Frontiers in Veterinary Science,
Journal Year:
2025,
Volume and Issue:
11
Published: Jan. 10, 2025
Highly
pathogenic
avian
influenza
(HPAI)
subtype
H5Nx
viruses
of
the
A/Goose/Guangdong/1/1996
(Gs/Gd)
lineage
have
led
to
substantial
economic
losses
within
poultry
industry
and
represent
an
ongoing
public
health
threat
[1].
The
Gs/Gd
H5
not
only
evolved
into
ten
primary
clades
0-9
with
their
subclades
but
also
reassorted
other
A
[2;
3;
4].
Notably,
since
2020,
clade
2.3.4.4b
HPAI
H5N1
caused
outbreaks
across
a
broad
geographic
range,
including
Asia,
Europe,
Africa,
North
America,
South
Antarctica
[5;
6;
7].
infections
in
mammals
wild,
domestic
humans
underscore
potential
zoonotic
risk
pandemic
these
evolving
[8].In
Korea,
multiple
outbreaks.
During
October
2022-March
2023,
total
16
different
genotypes
HPAIV,
Kor22-23A-P,
were
reported
wild
birds,
showing
high
genetic
diversity
HPAIVs
generated
through
frequent
reassortment
[9].
December
2023-May
2024,
H5N6
[10;
11]
32
cases
farms
(home.kahis.go.kr)
19
birds
(http://wadis.go.kr).
No
virus
had
been
detected
Korea
June
despite
large-scale
active
surveillance
targeting
both
poultry.
Here,
we
report
detection
isolated
from
captured
Mandarin
duck
(Aix
galericulata)
on
15,
Northern
pintail
(Anas
acuta)
found
dead
17,
during
early-stage
fall
migration
waterfowl
Korea.
To
facilitate
timely
information
sharing,
conducted
complete
genome
sequencing
using
Illumina
next-generation
(NGS)
technology
submitted
sequences
GISAID
database
(https://www.gisaid.org).
comparative
phylogenetic
analysis
was
carried
out
determine
virus's
origin
genotype.Materials
MethodsOn
15
8
ducks
along
Cheongmicheon
stream
Gyeonggi-do
Province,
(GPS
coordinate:
37°8'31.25"N,
127°22'52.23"E)
as
part
national
bird
program
(Supplementary
Figure
1).
On
17
at
Yongsu
reservoir
Jeju
island
33°30'18.28"N,
126°53'33.4"E).
We
collected
oropharyngeal
cloacal
swabs
birds.
Swab
samples
placed
phosphate-buffered
saline
(PBS)
containing
400
mg/mL
gentamicin
thoroughly
homogenized
by
vortexing
for
1
min.
supernatant
filtered
0.45-μm
Minisart
Syringe
Filter
(Sartorius,
Göttingen,
Germany)
after
centrifugation
sample
3000
rpm
10
min
inoculated
10-day-old
specific-pathogen-free
(SPF)
embryonated
chicken
eggs.
After
72
h
incubation
37•C,
allantoic
fluids
harvested
tested
hemagglutination
activity
(HA)
10%
red
blood
cells.
RNA
extracted
hemagglutination-activity-positive
fluid
Maxwell
RSC
simply
Tissue
Kit
(Promega,
Madison,
WI,
USA)
according
manufacturer's
instructions
screened
matrix
(M)
genes
real-time
reverse
transcription-PCR
(rRT-PCR)
previously
described
[12].Complementary
DNA
SuperScript
III
First-Strand
Synthesis
system
(Invitrogen,
Carlsbad,
CA,
USA),
eight
gene
segments
amplified
AccuPrime
Pfx
Polymerase
[13].
libraries
prepared
Nextera
Flex
Library
Prep
(Illumina,
San
Diego,
which
utilizes
transposon-mediated
tagmentation
adapter
ligation,
dual-index
barcodes
instructions.
sequenced
paired-end
150
Miseq
sequencing-by-synthesis
platform.
NGS
raw
reads
trimmed
adapters
low-quality
bases
BBDuk
version
38.84
setting
minimum
quality
30
[14].
Trimmed
assembled
de
novo
SPAdes
assembler
3.15.5.
mapped
top
result
EpiFlu
database,
identified
contigs,
Minimap
2.24
(https://github.com/lh3/minimap2)
default
options
visualized
Geneious
Prime
software.The
produced
reference-guided
assembly
used
generate
final
consensus
sequences.
dataset
presented
this
study
can
be
online
repositories.
names
repositories
accession
ID
is
available
GISAID(https://www.gisaid.org)
EPIFlu
(accession
ID:
EPI_ISL_19528860
EPI_ISL_19531393).
classification
performed
subspecies
tool
BV-BRC
(https://www.bvbrc.org/app/SubspeciesClassification).
examined
identify
molecular
markers
associated
mammalian
host
adaptation,
pathogenicity,
drug
resistance.
utilized
FluSurver
mutation
Initiative
[15]
manual
screening
based
known
impacting
AIV
biological
properties
[16].
Identified
amino
acid
substitutions
HA
segment
are
referenced
numbering.All
analyzed
BLAST
query
function
(https://gisaid.org/).
From
500
hits,
identical
ElimDupes
software
(https://www.hiv.lanl.gov/content/sequence/elimdupesv2/elimdupes.html).
Genome
aligned
MAFFT
[17].
Phylogenetic
tree
construction
each
RAxML
v8.0
[18]
general
time
reversible
model
nucleotide
substitution
Gamma
rate
heterogeneity,
1,000
bootstrap
replicates.
Interactive
Tree
Life
(iTOL)
employed
visualize
[19].
cluster
regarded
distinct
when
it
support
value
>
70
sequence
identity
97%.
genotype
G2b
G2d
2021
2022
[20;
21]
verify
viruses.
A/goose/Hunan/SE284/2022(SE284)
(H5N1)
[22]
categorize
G2c
viruses.A
Bayesian
relaxed-clock
phylogeny
reconstructed
BEAST
1.10.4
[23],
applying
Hasegawa,
Kishino,
Yano
uncorrelated
log-normal
distribution
Gaussian
Markov
Random
Field
(GMRF)
skyride
coalescent
prior
[24].
Chain
Monte
Carlo
(MCMC)
process
run
parallel
three
chains,
50
million
iterations,
results
combined
burn-in.
All
parameters
achieved
effective
sizes
>200
TRACER
v1.5
(http://tree.bio.ed.ac.uk/software/tracer/)
[25].
maximum
credibility
(MCC)
created
TreeAnnotator
FigTree
v1.4.4
(http://tree.bio.ed.ac.uk/software/figtree/).
most
recent
common
ancestor
(tMRCA)
estimated
height
values
nodes.A
live
2024
positive
via
embryo
inoculation
rRT-PCR.We
successfully
viruses,
designated
A/Mandarin
duck/Korea/24WS005-2/H5N1/2024
(hereafter
MD/24WS005-2)
A/Northern
pintail/Korea/24WC025/H5N1/2024
NP/24WC025).
52,864
136,268
generated,
respectively,
resulting
coding
(CDS)
all
average
depth
(>350).
presence
basic
acids
proteolytic
cleavage
site
(PLREKRRKR/G)
[26]
classified
2.3.4.4b.The
NP/24WC025
MD/24WS005-2
constellations,
suggesting
independently
introduced
(Figure
that
circulating
Japan
2023-2024.The
belonged
sub-lineage
21].
shared
ancestry
A/white-tailed
eagle/Hokkaido/2024,
A/chicken/Hokkaido/E012/2024,
concurrently
Japan,
tMRCA
April
28,
(95%
BCI:
January
10,
-July
29,
2024),
descendants
circulated
early-mid
2024.For
virus,
HA,
NA,
M
clustered
mainly
2022-2024
Asia
did
form
wellsupported
monophyletic
1D,F,G).
[22].
In
gene,
1A,B,C,E,H).
PB1,
NP,
A/eagle/Korea/22WC464/2023(H5N1)
1B,
D-G)
minor
genotype,
Kor22-23P,
winter
season
2022-2023
PB2,
PA,
NS
derived
Eurasian
LPAI
pool
population
1A,C,H).
has
contributed
generation
donor
lineages
[27].
long
branch
length
new
suggest
undetected
approximately
two
years
undergone
prevailing
population.Genetic
mutations
increased
binding
affinity
α-2,6
sialic
receptors
protein,
N110S
S154N
S133A
T156A
(Table
Both
V588T
PB2
enhance
pathogenicity
mice.
K482R,
polymerase
cell
lines,
virus.
PB1
D3V
D622G,
enhanced
viral
replication
lines
virulence
mice,
present
viruses.Mutations
protein
(N30D,
I43M,
T215A,
P42S),
increase
murine
models,
ESEV
motif
C-terminal
NS1
protein.The
Eurosurveillance,
Journal Year:
2023,
Volume and Issue:
28(31)
Published: Aug. 3, 2023
Since
mid-July
2023,
an
outbreak
caused
by
highly
pathogenic
avian
influenza
A(H5N1)
virus
clade
2.3.4.4b
genotype
BB
is
ongoing
among
farmed
animals
in
South
and
Central
Ostrobothnia,
Finland.
Infections
foxes,
American
minks
raccoon
dogs
have
been
confirmed
on
20
farms.
Genetic
analysis
suggests
introductions
from
wild
birds
scavenging
for
food
farm
areas.
Investigations
point
to
direct
transmission
between
animals.
While
no
human
infections
detected,
control
measures
are
being
implemented
limit
spread
exposure.
EFSA Journal,
Journal Year:
2023,
Volume and Issue:
21(7)
Published: July 1, 2023
Between
29
April
and
23
June
2023,
highly
pathogenic
avian
influenza
(HPAI)
A(H5N1)
virus
(clade
2.3.4.4b)
outbreaks
were
reported
in
domestic
(98)
wild
(634)
birds
across
25
countries
Europe.
A
cluster
of
mulard
ducks
for
foie
gras
production
was
concentrated
Southwest
France,
whereas
the
overall
situation
poultry
Europe
worldwide
has
eased.
In
birds,
black-headed
gulls
several
new
seabird
species,
mostly
terns
(e.g.
sandwich
terns),
heavily
affected,
with
increased
mortality
being
observed
both
adults
juveniles
after
hatching.
Compared
to
same
period
last
year,
dead
seabirds
have
been
increasingly
found
inland
not
only
along
European
coastlines.
As
regards
mammals,
identified
24
cats
one
caracal
Poland
between
10
30
2023.
Affected
animals
showed
neurological
respiratory
signs,
sometimes
mortality,
widely
scattered
nine
voivodeships
country.
All
cases
are
genetically
closely
related
viruses
detected
(since
October
2022,
but
now
sporadic)
(December
2022-January
2023)
past.
Uncertainties
still
exist
around
their
possible
source
infection,
no
feline-to-feline
or
feline-to-human
transmission
so
far.
Since
May
2023
as
4
July
two
clade
2.3.4.4b
detections
humans
from
United
Kingdom,
A(H9N2)
A(H5N6)
human
infections
China.
addition,
person
infected
A(H3N8)
China
died.
The
risk
infection
currently
circulating
H5
remains
low
general
population
EU/EEA,
moderate
occupationally
otherwise
exposed
people
mammals
(wild
domesticated).
Eurosurveillance,
Journal Year:
2023,
Volume and Issue:
28(31)
Published: Aug. 3, 2023
Background
Over
a
3-week
period
in
late
June/early
July
2023,
Poland
experienced
an
outbreak
caused
by
highly
pathogenic
avian
influenza
(HPAI)
A(H5N1)
virus
cats.
Aim
This
study
aimed
to
characterise
the
identified
and
investigate
possible
sources
of
infection.
Methods
We
performed
next
generation
sequencing
phylogenetic
analysis
detected
viruses
Results
sampled
46
cats,
25
tested
positive
for
virus.
The
belong
clade
2.3.4.4b,
genotype
CH
(H5N1
A/Eurasian
wigeon/Netherlands/3/2022-like).
In
Poland,
this
was
responsible
several
poultry
outbreaks
between
December
2022
January
2023
has
been
only
sporadically
since
February
2023.
Viruses
from
cats
were
very
similar
each
other,
indicating
one
common
source
addition,
most
closely
related
dead
white
stork
early
June.
Influenza
possessed
two
amino
acid
substitutions
PB2
protein
(526R
627K)
which
are
molecular
markers
adaptation
mammals.
presented
those
mutations
(627K),
suggests
that
had
spilled
over
already
partially
adapted
mammalian
species.
Conclusion
scale
HPAI
H5N1
infection
is
worrying.
One
seems
be
meat,
but
date
no
such
meat
with
certainty.
Surveillance
should
stepped
up
on
poultry,
also
certain
species
farmed
mammals
kept
close
infected
farms.
Animals,
Journal Year:
2024,
Volume and Issue:
14(9), P. 1372 - 1372
Published: May 2, 2024
Avian
influenza
viruses
(AIVs)
are
highly
contagious
respiratory
of
birds,
leading
to
significant
morbidity
and
mortality
globally
causing
substantial
economic
losses
the
poultry
industry
agriculture.
Since
their
first
isolation
in
2013–2014,
Asian-origin
H5
pathogenic
avian
(HPAI)
clade
2.3.4.4b
have
undergone
unprecedented
evolution
reassortment
internal
gene
segments.
In
just
a
few
years,
it
supplanted
other
AIV
clades,
now
is
widespread
wild
migratory
waterfowl,
spreading
Asia,
Europe,
Africa,
Americas.
Wild
natural
reservoir
LPAIVs
generally
more
resistant
disease,
also
manifested
high
with
HPAIV
2.3.4.4b.
This
caused
overt
clinical
signs
mass
variety
mammalian
species
never
reported
before,
such
as
raptors,
seabirds,
sealions,
foxes,
others.
Most
notably,
recent
outbreaks
dairy
cattle
were
associated
emergence
critical
mutations
related
adaptation,
raising
concerns
about
possibility
jumping
acquisition
sustained
human-to-human
transmission.
The
main
anatomopathological
findings
virus
infection
birds
non-human
mammals
hereby
summarized.
Virus Evolution,
Journal Year:
2024,
Volume and Issue:
10(1)
Published: Jan. 1, 2024
Since
2016,
A(H5Nx)
high
pathogenic
avian
influenza
(HPAI)
virus
of
clade
2.3.4.4b
has
become
one
the
most
serious
global
threats
not
only
to
wild
and
domestic
birds,
but
also
public
health.
In
recent
years,
important
changes
in
ecology,
epidemiology,
evolution
this
have
been
reported,
with
an
unprecedented
diffusion
variety
affected
birds
mammalian
species.
After
two
consecutive
devastating
epidemic
waves
Europe
2020-2021
2021-2022,
second
recognized
as
largest
epidemics
recorded
so
far,
begun
circulate
endemically
European
bird
populations.
This
study
used
complete
genomes
1,956
HPAI
viruses
investigate
during
varying
epidemiological
outline.
We
investigated
spatiotemporal
patterns
to/from
within
2021-2022
waves,
providing
evidence
ongoing
transmission
dynamics
disease
epidemiology.
demonstrated
genetic
diversity
circulating
viruses,
which
undergone
frequent
reassortment
events,
for
first
time
a
overview
proposed
nomenclature
multiple
genotypes
2020-2022.
described
emergence
new
genotype
gull
adapted
genes,
offered
opportunity
occupy
ecological
niches,
driving
endemicity
population.
The
propensity
reassortment,
its
jumps
progressively
wider
number
host
species,
including
mammals,
rapid
acquisition
adaptive
mutations
make
trend
spread
difficult
predict
unfailing
evolving
scenario.
The Journal of Infectious Diseases,
Journal Year:
2024,
Volume and Issue:
230(3), P. 533 - 542
Published: Sept. 15, 2024
Abstract
Since
the
resurgence
of
highly
pathogenic
avian
influenza
(HPAI)
A(H5N1)
virus,
clade
2.3.4.4b,
during
2021,
these
viruses
have
spread
widely
among
birds
worldwide,
causing
poultry
outbreaks
and
infections
a
wide
range
terrestrial
marine
mammal
species.
During
2024,
HPAI
was
detected
in
dairy
cattle
for
first
time
caused
an
ongoing
multistate
outbreak,
with
high
levels
virus
documented
raw
cow
milk.
Human
2.3.4.4b
from
exposures
to
infected
or
resulted
spectrum
illness
severity,
conjunctivitis
mild
respiratory
severe
fatal
pneumonia
different
countries.
Vigilance,
stronger
global
virologic
surveillance
birds,
poultry,
mammals,
humans,
characterization
rapid
data
sharing,
is
needed
inform
threat
viruses,
as
they
continue
evolve,
public
health.
EFSA Journal,
Journal Year:
2024,
Volume and Issue:
22(4)
Published: April 1, 2024
Abstract
Avian
influenza
viruses
(AIV)
remain
prevalent
among
wild
bird
populations
in
the
European
Union
and
Economic
Area
(EU/EEA),
leading
to
significant
illness
death
of
birds.
Transmission
between
mammal
species
has
been
observed,
particularly
fur
animal
farms,
where
outbreaks
have
reported.
While
transmission
from
infected
birds
humans
is
rare,
there
instances
exposure
these
since
2020
without
any
symptomatic
infections
reported
EU/EEA.
However,
continue
evolve
globally,
with
migration
birds,
new
strains
carrying
potential
mutations
for
mammalian
adaptation
could
be
selected.
If
avian
A(H5N1)
acquire
ability
spread
efficiently
humans,
large-scale
occur
due
lack
immune
defences
against
H5
humans.
The
emergence
AIV
capable
infecting
mammals,
including
can
facilitated
by
various
drivers.
Some
intrinsic
drivers
are
related
virus
characteristics
or
host
susceptibility.
Other
extrinsic
may
increase
mammals
thereby
stimulating
mutation
mammals.
Extrinsic
include
ecology
species,
such
as
wildlife,
human
activities
like
farming
practices
use
natural
resources,
climatic
environmental
factors.
One
Health
measures
mitigate
risk
adapting
focus
on
limiting
preventing
spread.
Key
options
actions
enhancing
surveillance
targeting
animals,
ensuring
access
rapid
diagnostics,
promoting
collaboration
sectors,
considering
implementation
preventive
vaccination
poultry.
Effective
communication
different
involved
target
audiences
should
emphasised,
well
strengthening
veterinary
infrastructure,
enforcing
biosecurity
at
reducing
wildlife
contact
domestic
animals.
Careful
planning
poultry
farming,
especially
areas
high
waterfowl
density,
highlighted
effective
reduction.
EFSA Journal,
Journal Year:
2024,
Volume and Issue:
22(7)
Published: July 1, 2024
Abstract
Between
16
March
and
14
June
2024,
42
highly
pathogenic
avian
influenza
(HPAI)
A(H5)
virus
detections
were
reported
in
domestic
(15)
wild
(27)
birds
across
13
countries
Europe.
Although
the
overall
number
of
Europe
has
not
been
this
low
since
2019–2020
epidemiological
year,
HPAI
viruses
continue
to
circulate
at
a
very
level.
Most
poultry
due
indirect
contact
with
birds,
but
there
was
also
secondary
spread.
Outside
Europe,
situation
intensified
particularly
USA,
where
new
A(H5N1)
genotype
(B3.13)
identified
>130
dairy
herds
12
states.
Infection
cattle
appears
be
centred
on
udder,
milk
from
infected
animals
showing
high
viral
loads
representing
vehicle
transmission.
Apart
cattle,
two
other
mammal
species
(alpaca
walrus)
for
first
time.
20
human
cases
infection
Vietnam
(one
A(H5N1),
one
A(H9N2)),
Australia
(with
travel
history
India,
A(H5N1)),
USA
(three
China
(two
A(H5N6),
three
A(H9N2),
A(H10N3)),
India
Mexico
fatal
A(H5N2)
case).
The
latter
case
laboratory‐confirmed
subtype
A(H5N2).
had
exposure
poultry,
live
markets,
or
prior
detection
onset
illness.
Human
infections
remain
rare
no
human‐to‐human
transmission
observed.
risk
currently
circulating
clade
2.3.4.4b
remains
general
public
EU/EEA.
low‐to‐moderate
those
occupationally
otherwise
exposed
contaminated
environments.
EFSA Journal,
Journal Year:
2025,
Volume and Issue:
23(1)
Published: Jan. 1, 2025
A
risk
assessment
framework
was
developed
to
evaluate
the
zoonotic
potential
of
avian
influenza
(AI),
focusing
on
virus
mutations
linked
phenotypic
traits
related
mammalian
adaptation
identified
in
literature.
Virus
sequences
were
screened
for
presence
these
and
their
geographical,
temporal
subtype-specific
trends.
Spillover
events
mammals
(including
humans)
human
seroprevalence
studies
also
reviewed.
Thirty-four
associated
with
five
(increased
receptor
specificity,
haemagglutinin
stability,
neuraminidase
enhanced
polymerase
activity
evasion
innate
immunity)
shortlisted.
AI
viruses
(AIVs)
carrying
multiple
adaptive
belonged
both
low
highly
pathogenic
subtypes,
mainly
A(H9N2),
A(H7N9),
A(H5N6)
A(H3N8),
sporadic
primarily
detected
Asia.
In
EU/EEA,
H5Nx
clade
2.3.4.4b,
which
have
increased
opportunities
evolution
due
widespread
circulation
birds
occasional
cases/outbreaks
mammals,
acquired
highest
number
traits.
Adaptive
traits,
such
as
immune
evasion,
frequently
acquired,
while
receptor-specific
remained
rare.
Globally,
cases
remain
rare,
majority
overall
A(H5N1),
A(H5N6),
A(H7N9)
A(H9N2)
that
are
among
subtypes
tend
a
higher
The
main
drivers
include
host
characteristics,
external
factors
increasing
AIV
exposure
humans
wild
domestic
(e.g.
activities
ecological
factors).
Comprehensive
surveillance
AIVs
targeting
whole
genome
sequencing
animals
is
essential
early
detection
efficient
implementation
control
measures.
All
preparedness,
preventive
measures
must
be
implemented
under
One
Health
tailored
setting
epidemiological
situation;
particular,
monitoring,
biosecurity,
genomic
global
collaboration
critical
mitigating
risks
AIV.
Clade
2.3.4.4b
H5N1
high
pathogenicity
avian
influenza
virus
(HPAIV)
has
caused
a
panzootic
affecting
all
continents
except
Australia,
expanding
its
host
range
to
several
mammalian
species.
In
March
2024,
HPAIV
was
first
detected
in
dairy
cattle
and
goats
the
United
States.
Over
891
farms
across
16
states
have
tested
positive
until
25th
December
with
zoonotic
infections
reported
among
workers.
This
raises
concerns
about
undergoing
evolutionary
changes
that
could
enhance
potential.
The
Influenza
glycoprotein
haemagglutinin
(HA)
facilitates
entry
into
cells
through
receptor
binding
pH-induced
fusion
cellular
membranes.
Adaptive
HA
modulate
virus-host
cell
interactions.
study
compared
genes
of
goat
viruses
dominant
avian-origin
clade
Kingdom,
focusing
on
binding,
pH
fusion,
thermostability.
All
showed
exclusively
avian-like
receptors,
5.9,
outside
associated
efficient
human
airborne
transmissibility
(pH
5.0
5.5).
We
further
investigated
impact
emerging
substitutions
seen
ongoing
outbreaks,
but
saw
little
phenotypic
difference,
continued
exclusive
analogues
pHs
above
5.8.
suggests
from
outbreaks
do
not
pose
an
enhanced
threat
circulating
viruses.
However,
given
rapid
evolution
H5
viruses,
continuous
monitoring
updated
risk
assessments
remain
essential
understanding
pandemic
risks.