Genetic Characterization of Kazakhstan Isolates: Avian Influenza H9N2 Viruses Demonstrate Their Potential to Infect Mammals

Abstract

Low pathogenic H9N2 avian influenza viruses have become widespread in wild birds and poultry worldwide, raising concerns about their potential to spark pandemics or their role in enhancing the virulence and infectivity of H5Nx viruses through genetic reassortment. Therefore, influenza monitoring studies, including those of H9N2 viruses, are crucial for understanding, evaluating, and mitigating the risks associated with avian infections, and have broader implications for global public health. Although H9N2 viruses are not considered enzootic in Kazakhstan, they have been repeatedly detected in wild waterfowls and domestic poultry. In this study, all eight gene segments of influenza A/H9N2 viruses isolated in various regions of Kazakhstan between 2014 and 2020 were sequenced and analyzed. Molecular characterization revealed the presence of genetic markers associated with mammalian infectivity and disease potential. Furthermore, their predicted receptor binding site sequences indicate their potential capacity to attach to human-type receptors. These findings highlight the importance of continued surveillance and molecular investigation to better understand the evolution and zoonotic potential of H9N2 viruses in Kazakhstan.

Keywords: H9N2 subtype; adaptation; bird; genome; hemagglutinin; influenza A virus; phylogenesis; transmission; variability.

Figure 1

Map of sampling sites in this study. The two regions where samples were collected are indicated by a red dash (the map was modified from [24]). The blue circles indicate the locations of lakes where H9N2 strains were isolated from waterfowl in the North Kazakhstan and Almaty regions. The red triangle indicates the location of the poultry farm in the Almaty region.

Figure 2

Phylogenetic tree of the full-length HA sequence of the H9N2 isolates from Kazakhstan, including A/Chicken/Almaty/220/2020 (marked with red triangle) and wild bird’s isolates (marked with blue dots) and other related sequences from wild and domestic birds (GenBank and GISAID databases). Numbers at nodes indicate maximum likelihood bootstrap values of 1000 replicates under the specified model. Only the bootstrap values above 70 are shown. The tree is rooted to A/turkey/Wisconsin/1966. Bar, 0.05 nt substitutions per site.

Figure 3

Phylogenetic tree of the full-length NA sequence of the H9N2 isolates from Kazakhstan, including A/Chicken/Almaty/220/2020 (marked with red triangle) and wild birds isolates (marked with blue dots) and other related sequences from wild and domestic birds (GenBank and GISAID databases). Numbers at nodes indicate maximum likelihood bootstrap values of 1000 replicates under the specified model. Only the bootstrap values above 70 are shown. The tree is rooted to A/turkey/Wisconsin/1966. Bar, 0.02 nt substitutions per site.

Figure 4

Alignment of HA aa sequences of four of Kazakhstan’s IAVs and their consensus sequence in comparison with the reference viruses’ IAVs and their consensus sequence. Cleavage and glycosylation sites are indicated in blue and red boxes, respectively.