H5N1 clade 2.3.4.4b avian influenza viruses replicate in differentiated bovine airway epithelial cells cultured at air-liquid interface.

Luca Bordes1 Nora M Gerhards1 Stan Peters1 Sophie van Oort1 Marit Roose1 Romy Dresken1 Sandra Venema1 Manouk Vrieling1 Marc Engelsma1 Wim H M van der Poel1 Rik L de Swart1
Affiliations 1 institutions
  1. Wageningen Bioveterinary Research, Lelystad, Netherlands.

Abstract

Highly pathogenic avian influenza (HPAI) H5N1 viruses are responsible for disease outbreaks in wild birds and poultry, resulting in devastating losses to the poultry sector. Since 2020, an increasing number of outbreaks of HPAI H5N1 was seen in wild birds. Infections in mammals have become more common, in most cases in carnivores after direct contact with infected birds. Although ruminants were previously not considered a host species for HPAI viruses, in March 2024 multiple outbreaks of HPAI H5N1 were detected in goats and cattle in the United States. Here, we have used primary bronchus-derived well-differentiated bovine airway epithelial cells (WD-AECs) cultured at air-liquid interface to assess the susceptibility and permissiveness of bovine epithelial cells to infection with European H5N1 virus isolates. We inoculated bovine WD-AECs with three low-passage HPAI clade 2.3.4.4b H5N1 virus isolates and detected rapid increases in viral genome loads and infectious virus during the first 24 h post-inoculation, without substantial cytopathogenic effects. Three days post-inoculation infected cells were still detectable by immunofluorescent staining. These data indicate that multiple lineages of HPAI H5N1 may have the propensity to infect the respiratory tract of cattle and support extension of avian influenza surveillance efforts to ruminants. Furthermore, this study underscores the benefit of WD-AEC cultures for pandemic preparedness by providing a rapid and animal-free assessment of the host range of an emerging pathogen.

Supporting text Virus Host Location
air-liquid interface 1 airway epithelial cells 1 cattle 132 cow 1 H5N1 82 highly pathogenic avian influenza 15 ruminants 9 Epithelial Cells 27 Influenza A Virus, H5N1 Subtype 300 Virus Replication 191 Animals 1948 Cattle 126 Cells, Cultured 26

Evidence records

3 total
Zoonotic Surveillance
1 records · 1 evidence types
Evidence type
1 records
OVE8158
Key finding

Infectious HPAI clade 2.3.4.4b H5N1 virus was recovered and propagated following inoculation of primary well-differentiated bovine airway epithelial cells cultured at air–liquid interface.

Virus
Host
Location
Not specified
Supporting text

We inoculated bovine WD-AECs with three low-passage HPAI clade 2.3.4.4b H5N1 virus isolates and detected rapid increases in viral genome loads and infectious virus during the first 24 h post-inoculation, without substantial cytopathogenic effects.

Sample type
primary bovine airway epithelial cells cultured at air–liquid interface
Transmission Evidence
1 records · 1 evidence types
Evidence type
1 records
OVE8160
Key finding

Multiple outbreaks of highly pathogenic avian influenza (HPAI) H5N1 were detected in goats and cattle in the United States in March 2024.

Virus
Host
Location
Supporting text

In March 2024 multiple outbreaks of HPAI H5N1 were detected in goats and cattle in the United States.

Method
outbreak detection
Transmission direction
unknown
Geographic raw
United States
Country inferred
USA
Outbreak time
March 2024
Outbreak scale
multiple outbreaks
Experimental Infection
1 records · 1 evidence types
Evidence type
1 records
OVE8159
Key finding

European H5N1 clade 2.3.4.4b avian influenza virus isolates were able to replicate in differentiated bovine airway epithelial cells, demonstrating ruminant cell susceptibility and permissiveness.

Virus
Host
Location
Not specified
Supporting text

We used primary bronchus-derived well-differentiated bovine airway epithelial cells (WD-AECs) cultured at air-liquid interface to assess the susceptibility and permissiveness of bovine epithelial cells to infection with European H5N1 virus isolates. Inoculated bovine WD-AECs supported rapid increases in viral genome loads and infectious virus during the first 24 h post-inoculation, and infected cells remained detectable by immunofluorescent staining at three days post-inoculation.

Method
in vitro infection assay | viral genome load quantification | infectious virus titration | immunofluorescent staining
Sample type
airway epithelial cell cultures
Experimental system
air-liquid interface cell culture system using well-differentiated bovine airway epithelial cells