The recent H5N1 panzootic has expanded to include sustained circulation in US dairy cattle. The haemagglutinin (HA) protein is a primary determinant of host receptor recognition and a key factor in assessing zoonotic potential. The study assessed receptor-binding specificity of HA variants identified in viruses circulating in Californian dairy cattle using bio-layer interferometry and pseudotype virus entry assays.
All tested HA variants retained a clear preference for avian-type α2,3-linked sialic acid receptors. No switch to human-type α2,6-linked sialic acid binding was reported in the tested substitutions. These findings indicate that, despite ongoing circulation in a mammalian host and repeated spillover into humans, the studied HA variants have not acquired receptor specificity changes associated with efficient human upper respiratory tract infection.
Beyond receptor binding, HA stability—measured by fusion pH and thermostability—affects transmission dynamics and environmental persistence. The investigators evaluated HA stability using cell‑based fusion assays and thermostability measurements.
All mutant HAs examined exhibited fusion pH values greater than 5.5. This is above the fusion pH range (pH 5.0–5.5) that has been associated with efficient airborne transmission in humans. Therefore, the stability phenotypes measured in these cattle-origin H5N1 viruses do not fall within the range that has been linked to enhanced airborne transmissibility in humans.
However, some substitutions did alter HA stability modestly. Two specific mutations increased pH stability compared with the wild-type HA, and viruses carrying combinations of mutations demonstrated increased resistance to thermal inactivation.
Two substitutions in the HA sequence—D88G and S94N—were highlighted for their effects on stability. Compared with the wild-type HA fusion pH of 5.9, the D88G and S94N substitutions lowered the fusion pH to 5.6 and 5.7, respectively, indicating increased pH stability. When both mutations were present in the same HA background, the viruses displayed increased thermostability in the assays performed.
Although these changes reflect enhanced stability relative to the wild type, the resulting fusion pH values remained above pH 5.5 and therefore outside the window associated with efficient airborne human transmission. The report does not describe additional functional consequences beyond the measured receptor binding and stability phenotypes.
The published data place these molecular findings in a broader epidemiologic context. The H5N1 event in US dairy cattle was first detected in March 2024. By July 2026, infections had been reported on more than 1,150 dairy farms across 19 states. While the initial outbreak centred in Texas, California emerged as the principal focus for onward transmission and accounted for most human infections attributed to exposure to infected dairy cattle.
Continued transmission within cattle populations and repeated spillover events into humans increase the opportunities for selection of mammalian-adaptive mutations. The study emphasizes that even modest changes in HA stability can arise during circulation in a mammalian host, warranting attention.
The key conclusion is that cattle-origin H5N1 viruses in this investigation retained avian-like receptor specificity while acquiring mutations that modestly enhance HA stability. Because increased HA stability can be one component of adaptations that facilitate mammalian transmission, the authors stress the importance of integrated genomic and phenotypic surveillance in animal reservoirs.
Detecting and characterizing emerging HA mutations—both sequence changes and their phenotypic effects on receptor binding, fusion pH, and thermostability—will help identify variants that could increase zoonotic or pandemic risk. The source notes these findings without claiming that the studied mutations have produced human-adapted transmission; rather, they underscore a surveillance imperative as H5N1 continues to circulate in dairy cattle and spill over into humans.
Competing interests were declared absent by the authors in the source. Funding sources were reported but specific grant details are listed in the original report and are not expanded here.