Long-term immune responses induced by low-dose infection with high pathogenicity avian influenza viruses can protect mallards from reinfection with a heterologous strain

Migratory water birds are considered to be carriers of high pathogenicity avian influenza viruses (HPAIVs). In Japan, mallards are often observed during winter, and HPAIV-infected mallards often shed viruses asymptomatically. In this study, we focused on mallards as potential carriers of HPAIVs and investigated whether individual wild mallards are repeatedly infected with HPAIVs and act as HPAIV carriers multiple times within a season. Mallards were experimentally infected with H5N1 and H5N8 HPAIVs that were isolated recently in Japan and phylogenetically belong to different hemagglutinin groups (G2a, G2b, and G2d). All of these strains are more infectious to mallards than to chickens, and the infected mallards shed enough virus to infect others, regardless of whether they exhibited clinical signs. Serum antibodies to the homologous antigen, induced by a single infection with a low virus dose (10 times the 50% mallard infectious dose), were maintained at detectable levels for 84 days. Immunity at 84 days post-inoculation fully protected the mallards from a challenge with the homologous strain, as demonstrated by a lack of viral shedding, and antibody levels did not increase significantly in most of these birds. Protection against heterologous challenge was also observed despite undetectable levels of antibodies to the challenge strain. Our findings suggest that repeated infections with homologous and heterologous HPAIV strains do not occur frequently in individual wild mallards within a season, particularly at low viral doses, and the frequency with which they act as carriers may be limited.