August 1, 2025 • 19 mins
🎙️ Episode 93: Neuroinvasive and Pulmonary Virulence of H5N1 Clade 2.3.4.4b Genotype B3.13
🧬 In this episode of PaperCast Base by Base, we explore how a bovine-derived H5N1 virus from clade 2.3.4.4b genotype B3.13 exhibits enhanced pulmonary and neurovirulent properties in mouse models, highlighting its potential implications for zoonotic transmission and public health.
🔍 Study Highlights:
Using intranasal and orogastric inoculation of C57BL/6J and BALB/c mice with four clade 2.3.4.4b H5N1 isolates, the researchers assessed weight loss, survival, tissue viral loads, histopathology and cytokine responses
The bovine B3.13 isolate caused rapid uniform lethality at low doses and induced severe respiratory disease accompanied by neurologic signs in C57BL/6J mice
High viral titers were observed in lung and brain tissues, supporting systemic spread and neurotropism of the bovine isolate
Histological analyses revealed abundant viral antigen in respiratory and neural cells without overt brain lesions, accompanied by robust pro-inflammatory cytokine profiles in the central nervous system
Comparative analysis showed that the bovine genotype B3.13 strain possessed greater neuroinvasive and neurovirulent capacity than mink, mountain lion and reference human isolates
🧠 Conclusion:
These findings underscore the importance of monitoring emergent HPAI H5N1 genotypes in mammals and support the use of these mouse models for evaluating vaccines and therapeutics targeting neuroinvasive influenza strains
📖 Reference:
Tipih T, Mariappan V, Yinda KC, Meade-White K, Lewis M, Okumura A, McCarthy N, Altynova E, Leventhal SS, Bushmaker T, Clancy CS, de Wit E, Munster VJ, Feldmann H & Rosenke K. Highly pathogenic avian influenza H5N1 clade 2.3.4.4b genotype B3.13 is highly virulent for mice, rapidly causing acute pulmonary and neurologic disease. Nat Commun. 2025;16:5738. doi:10.1038/s41467-025-60407-y
📜 License:
This episode is based on an open-access article published under the Creative Commons Attribution 4.0 International License (CC BY 4.0) – https://creativecommons.org/licenses/by/4.0/
Mark as Played
Transcript
Episode Transcript
Available transcripts are automatically generated. Complete accuracy is not guaranteed.
(00:14):
Welcome to Base by Base, the paper cast that brings genomics
to you wherever you are. Imagine a scenario, one that
really keeps public health experts on edge.
A highly contagious animal virus.
It's already causing huge outbreaks in livestock, and then
suddenly it develops. This, this terrifying new
ability can attack the brain. What fundamentally shifts when a
(00:35):
virus, one we mostly know for inrespiratory illness, starts
showing clear signs of neurological impact?
How would that kind of change force us to completely rethink,
well, everything our understanding of the risks and,
crucially, our whole strategy for protection?
Well, this isn't just hypothetical anymore.
This is precisely the concern, the escalating concern around
(00:55):
the highly pathogenic avian influenza AH5N1 Clay 2.3 point
4.4 B viruses. These strains, they're already
fueling a significant ongoing outbreak in dairy cows across
the United States. And while the human cases we've
seen linked to H5 N 1, they've largely been mild so far.
The really unsettling question is, what if some of these
(01:17):
circulating strains actually possess traits that could
rapidly, dramatically change that picture, make human
infections far more severe? And that's exactly where today's
deep dive takes a critical turn.We're getting into some new,
really groundbreaking research that unpacks precisely this.
It reveals a specific H5 N 1 isolate originally from a cow,
(01:38):
actually with a surprising and frankly, deeply concerning
ability to cause severe, rapidlyfatal neurologic disease in a
key animal model. So we're not just talking
respiratory distress now. It's like a direct assault on
the brain. But before we delve deeper into
the, well, the alarming specifics of this research, it's
really essential we shine a spotlight on the team, the
dedicated team whose critical findings are shaping how we
(02:00):
understand this evolving. Who are the minds behind this
work? Yeah, absolutely.
We really celebrate the work of Thomas Tippevignesh, Mary Eppen,
Kaywe Sienda, Kimberly Mead White, Matthew Lewis at Sushi
Okamura, Natalie McCarthy, Ekaterina Elton, Nova Shenna S
Leventhal, Trenton Bushmaker, Chad S Clancy, Emmy DeWitt,
Vincent J Munster, Heinz Feldmanand Kyle Rosenke.
(02:20):
They're from the Laboratory of Virology, Division of Intramural
Research. NIAINIH up in Hamilton, Mt Their
work, it's significantly advanlyadvancing our understanding of
HPAIA H5, N1 virulence, and you know what it could all mean.
OK, so let's set. The stage a bit what?
Exactly Is HPAI AH5N1 and why has it become such an urgent
(02:41):
global concern like right now? Right.
Well, the highly pathogenic avian influenza AH5N1,
specifically what's known as thegoose Guangdong lineage.
That's basically a particular ancestral strain they found in
geese at first, popped up in China back in 1996.
Since then, this virus has just been a master of evolution.
It's constantly adapting, branching out into 10 main
phylogenetic clades. You can think of these clades as
(03:02):
sort of distinct genetic families, right?
Subgroups, each with slightly different traits and behaviors.
And clade 2 has really proven tobe the most dominant, the most
widespread. Starting around 2005, these HPA
IH5 N 1 Clade 2 viruses began tospread well pretty aggressively
across Asia, Europe, Africa, andthen eventually they made their
(03:23):
way to North and South America. And it's not just a threat to
birds anymore, is it? We've seen this worrying trend,
this spillover into mammals. Absolutely.
Since late 2021, these H5 N 1 clay 2.3 point 4.4 B viruses
have been continuously circulating in North America.
Huge outbreaks in wild birds, domestic poultry.
But what's really alarming is the increasing spillover, you
(03:45):
know, the virus jumping from birds into wild mammals.
We've documented cases in farmedmink in Spain that was 2022,
Arctic foxes, raccoon dogs in Finland, domestic cats in
Poland, even green mammals down across South America.
And just recently a Mountain Lion was found dead in Montana,
also infected with H5 N one. It just highlights how broad its
impact on mammals can be. And then things really escalated
(04:08):
with the dairy cows. That really caught.
Everyone's attention didn't. It almost became a household
name. It really did.
In March 2024, HPAIA H5 N 1 Clayed 2.3 point 4.4 B viruses
were found in dairy cows and crucially, in unpasteurized milk
across the US. Now, as of when this paper came
out, 70 human cases of HPA IH5 None had been reported in the
(04:31):
United States. These were caused by either the
B 3.13 genotype from the dairy cows or the D 1.1 genotype from
birds. And while most human cases have
been mild, often just conjunctivitis or, you know,
pink eye respiratory symptoms have definitely been described.
And in a pretty sobering case upin Canada, a teenage girl
actually needed hospitalization for severe respiratory failure.
(04:51):
Thankfully, she did recover. So the main risk for humans
getting infected here is mostly occupational right for people
working closely with these animals.
That's a big part of it. Yes, occupational exposure is
common, especially for folks on dairy and poultry farms.
But we've also seen severe humancases from exposure to sick and
dead birds in just backyard flocks.
(05:11):
So the exposure sources aren't strictly limited to large
commercial operations. And a really critical point here
is that there's clear evidence of mammal to mammal transmission
happening in some cases. Yet despite all this, the USCDC
currently categorizes the overall public health risk for
human infection as low. But the big So what?
Here, the really profound point is that this low risk could
(05:33):
change, and it could change veryquickly if the virus keeps
evolving, picking up new geneticchanges.
This particular study, it seems it dies right into that critical
gap in our understanding of the virus's virulence, its potential
punch. Precisely, it fills a really
crucial knowledge gap. OK, so with such a pressing
evolving threat, how did the researchers actually go about
investigating the virulence these specific H5 N 1 strains?
(05:55):
What was their their approach? Well, they started with a very
clear comparative goal. They wanted to assess the
virulence, basically how harmfulthe virus is of a well known
reference HPAIA H5 N 1 clade onevirus that's Vietnam 1203, 2004
or VN123. And they compared this benchmark
against three very recent HPII AH5N1 clade 2.3 point 4.4 B
(06:20):
isolates. These newer ones were sourced
from, you know, highly relevant situations, one from a cow, one
from a Mountain Lion and anotherfrom mink.
The bovine isolate, a bovine OHB24 O SU-342200 four that was
obviously of particular interestgiven the ongoing dairy cow
outbreaks. And they use mouse models for
this, right? Which specific strains did they
use and why are those models important here?
(06:41):
That's right, they use two established mouse strains,
C57BL6J and BALBC mice. These are pretty standard in
virology because we know a lot about how they respond to
influenza viruses. It makes them reliable for these
kinds of comparative studies andto mimic how exposure might
happen in the real world. They compared 2 main ways of
giving the virus intranasal, which is like inhaling virus
particles, and orgastric, where the virus goes in via a feeding
(07:03):
tube simulating eating or drinking something contaminated.
Right, that orgastric route seems particularly relevant for
how it might spread through contaminated milk, for example.
Exactly. It's key for understanding that
potential pathway. So once the mice were exposed,
what kind of data were they collecting?
What were they measuring to see the impact?
They tracked several key things really meticulously, daily
(07:24):
monitoring for clinical signs, so the visible symptoms of
sickness, weight loss, ruffled fur, hunched posture, rapid
breathing, that sort of thing. And critically, they watched
very closely for neurological signs, tremors, ataxia.
That's a loss of coordination, hyperactivity, circling
behaviors. They also tracked survival rates
for up to 28 days. And then to understand how the
(07:47):
virus was replicating and spreading inside the body, they
measured virus titers. That's the amount of virus
present in organs like the lung,liver and brain, and also in the
blood. They collected these at the peak
of the disease. So they weren't just watching
symptoms, they were digging deeper, looking at what was
happening at the cellular level systemically.
Exactly. They used histopathology, which
(08:07):
is like doing microscopic forensics on the tissues looking
for any cell damage or changes caused by the virus.
They also used immunohistochemistry.
That's a technique using specialmarkers to basically light up
exactly where the virus parts, its antigens are located within
the brain and lung cells. And to understand the immune
response. They did cytokine analysis in
brain and lung tissue. Cytokines are like the immune
(08:30):
system's alarm signals. They looked at specific pro
inflammatory ones like IL 1, GMCSF and MIP one which tell you
the body's mounting a strong, maybe even overwhelming
inflammatory response. And finally, specifically for
that bovine isolate, they did low dose studies to figure out
its median lethal dose, the LD 50.
That tells you how little virus it takes to be fatal to half the
(08:51):
animals, giving you a real senseof its potency.
Wow, quite comprehensive. So after all this meticulous
work, what did they find was thebig discovery?
This is where the H5 N one storyseems to take a really
concerning turn. Yeah, the most impactful result,
no question, revolves around that bovine isolate.
It showed dramatically enhanced virulence and really an
unprecedented ability to invade the brain, what we call
(09:14):
neuroinvasion. Compared to all the other
strands they tested in the C57BL6J mice, the bovine isolate
caused uniform lethality. Didn't matter if it was given
intranasally through the nose oror gastricly by ingestion.
Uniform lethality. Yep, intranasally mice died by
day four or gastricly by day 6. This disease progression was
(09:34):
significantly faster and more severe than what they saw with
the mink Mountain Lion or even that reference VN1 Toll 3
string. And those neurological signs you
mentioned, That sounds like the really striking and honestly
quite terrifying part. It absolutely is.
Crucially, every single C57BL6J mouse given the bovine isolate
developed clear, undeniable neurologic signs.
(09:55):
We're talking tremors, ataxia, that loss of coordination again,
hyperactivity circling. This level of neuroinvasion was
largely unique to the bovine isolate.
Only two of the Mountain Lion infected mice showed just minor
temporary hyperactivity. It was a really stark contrast
and matching up with these severe neurological signs.
The researchers found extremely high virus titers, huge amounts
(10:16):
of virus in the brains of those bovine infected C57BL6J mice,
way higher than any other isolate where the brain either
had no virus or very low levels.Plus the bovine isolate showed
enhanced replication and other organs too, and cause systemic
infection. They found significantly higher
levels of virus circulating in the blood.
That's viremia. It shows the virus wasn't just
(10:38):
staying put, it was spreading all through the body.
OK, so we've got high viral loads in the brain, unmistakable
neurological symptoms. But here's the really
groundbreaking part of the paradox you mentioned earlier.
What about the actual damage to the brain tissue itself?
That's where things got weird, right?
This was a genuinely perplexing finding.
Yeah. And it forces us to sort of
rethink how these viruses can cause trouble.
(11:00):
Despite those incredibly high viral loads and the clear
presence of viral antigen, the virus bits inside the neurons
and glial cells in the brain, when they looked at the brain
tissue using histopathology, they saw no significant physical
tissue damage or inflammation. No damage but clear symptoms and
virus present. Exactly.
It's not just a fascinating biological puzzle, it really
(11:21):
shifts our understanding of how viruses can cause neurological
disease. It suggests the neurological
signs might not be from direct cell destruction or a big
inflammatory attack, but maybe from more subtle neuronal
metabolic disturbances. Like imagine the virus subtly
messing with the brain cells energy systems, causing
dysfunction without obvious structural damage.
(11:44):
This invisible damage mechanism is a critical new piece of the
puzzle. That's wild.
It is, but it's important to note while they didn't see the
physical damage, the bovine isolate did trigger a
significant pro inflammatory cytokine response in the brain
tissue. Those elevated Illinois 1GMC S
FM IP110 levels. So the brain's immune system was
definitely reacting, even if thestructure looked OK.
(12:05):
And you mentioned differences between the mouse strains.
Did one turn out to be more useful for studying these
neurological effects? Yes.
Interestingly, while the Bao BC mice actually died a day earlier
from the bovine strain than the C57BL6J mice, it was only the
C57BL6J mice that consistently showed those obvious severe
(12:26):
neurologic signs. This happened even though both
strains had similar amounts of virus in their brains at later
stages. So it might be down to different
immune responses between the strains, or maybe subtle
differences in how the disease progresses overall.
Perhaps the BLLBC mice just succumbed faster to the
respiratory illness before the neurological signs really had a
chance to fully develop. Interesting.
(12:48):
So yeah, the C57BL6J model proved particularly valuable for
directly studying this neuro invasion and neurovirulence.
And let's just circle back to that orgastric route, ingesting
the virus. How significant was that finding
overall? It was very significant.
This study strongly reinforced its importance.
It confirmed it as a pertinent way for the bovine isolate to
cause infection, leading to rapid disease and death.
(13:10):
This has really direct implications for how this virus
might spread in the real world, you know, beyond just
respiratory transmission. OK, so let's try and pull this
all together. What does this all mean for us?
For the ongoing H5 N 1 situation?
For how we think about viral threats?
What are the big takeaways? Well, the central implication is
pretty stark, really. The increased virulence and the
(13:30):
neuroinvasion shown by this specific bovine H5 N 1 isolate
in these mouse models is franklydeeply concerning for both
animal and public health. It tells us this virus is
clearly evolving in ways that could lead to more severe
disease. It's a serious wake up call that
the virus isn't static, it's changing.
We saw the. Virus.
Got into the brain, but the exact mechanism, the how isn't
(13:53):
totally. Clear yet right?
That's right, The precise entry points aren't fully nailed down.
But the presence of viremia virus in the blood strongly
suggests hematogynous spread, like the virus gets into the
bloodstream and then has to somehow cross the blood brain
barrier. That protective shield around
the brain for viruses entering through the nose like in the
intranasal tests, direct spread along cranial nerves, the nerves
(14:15):
connecting the nose straight to the brain, seems like a very
plausible route. And what's really fascinating
for the orgastric route, when the virus is swallowed,
researchers are looking at thesenerve networks in the gut wall,
the submucosal and myenteric plexi.
These could potentially act as direct neural pathways from the
gut right up to the brain. That would be a pretty novel way
(14:35):
for a virus like this to cause neuroinvasion.
Definitely needs more investigation.
And that paradox you described, the neurological signs without
obvious brain lesions, What's the thinking there?
How does that change things? Yeah, the absence of those clear
histological lesions, despite the strong neurological signs in
the virus being present, it is really intriguing.
It strongly supports the idea that the symptoms might be due
(14:58):
to those neuronal metabolic disturbances, not direct cell
killing or massive inflammation.This has big implications for a
diagnosis and maybe even treatment.
If doctors are only looking for physical brain damage to confirm
neurological H5 N one they mightmiss cases.
It suggests we might need to focus more on functional
changes, maybe biochemical markers of neuronal stress.
(15:19):
It's an area needing a lot more research.
Now, we often hear about specific genetic mutations
making viruses nastier. Did this study find any clues
about what might be making this bovine isolate so aggressive and
neuroinvasive? It did, but with an interesting
twist. Some mutations that we've
previously linked to higher virulence and other H5N1
(15:39):
strains, things like the NA stock deletion, NS-1 deletion or
that PB2E627K mutation. Specific genetic tweets known to
boost disease severity, right? They were actually missing in
the recent isolate studied here,including the Bovine 1.
This suggests that other may be yet to be defined molecular
markers or perhaps combinations of genetic factors are driving
(16:01):
their enhanced virulence in mammals.
However, these recent isolates do have mutations in the matrix
protein M1 and NS-1 that are known to increase virulence in
mice. So while some expected flags
weren't there, other subtle changes are clearly playing a
role. It just highlights how complex
viral evolution is and why we need constant surveillance.
And let's hit that orgastric exposure route one more time,
(16:23):
because it's confirmation here seems incredibly relevant right
now. It is absolutely.
This study really hammers home its importance as a potent
exposure route. It gives us a very plausible
explanation for infections we'reseeing in wild and domestic
carnivores like that Mountain Lion, who probably get infected
by eating infected prey. But critically, it also
(16:43):
highlights a potential infectionroute for other mammals,
including humans, including cats, from drinking raw,
unpasteurized milk from infecteddairy cows.
That's a major point of concern this research underscores,
emphasizing the public health advice about avoiding
unpasteurized dairy. So these mouse models, even with
their slight differences, they're proving to be really
invaluable tools in this whole H5 N 1 situation, aren't they?
(17:05):
Oh immensely valuable the C57BL6J and BALBC mice are
excellent in vivo meaning in a living Organism screening
models, they're just crucial forquickly evaluating potential
antiviral drugs, testing different therapies and checking
how well new vaccine candidates work against these emerging H5 N
1 viruses. And specifically, like we
(17:26):
discuss the C57BL6J model is turning out to be particularly
useful for digging into neuroinvasion and the mechanisms
behind neurovirulence. They're giving us critical fast
insights that directly help our public health responses and
preparedness. So I think the central insight
from this deep dive, it's just unequivocally clear this highly
pathogenic avian influenza AH5 N1 bovine isolate, the one
(17:48):
actually circulating out there, shows significantly enhanced
virulence in these mouse models.It rapidly causes fatal
respiratory disease and severe neurologic disease.
This unique ability to invade the nervous system even without
causing obvious visible brain damage really underscores the
dynamic evolving nature of this virus.
It shows a concerning potential for unexpected and severe
impacts. So what does this powerful
finding really mean for how we respond to the ongoing H5 N 1
(18:10):
outbreak and how urgently shouldwe be preparing for potential,
maybe even drastic shifts and how this virus behaves,
especially thinking about transmission routes like raw
milk? This episode was based on an
Open Access article under the CCPY 4 point O license.
You can find a direct link to the paper and the license in our
episode description. If you enjoyed this analysis,
(18:31):
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Thanks for listening and join usnext time as we explore more
science base by base.
Welcome to Base by Base, the paper cast that brings genomics
to you wherever you are. Imagine a scenario, one that
really keeps public health experts on edge.
A highly contagious animal virus.
It's already causing huge outbreaks in livestock, and then
suddenly it develops. This, this terrifying new
ability can attack the brain. What fundamentally shifts when a
(00:35):
virus, one we mostly know for inrespiratory illness, starts
showing clear signs of neurological impact?
How would that kind of change force us to completely rethink,
well, everything our understanding of the risks and,
crucially, our whole strategy for protection?
Well, this isn't just hypothetical anymore.
This is precisely the concern, the escalating concern around
(00:55):
the highly pathogenic avian influenza AH5N1 Clay 2.3 point
4.4 B viruses. These strains, they're already
fueling a significant ongoing outbreak in dairy cows across
the United States. And while the human cases we've
seen linked to H5 N 1, they've largely been mild so far.
The really unsettling question is, what if some of these
(01:17):
circulating strains actually possess traits that could
rapidly, dramatically change that picture, make human
infections far more severe? And that's exactly where today's
deep dive takes a critical turn.We're getting into some new,
really groundbreaking research that unpacks precisely this.
It reveals a specific H5 N 1 isolate originally from a cow,
(01:38):
actually with a surprising and frankly, deeply concerning
ability to cause severe, rapidlyfatal neurologic disease in a
key animal model. So we're not just talking
respiratory distress now. It's like a direct assault on
the brain. But before we delve deeper into
the, well, the alarming specifics of this research, it's
really essential we shine a spotlight on the team, the
dedicated team whose critical findings are shaping how we
(02:00):
understand this evolving. Who are the minds behind this
work? Yeah, absolutely.
We really celebrate the work of Thomas Tippevignesh, Mary Eppen,
Kaywe Sienda, Kimberly Mead White, Matthew Lewis at Sushi
Okamura, Natalie McCarthy, Ekaterina Elton, Nova Shenna S
Leventhal, Trenton Bushmaker, Chad S Clancy, Emmy DeWitt,
Vincent J Munster, Heinz Feldmanand Kyle Rosenke.
(02:20):
They're from the Laboratory of Virology, Division of Intramural
Research. NIAINIH up in Hamilton, Mt Their
work, it's significantly advanlyadvancing our understanding of
HPAIA H5, N1 virulence, and you know what it could all mean.
OK, so let's set. The stage a bit what?
Exactly Is HPAI AH5N1 and why has it become such an urgent
(02:41):
global concern like right now? Right.
Well, the highly pathogenic avian influenza AH5N1,
specifically what's known as thegoose Guangdong lineage.
That's basically a particular ancestral strain they found in
geese at first, popped up in China back in 1996.
Since then, this virus has just been a master of evolution.
It's constantly adapting, branching out into 10 main
phylogenetic clades. You can think of these clades as
(03:02):
sort of distinct genetic families, right?
Subgroups, each with slightly different traits and behaviors.
And clade 2 has really proven tobe the most dominant, the most
widespread. Starting around 2005, these HPA
IH5 N 1 Clade 2 viruses began tospread well pretty aggressively
across Asia, Europe, Africa, andthen eventually they made their
(03:23):
way to North and South America. And it's not just a threat to
birds anymore, is it? We've seen this worrying trend,
this spillover into mammals. Absolutely.
Since late 2021, these H5 N 1 clay 2.3 point 4.4 B viruses
have been continuously circulating in North America.
Huge outbreaks in wild birds, domestic poultry.
But what's really alarming is the increasing spillover, you
(03:45):
know, the virus jumping from birds into wild mammals.
We've documented cases in farmedmink in Spain that was 2022,
Arctic foxes, raccoon dogs in Finland, domestic cats in
Poland, even green mammals down across South America.
And just recently a Mountain Lion was found dead in Montana,
also infected with H5 N one. It just highlights how broad its
impact on mammals can be. And then things really escalated
(04:08):
with the dairy cows. That really caught.
Everyone's attention didn't. It almost became a household
name. It really did.
In March 2024, HPAIA H5 N 1 Clayed 2.3 point 4.4 B viruses
were found in dairy cows and crucially, in unpasteurized milk
across the US. Now, as of when this paper came
out, 70 human cases of HPA IH5 None had been reported in the
(04:31):
United States. These were caused by either the
B 3.13 genotype from the dairy cows or the D 1.1 genotype from
birds. And while most human cases have
been mild, often just conjunctivitis or, you know,
pink eye respiratory symptoms have definitely been described.
And in a pretty sobering case upin Canada, a teenage girl
actually needed hospitalization for severe respiratory failure.
(04:51):
Thankfully, she did recover. So the main risk for humans
getting infected here is mostly occupational right for people
working closely with these animals.
That's a big part of it. Yes, occupational exposure is
common, especially for folks on dairy and poultry farms.
But we've also seen severe humancases from exposure to sick and
dead birds in just backyard flocks.
(05:11):
So the exposure sources aren't strictly limited to large
commercial operations. And a really critical point here
is that there's clear evidence of mammal to mammal transmission
happening in some cases. Yet despite all this, the USCDC
currently categorizes the overall public health risk for
human infection as low. But the big So what?
Here, the really profound point is that this low risk could
(05:33):
change, and it could change veryquickly if the virus keeps
evolving, picking up new geneticchanges.
This particular study, it seems it dies right into that critical
gap in our understanding of the virus's virulence, its potential
punch. Precisely, it fills a really
crucial knowledge gap. OK, so with such a pressing
evolving threat, how did the researchers actually go about
investigating the virulence these specific H5 N 1 strains?
(05:55):
What was their their approach? Well, they started with a very
clear comparative goal. They wanted to assess the
virulence, basically how harmfulthe virus is of a well known
reference HPAIA H5 N 1 clade onevirus that's Vietnam 1203, 2004
or VN123. And they compared this benchmark
against three very recent HPII AH5N1 clade 2.3 point 4.4 B
(06:20):
isolates. These newer ones were sourced
from, you know, highly relevant situations, one from a cow, one
from a Mountain Lion and anotherfrom mink.
The bovine isolate, a bovine OHB24 O SU-342200 four that was
obviously of particular interestgiven the ongoing dairy cow
outbreaks. And they use mouse models for
this, right? Which specific strains did they
use and why are those models important here?
(06:41):
That's right, they use two established mouse strains,
C57BL6J and BALBC mice. These are pretty standard in
virology because we know a lot about how they respond to
influenza viruses. It makes them reliable for these
kinds of comparative studies andto mimic how exposure might
happen in the real world. They compared 2 main ways of
giving the virus intranasal, which is like inhaling virus
particles, and orgastric, where the virus goes in via a feeding
(07:03):
tube simulating eating or drinking something contaminated.
Right, that orgastric route seems particularly relevant for
how it might spread through contaminated milk, for example.
Exactly. It's key for understanding that
potential pathway. So once the mice were exposed,
what kind of data were they collecting?
What were they measuring to see the impact?
They tracked several key things really meticulously, daily
(07:24):
monitoring for clinical signs, so the visible symptoms of
sickness, weight loss, ruffled fur, hunched posture, rapid
breathing, that sort of thing. And critically, they watched
very closely for neurological signs, tremors, ataxia.
That's a loss of coordination, hyperactivity, circling
behaviors. They also tracked survival rates
for up to 28 days. And then to understand how the
(07:47):
virus was replicating and spreading inside the body, they
measured virus titers. That's the amount of virus
present in organs like the lung,liver and brain, and also in the
blood. They collected these at the peak
of the disease. So they weren't just watching
symptoms, they were digging deeper, looking at what was
happening at the cellular level systemically.
Exactly. They used histopathology, which
(08:07):
is like doing microscopic forensics on the tissues looking
for any cell damage or changes caused by the virus.
They also used immunohistochemistry.
That's a technique using specialmarkers to basically light up
exactly where the virus parts, its antigens are located within
the brain and lung cells. And to understand the immune
response. They did cytokine analysis in
brain and lung tissue. Cytokines are like the immune
(08:30):
system's alarm signals. They looked at specific pro
inflammatory ones like IL 1, GMCSF and MIP one which tell you
the body's mounting a strong, maybe even overwhelming
inflammatory response. And finally, specifically for
that bovine isolate, they did low dose studies to figure out
its median lethal dose, the LD 50.
That tells you how little virus it takes to be fatal to half the
(08:51):
animals, giving you a real senseof its potency.
Wow, quite comprehensive. So after all this meticulous
work, what did they find was thebig discovery?
This is where the H5 N one storyseems to take a really
concerning turn. Yeah, the most impactful result,
no question, revolves around that bovine isolate.
It showed dramatically enhanced virulence and really an
unprecedented ability to invade the brain, what we call
(09:14):
neuroinvasion. Compared to all the other
strands they tested in the C57BL6J mice, the bovine isolate
caused uniform lethality. Didn't matter if it was given
intranasally through the nose oror gastricly by ingestion.
Uniform lethality. Yep, intranasally mice died by
day four or gastricly by day 6. This disease progression was
(09:34):
significantly faster and more severe than what they saw with
the mink Mountain Lion or even that reference VN1 Toll 3
string. And those neurological signs you
mentioned, That sounds like the really striking and honestly
quite terrifying part. It absolutely is.
Crucially, every single C57BL6J mouse given the bovine isolate
developed clear, undeniable neurologic signs.
(09:55):
We're talking tremors, ataxia, that loss of coordination again,
hyperactivity circling. This level of neuroinvasion was
largely unique to the bovine isolate.
Only two of the Mountain Lion infected mice showed just minor
temporary hyperactivity. It was a really stark contrast
and matching up with these severe neurological signs.
The researchers found extremely high virus titers, huge amounts
(10:16):
of virus in the brains of those bovine infected C57BL6J mice,
way higher than any other isolate where the brain either
had no virus or very low levels.Plus the bovine isolate showed
enhanced replication and other organs too, and cause systemic
infection. They found significantly higher
levels of virus circulating in the blood.
That's viremia. It shows the virus wasn't just
(10:38):
staying put, it was spreading all through the body.
OK, so we've got high viral loads in the brain, unmistakable
neurological symptoms. But here's the really
groundbreaking part of the paradox you mentioned earlier.
What about the actual damage to the brain tissue itself?
That's where things got weird, right?
This was a genuinely perplexing finding.
Yeah. And it forces us to sort of
rethink how these viruses can cause trouble.
(11:00):
Despite those incredibly high viral loads and the clear
presence of viral antigen, the virus bits inside the neurons
and glial cells in the brain, when they looked at the brain
tissue using histopathology, they saw no significant physical
tissue damage or inflammation. No damage but clear symptoms and
virus present. Exactly.
It's not just a fascinating biological puzzle, it really
(11:21):
shifts our understanding of how viruses can cause neurological
disease. It suggests the neurological
signs might not be from direct cell destruction or a big
inflammatory attack, but maybe from more subtle neuronal
metabolic disturbances. Like imagine the virus subtly
messing with the brain cells energy systems, causing
dysfunction without obvious structural damage.
(11:44):
This invisible damage mechanism is a critical new piece of the
puzzle. That's wild.
It is, but it's important to note while they didn't see the
physical damage, the bovine isolate did trigger a
significant pro inflammatory cytokine response in the brain
tissue. Those elevated Illinois 1GMC S
FM IP110 levels. So the brain's immune system was
definitely reacting, even if thestructure looked OK.
(12:05):
And you mentioned differences between the mouse strains.
Did one turn out to be more useful for studying these
neurological effects? Yes.
Interestingly, while the Bao BC mice actually died a day earlier
from the bovine strain than the C57BL6J mice, it was only the
C57BL6J mice that consistently showed those obvious severe
(12:26):
neurologic signs. This happened even though both
strains had similar amounts of virus in their brains at later
stages. So it might be down to different
immune responses between the strains, or maybe subtle
differences in how the disease progresses overall.
Perhaps the BLLBC mice just succumbed faster to the
respiratory illness before the neurological signs really had a
chance to fully develop. Interesting.
(12:48):
So yeah, the C57BL6J model proved particularly valuable for
directly studying this neuro invasion and neurovirulence.
And let's just circle back to that orgastric route, ingesting
the virus. How significant was that finding
overall? It was very significant.
This study strongly reinforced its importance.
It confirmed it as a pertinent way for the bovine isolate to
cause infection, leading to rapid disease and death.
(13:10):
This has really direct implications for how this virus
might spread in the real world, you know, beyond just
respiratory transmission. OK, so let's try and pull this
all together. What does this all mean for us?
For the ongoing H5 N 1 situation?
For how we think about viral threats?
What are the big takeaways? Well, the central implication is
pretty stark, really. The increased virulence and the
(13:30):
neuroinvasion shown by this specific bovine H5 N 1 isolate
in these mouse models is franklydeeply concerning for both
animal and public health. It tells us this virus is
clearly evolving in ways that could lead to more severe
disease. It's a serious wake up call that
the virus isn't static, it's changing.
We saw the. Virus.
Got into the brain, but the exact mechanism, the how isn't
(13:53):
totally. Clear yet right?
That's right, The precise entry points aren't fully nailed down.
But the presence of viremia virus in the blood strongly
suggests hematogynous spread, like the virus gets into the
bloodstream and then has to somehow cross the blood brain
barrier. That protective shield around
the brain for viruses entering through the nose like in the
intranasal tests, direct spread along cranial nerves, the nerves
(14:15):
connecting the nose straight to the brain, seems like a very
plausible route. And what's really fascinating
for the orgastric route, when the virus is swallowed,
researchers are looking at thesenerve networks in the gut wall,
the submucosal and myenteric plexi.
These could potentially act as direct neural pathways from the
gut right up to the brain. That would be a pretty novel way
(14:35):
for a virus like this to cause neuroinvasion.
Definitely needs more investigation.
And that paradox you described, the neurological signs without
obvious brain lesions, What's the thinking there?
How does that change things? Yeah, the absence of those clear
histological lesions, despite the strong neurological signs in
the virus being present, it is really intriguing.
It strongly supports the idea that the symptoms might be due
(14:58):
to those neuronal metabolic disturbances, not direct cell
killing or massive inflammation.This has big implications for a
diagnosis and maybe even treatment.
If doctors are only looking for physical brain damage to confirm
neurological H5 N one they mightmiss cases.
It suggests we might need to focus more on functional
changes, maybe biochemical markers of neuronal stress.
(15:19):
It's an area needing a lot more research.
Now, we often hear about specific genetic mutations
making viruses nastier. Did this study find any clues
about what might be making this bovine isolate so aggressive and
neuroinvasive? It did, but with an interesting
twist. Some mutations that we've
previously linked to higher virulence and other H5N1
(15:39):
strains, things like the NA stock deletion, NS-1 deletion or
that PB2E627K mutation. Specific genetic tweets known to
boost disease severity, right? They were actually missing in
the recent isolate studied here,including the Bovine 1.
This suggests that other may be yet to be defined molecular
markers or perhaps combinations of genetic factors are driving
(16:01):
their enhanced virulence in mammals.
However, these recent isolates do have mutations in the matrix
protein M1 and NS-1 that are known to increase virulence in
mice. So while some expected flags
weren't there, other subtle changes are clearly playing a
role. It just highlights how complex
viral evolution is and why we need constant surveillance.
And let's hit that orgastric exposure route one more time,
(16:23):
because it's confirmation here seems incredibly relevant right
now. It is absolutely.
This study really hammers home its importance as a potent
exposure route. It gives us a very plausible
explanation for infections we'reseeing in wild and domestic
carnivores like that Mountain Lion, who probably get infected
by eating infected prey. But critically, it also
(16:43):
highlights a potential infectionroute for other mammals,
including humans, including cats, from drinking raw,
unpasteurized milk from infecteddairy cows.
That's a major point of concern this research underscores,
emphasizing the public health advice about avoiding
unpasteurized dairy. So these mouse models, even with
their slight differences, they're proving to be really
invaluable tools in this whole H5 N 1 situation, aren't they?
(17:05):
Oh immensely valuable the C57BL6J and BALBC mice are
excellent in vivo meaning in a living Organism screening
models, they're just crucial forquickly evaluating potential
antiviral drugs, testing different therapies and checking
how well new vaccine candidates work against these emerging H5 N
1 viruses. And specifically, like we
(17:26):
discuss the C57BL6J model is turning out to be particularly
useful for digging into neuroinvasion and the mechanisms
behind neurovirulence. They're giving us critical fast
insights that directly help our public health responses and
preparedness. So I think the central insight
from this deep dive, it's just unequivocally clear this highly
pathogenic avian influenza AH5 N1 bovine isolate, the one
(17:48):
actually circulating out there, shows significantly enhanced
virulence in these mouse models.It rapidly causes fatal
respiratory disease and severe neurologic disease.
This unique ability to invade the nervous system even without
causing obvious visible brain damage really underscores the
dynamic evolving nature of this virus.
It shows a concerning potential for unexpected and severe
impacts. So what does this powerful
finding really mean for how we respond to the ongoing H5 N 1
(18:10):
outbreak and how urgently shouldwe be preparing for potential,
maybe even drastic shifts and how this virus behaves,
especially thinking about transmission routes like raw
milk? This episode was based on an
Open Access article under the CCPY 4 point O license.
You can find a direct link to the paper and the license in our
episode description. If you enjoyed this analysis,
(18:31):
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