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

## TL;DR

This study investigates three avian avulaviruses in Antarctic penguins, finding limited replication and environmental spread.

## Contribution

The study provides novel insights into the replication dynamics and ecological spread of Antarctic penguin avulaviruses.

## Key findings

- AVV18 showed productive replication in REM 134 cells and embryonated chicken eggs.
- AVV17 and AVV19 did not replicate efficiently in tested cell lines or eggs.
- Field surveillance detected low-level circulation of AVV17–19 only in penguin fecal samples, not in environmental samples.

## Abstract

In Antarctic ecosystems, avian avulaviruses (AVVs) contribute to viral diversity in bird populations, particularly penguins. However, little is known about their biological properties and circulation patterns. This study analyzed three historical penguin avulavirus strains (AVV17, AVV18, and AVV19) to better understand their replication and distribution. Using molecular techniques, the viruses were identified and tested for replication ability in cell cultures and embryonated chicken eggs. Only for the AVV18 productive replication in cells was evidenced, while AVV17 and AVV19 showed no efficient replication. Field surveillance in the South Shetland Islands revealed low-level circulation of AVV17, AVV18, and AVV19, detectable only in penguin fecal samples, with no viral presence in environmental samples. These findings provide valuable insight into the replication dynamics and limited ecological spread of AVVs in penguin populations, suggesting that while these viruses circulate, their dissemination in the environment is restricted.

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.