Abstract:
Influenza A viruses (IAVs) are typically isolated and cultured by successive passages using
9- to 11-day-old embryonated chicken eggs (ECEs) and in 14-day old ECEs for virus mutational
studies. Real-time reverse transcription-polymerase chain reaction tests (RT-PCRs) are
commonly used for IAV diagnosis, but virus isolation remains invaluable in terms of its high
sensitivity, providing viable isolates for further studies and the ability to distinguish between
viable and nonviable virus. Efforts at isolating ostrich-origin IAVs from RT-PCR positive
specimens using ECEs have often been unsuccessful, raising the possibility of a species
bottleneck, whereby ostrich-adapted IAVs may not readily infect and replicate in ECEs, yet the
capacity of an ostrich embryo to support the replication of influenza viruses has not been
previously demonstrated. This study describes an optimised method for H5 and H7 subtype
IAV isolation and propagation in 28-day old embryonated ostrich eggs (EOEs), the biological
equivalent of 14-day old ECEs. The viability of EOEs transported from breeding sites could be
maximised by pre-incubating the eggs for 12 to 14 days prior to long-distance transportation.
This method applied to studies for ostrich-adapted virus isolation and in ovo studies will
enable better understanding of the virus-host interaction in ostriches and the emergence of
potentially zoonotic diseases.
