Phylogeographic analysis of H5N1 highly pathogenic avian influenza virus isolated in Cambodia from 2018 to 2019

Since 2004, several outbreaks of highly pathogenic avian influenza (HPAI) have been reported in Cambodia. Until 2013, all H5N1 viruses identified in Cambodia belonged to clade 1 and its subclades. H5N1 HPAI viruses belonging to clade 2.3.2.1c have been dominant since the beginning of 2014, with various genotypes (KH1-KH5) reported. Here, we isolated nine H5N1 HPAI viruses from domestic poultry farms and slaughterhouses in Cambodia during 2018-2019 and performed phylogenetic analysis of whole genome sequences. All isolates were classified as H5 clade 2.3.2.1c viruses and all harbored multi-basic amino acid sequences (PQRERRRKR/GLF) at the haemagglutinin (HA) cleavage site. Phylogenetic analysis revealed that the H5N1 isolates in this study belonged to the KH2 genotype, the dominant genotype in Cambodia in 2015. Phylogenetic analysis of the HA gene showed that the isolates were divided into two groups (A and B). The results of Bayesian discrete phylogeography analysis revealed that the viral migration pathways from Vietnam to Cambodia (Bayes factor value: 734,039.01; posterior probability: 1.00) and from Cambodia to Vietnam (Bayes factor value: 26,199.95; posterior probability: 1.00) were supported by high statistical values. These well-supported viral migrations between Vietnam and Cambodia demonstrate that viral transmission continued in both directions. Several factors may have contributed to this, including the free-grazing duck system and movement of poultry-related products. Thus, the results emphasize the need for an enhanced international surveillance program to better understand transboundary infection and evolution of H5N1 HPAI viruses, along with implementation of more stringent international trade controls on poultry and poultry products.