Avian influenza viruses (AIVs) continue to pose global threats mainly due to their rapid evolving nature and the dynamic range of susceptible hosts. Zoonotic highly pathogenic avian influenza virus (HPAIV) H5N1 of clade 2.2.1 has recently diversified into two distinct genetic subclades (18.104.22.168 and 22.214.171.124) in Egypt. This study was conducted as part of routine surveillance activities in Egypt; 5202 cloacal and oropharyngeal swabs were collected from live/dead birds, including commercial and backyard flocks, in five different locations in Egypt between the years of 2015 and 2016. All samples were screened using virus isolation, as well as antigenic and molecular detection methods. Full-length hemagglutinin (HA) sequences of six representative H5N1 isolates were analysed to study their genetic evolution followed by estimation of their evolutionary rates among different virus clusters. This analysis revealed a high evolution rate for clade 126.96.36.199. Additionally, analysis of selection pressures in the HA gene revealed a positive selection pressure. Deduced amino acid analysis revealed characteristic mutations at HA antigenic sites besides two other mutations (129∆, I151T) that were found to be stable in recent subclade 188.8.131.52 isolates from humans and chickens. Our results revealed that linking the epidemiological and sequence data is important to understand the prevalence, transmission, persistence and evolution of the virus, and to monitor the circulating AIV strains and emergence of new AIV subtypes in Egypt. The wide circulation of the 184.108.40.206 subclade, carrying triple mutations (120, 129∆, I151T) associated with increased binding affinity to human receptors, is an alarming finding with public health importance.
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