All right. Let's talk a little bit about the disease in terms of how it's manifested. So what are the symptoms of avian influenza? is there any data that's available about

Balaji Narasimhan (09:13.516)

age distributions and exposures? Yeah, so the symptoms of flu, avian flu or seasonal flu are roughly the same. What we've seen at least early on was that there seemed to be a higher incidence of folks that actually got infected having more severe outcomes. And if you actually look at the data from early cases from say 2003 to 2013, there were roughly 868 cases in humans that were associated with about 457 deaths. So about 53 % mortality rate, which was very concerning to the community. Now, what we never knew was what the true denominator of that was, right? If there were asymptomatic people that had been infected, it didn't have symptoms, didn't end up in the hospital, we wouldn't really know what the denominator is. And he put it in context, a seasonal influenza infection has a mortality rate of about .2%.

And so 53 % was concerning, but again, we thought like this was probably not true because the denominator was much, much greater than what we knew. And so bringing that back down to more levels and what we've seen with recent outbreaks here in human cases have been relatively mild disease conjunctivitis, those types of diseases. We haven't seen the lethalities that we've seen in these recent cases. ACE distribution is a little bit tough again, from 2003 to 2013, when it was really, really well studied.

It was individuals that were being exposed to infected animals. And in fact, the age distribution of that was young children. And it turns out those young children were the ones that were actually the ones tending the animals. And so the question was always, were young children more susceptible or was it because they were the ones being exposed to the animals? And I think the recent cases have suggested more that in fact, it's exposure to those animals as opposed to maybe age distribution, but we still don't know. 

53 % mortality would be absolutely catastrophic, obviously. So let's hope that we never get to a virus that has that kind of potential. So what evidence is there, Kevin, of probable person-to-person transmission? Yeah, I'd say the best case that's been out there is something in 2006. So the WHO reported a case of what was probably human-to-human transmission.

Balaji Narasimhan (11:35.02)

in Indonesia. There were eight individuals in one family that became infected. First one was somebody that was exposed to infected animals. Then six other members of the family got infected. And then one of them, a young child, we believe infected the father. And based on the serial natures of the infection, as well as the sequencing, it suggests that there was, again, human to human transmission. But once again, we think that it takes 10 times as much virus really to infect human to human transmission. And we've really not seen that occur since.

These individuals, it was reported that they seem to have higher viral loads than other individuals in the throats and the nasal passages, which may have resulted in this. But again, since 2006, there's not been real good evidence for human to human transmission. So given what we saw, you know, during the COVID-19 pandemic and how quickly viruses like these have the potential to have global impact what is your perspective on sort of the global significance of an avian influenza type virus? Well, I think there's both an economic but also a personal health impact as well. So just to put some economics into context, in 2022, we had a really strong avian influenza outbreak here in the United States that resulted in something like 40 million animals that were lost to the outbreak and an economic burden rating between around 2.5 to $3 billion. So obviously that has impacts on food safety, right? And downstream economic effects. As far as human disease right now, the cases have been extremely mild. Obviously we know that there needs to be some sort of mutations in this virus to allow widespread human to human transmission. That will probably come with some dampening down, typically of symptoms.

and lethalities, that's what we've seen in the past. And right now those cases have been really mild here in recent times. So obviously spreading of a brand new virus in a immunologically naive population or population that doesn't have immunity to that could be devastating. But right now all evidence suggests that we have some limited immunity to the virus as well as we have countermeasures that we'll be able to fight back against it. You mentioned 40 million birds. I suspect that

Balaji Narasimhan (13:55.84)

that would also have a significant maybe downstream supply chain impact on seasonal influenza vaccines, many of which are egg-based, right? So if we had an outbreak like this, that would also presumably have an impact on our seasonal influenza vaccine supply, right? It very well could for those egg-based produced vaccines. Obviously we have cell culture-based vaccine production.

as well now and other technologies that are coming down the pipeline. But certainly we grow our virus, our vaccine viruses right now that make the vaccine and in your native chicken eggs. And so if you lose those eggs, then they would have a problem. And in fact, that was one of the original issues in making a high path avian influenza vaccine, which we do have and have stockpiled was that poly basic amino acid residue that causes systemic release was killing those eggs. And so if you kill the eggs, you can't make the virus that you need for the vaccine. And so they've been able to remove that to create this stockpile vaccine, which has been shown to be pretty effective with antibodies reacting against the currently circulating strains. Right. So what are what are the currently available prevention strategies? What what can people be assured of with respect to what's out there? Are there vaccines? Are there other strategies that are in place to to help us deal with this? Yeah, think, yes, we have currently stockpiled vaccines that we know and they were tested in the early 2000s against H5N1 and high path avian influenza. And looking at those stockpiled vaccines, are individuals that got those vaccines during the trial. They recently revisited those individuals and looked and they have antibodies that would cross protect against the currently circulating strain. So we have vaccines ready to go.

The other really good thing is that we have antiviral drugs against influenza. And really there are three types of antiviral drugs that we think of with flu. The ones that inhibit the neuromidase, or release of the virus from one cell to another. We have one that interferes with this really interesting, if you will, cap snatching ability that the virus use where it takes our own mRNA and uses it for its own good. And then a third is this ion channel blockers.

Balaji Narasimhan (16:18.38)

And two of those three we know still work against the avian influenza. So the neuromenedidase blockers, things you may have heard of like Oseltamabir or Tamiflu. The RNA cap snatching, which is Oxfluza, is also very effective as well against this virus. So we have antivirals ready to go. We have vaccines that have been stockpiled and ready to go. And of course, there are a lot of folks now trying to get better vaccines ready to go in case we need them. 

What about animal control? Yeah, so animal control is another way. It has been a way of trying to cull and stop the spread. I certainly have seen that around the world early on in these outbreaks, where they would cull off poultry. They would cull off whatever wild birds, whatever they could needed to do to try to stop the spread. That is, of course, an economic burden, but also a food issue burden as well.

And in fact, here in the U.S., something like since 2016, there've been about 100 million birds that have been culled to try to stop the spread. And in fact, we've seen this around the world. We've seen this in Vietnam. We've seen this in Thailand. And in fact, the first evidence of this virus ever infecting cats, actually because they were fed infected poultry accidentally.

and some tigers at the zoo in Thailand actually came down with the virus. And so all this virus initially started out in migratory birds and, know, poultry, if you will. It's gone silent in migratory birds, but it's still circulating in migratory birds. And around the world, as it spread from where we first saw it, as it started to spread, what we would normally see is migratory birds that had died and then cats that were dying from eating the

the dead migratory birds and this would tell you that you probably had avian flu influenza in the area. What about the risk to urban chickens? The risk to urban chickens and as well as any chicken plots are really the same, right? So if they were exposed because of a migratory bird in the area that had the virus, they would be able to come down with it. And yes, backyard chickens have come down with that and have had to be culled.

Balaji Narasimhan (18:37.236)

So thinking about the impact about egg availability on vaccine production, it is very heartening to know that the food supply, the safety of our food supply is in place, intact, right? Because of the excellent quality control and quality assurance measures that are in place as we think about it from a food safety perspective for humans. Yeah, I mean, there's a lot of surveillance, right? So there's a lot of surveillance both in our food supply

both in farms, whether that be poultry, whether that be cows, and to stop the spread before it would ever get into the food supply. There's certainly surveillance in human populations as well to try to prevent the spread of the virus. But certainly in the food supply, we talked about culling of birds and to try to stop the spread, but that also means it doesn't get into our food supply. So that meat is not used if it's infected poultry, those eggs are not used, they stop before it ever gets to humans.

Certainly, I would advocate that you cook your poultry, that you cook your beef. And certainly, the CDC would certainly recommend right now that you don't drink unpasteurized milk. So that certainly would reduce any risk if something were to somehow slip through, which we don't think it's doing. So this is a nice segue into the research that's happening in your own lab. So can you talk a little bit about some of the newer types of, or maybe next generation,

vaccines that your lab is working on? Yeah, so Balaji, as you know, we've been working on a particle-based flu vaccine for a number of years now. if you think about the current seasonal flu vaccines, they're largely geared at driving one type of immune response. That's an antibody response. And that antibody response is geared to only one protein in the virus. That's that HA protein that I talked about earlier. When we get infected, though,

By natural infection, we mount a very diverse immune response. So we certainly mount antibodies against HA, but we mount antibodies against other flu proteins. And then we have another type of immunity called T cells. These T cells will find virally infected cells and kill those virally infected cells, thereby limiting the spread of the virus. And our current vaccines are not geared to drive T cell based immunity. And so we've thought for years if we could get T cell based immunity on top of antibody responses, it would confer greater levels of protection.

Balaji Narasimhan (20:56.844)

The other thing that the vaccines don't do currently is they don't drive mucosal-based protection. And so if you get a shot in the arm, you generate an antibody response and antibody circulates around, but you're really not parking cells within the lungs and nasal passages waiting to fight the virus when it shows up. So one of the ideas with this particle-based vaccination was to be needle-free. We'd be able to give it a nasal passages and drive local-based protection, both T and B cell or antibody-based protection, and give level

of higher levels of protection than what's currently seen with the current seasonal flu vaccines. And that's what we've been able to show in animal models. And as we started out with vaccines against seasonal flu, about a year and a half ago, as you know, we were funded to look at trying to drive a better vaccine against avian flu. At the time, we weren't having the major outbreaks in cattle and others that we've seen here, but it was certainly something that was on the horizon as a possibility. And so,

We've done, developed a vaccine against H5N1 and we've started to test that vaccine in preclinical animal models. That vaccine is showing very good effects to you driving both T cell and B cell, our antibody-based immune responses, systemic immunity as well as immunity within the nasal and respiratory tree, giving this that local as well as systemic based levels of protection. And interesting enough, we've now done some trials as to whether that vaccine would confer protection.

against high path avian flu, but also seasonal, if you will, H5N1 or low path flu. And the vaccine has performed very, very well. In fact, right now we've given what would be considered an excessive lethal dose in these animals of a challenge dose. And the vaccine has protected those animals from any type of disease, whether morbidity or mortality. Interestingly enough, we've also tested our H1N1 based vaccine or seasonal based vaccine. And that is also providing

protection against this high path avian strain. So this is just having this multifactored immune response, both T and antibody and B cell base, but also local and systemic types of protection again, for confer greater levels of protection than what seasonal vaccines may. That's very exciting, Kevin. So a related question along those lines is, while this type of broad based and immunity in both lungs as well as systemically,

Balaji Narasimhan (23:20.236)

is certainly exciting to see. What can we say about the ability of these novel sort of particle-based vaccines to potentially maybe prevent transmission of the disease? So how would a vaccinated subject be able to stop getting the disease if they're exposed to the virus? Yeah, so we don't have any evidence one way or the other yet, but certainly right now having mucosal-based protection

we suggest that you would have lower levels of transmission, right? If you stop the virus where it's replicating and where it leaves from, then you would be able to prevent transmission. Also, if you had protection, since we're inhaling the virus and you have local based protection, the idea that you would be better protected from transmission would be much higher. But right now we need to still formally test that to be able to know for sure. But certainly having systemic protection will give you longer term protection.

throughout the body, but also having local protection has a real possibility of providing prevention, if you will, from transmission. Great. So just to finish this off then, having learned about this exciting new approach to flu vaccines, both seasonal and avian, what do you see as the horizon for this going forward in terms of what it's going to take to get this vaccine through the approval process?

Well, I think several things. One, if we're talking specifically about the high path avian influenza based vaccine, there's more testing that needs to be done. If you're talking about the seasonal based vaccine or the particle based vaccination, certainly have tested in several preclinical models that have shown very good efficacy, but also safety. And I think the next step for that is obviously a phase one clinical trial and getting the approval from the FDA to be able to do that sort of trial. Obviously this next

goal for that would be able to move that into this H5N1-based vaccine technology as well. But given what we've seen in the preliminary data that with the H1N1 vaccine providing protection in these preclinical models, that might confer protection as well against these high-pass strains. Well, thank you very much, Kevin, for a very stimulating discussion. That's certainly gratifying to know that lots of exciting work is on the way, and hopefully we'll be able to even further.

Balaji Narasimhan (25:46.73)

enhance our preparedness for these types of viruses. So thank you everyone for joining us in this discussion. Other episodes this season will explore other facets of respiratory disease in livestock and humans. And later this season, antimicrobial resistance. Please check out the show notes to learn more about the research discussed in this episode. And please be sure to like and subscribe to the Nanovax podcast in your preferred app or media player.