Abstract
During the epizootic of highly pathogenic avian influenza (HPAI) H5N8 virus in Europe in 2016-2017, HPAI viruses of subtype H5N5 were also isolated. However, the detection of H5N5 viruses was limited compared to H5N8. In this study, we show that the genetic constellation of a newly isolated H5N5 virus is different from two genotypes previously identified in the Netherlands. The introduction and spread of the three H5N5 genotypes in Europe was studied using spatiotemporal and genetic analysis. This demonstrated that the genotypes were isolated in distinguishable phases of the epizootic, and suggested multiple introductions of H5N5 viruses into Europe followed by local spread. We estimated the timing of the reassortment events, which suggested that the genotypes emerged after the start of autumn migration. This may have prevented large-scale spread of the H5N5 viruses on wild bird breeding sites before introduction into Europe. Experiments in primary chicken and duck cells revealed only minor differences in cytopathogenicity and replication kinetics between H5N5 genotypes and H5N8. These results suggest that the limited spread of HPAI H5N5 viruses is related to the timing of the reassortment events rather than changes in virus pathogenicity or replication kinetics.