Molecular, Functional, and Ecological Characterization of Antarctic Penguin Orthoavulaviruses (AVV17–19)

Emerging avian avulaviruses (AVVs) have been identified in Antarctic ecosystems; however, their biological properties and current circulation remain poorly understood. This study utilized historical egg-isolated Antarctic penguin avulavirus strains (AVV17, AVV18, and AVV19) and combined molecular characterization with field surveillance during the 2024-2025 Antarctic season. The historical isolates were molecularly confirmed using RT-qPCR targeting the L gene, confirming their classification as AVV17, AVV18, or AVV19. These isolates were tested for replication indicators in REM 134, MDCK cells and embryonated chicken eggs. Only AVV18 showed evidence of productive replication in REM 134 cells, as indicated by decreasing Ct values and the presence of cytopathic effects.AVV17 and AVV19 were detectable by RT-qPCR but did not exhibit cytopathic changes or replication dynamics. None of the viruses replicated efficiently in MDCK cells. Further propagation of AVV18 in embryonated chicken eggs showed viral amplification in some eggs. Concurrent surveillance at five sites in the South Shetland Islands detected low-level circulation of AVV17-19, exclusively in cloacal swabs, with no viral RNA found in environmental samples. These findings link historical isolates to current circulation, highlighting selective replication among Antarctic AVVs and limited ecological spread in penguin hosts.