In this study, we investigated the clinical response, viral shedding, transmissibility, pathologic lesions, and tropism of HPAIV Gs/Gd H5N8 subtype (clade 2.3.4.4b), following experimental infection of three groups of captive mallards (Anas platyrhynchos): (i) fully susceptible, (ii) pre-exposed to low pathogenic avian influenza virus (LPAIV) H5N1 subtype, and (iii) pre-exposed to LPAIV H3N8 subtype. Infection of naïve mallards with HPAIV H5N8 resulted in ~60% mortality, neurological signs, abundant shedding, and transmission to contact ducks, who also became sick and died. High amounts of viral RNA were found in all collected organs, with the highest RNA load recorded in the brain. The IHC examinations performed on tissues collected at 4 and 14 days post-infection (dpi) revealed tropism to nervous tissue, myocardium, respiratory epithelium, and hepatic and pancreatic cells. The mallards pre-exposed to LPAIV H5N1 and challenged with HPAIV H5N8 were asymptomatic and showed a significant reduction of viral RNA shedding, yet still sufficient to cause infection (but no disease) in the contact ducks. The AIV antigen was not detected in organs at 4 and 14 dpi, and microscopic lesions were mild and scarce. Similarly, mallards previously inoculated with LPAIV H3N8 remained healthy after challenge with HPAIV H5N8, but viral RNA was detected in large quantities in swabs and organs, particularly in the early phase of infection. However, in contrast to mallards from group I, the IHC staining yielded negative results at the selected timepoints. The virus was transmitted to contact birds, which remained symptomless but demonstrated low levels of viral RNA shedding and mild- to moderate tissue damage despite negative IHC staining. The results indicate that naïve mallards are highly susceptible to HPAIV H5N8 clade 2.3.4.4b and that homo- and heterosubtypic immunity to LPAIV can mitigate the clinical outcomes of infection.
Keywords: highly pathogenic avian influenza; immunity; low pathogenic avian influenza; mallards; susceptibility.