Background <p>HA and NA are important surface proteins of influenza viruses. The purpose of this study was to explore the molecular evolution and amino acid mutations of HA and NA of H1N1, H3N2 and Influenza B Victoria lineage (B/Victoria) viruses in Qingyang from 2020 to 2023.</p> Methods <p>Through the China Influenza Surveillance Information System, the positive data of influenza etiology surveillance in Qingyang from 2020 to 2023 were screened. A total of 19 samples of H1N1, 47 samples of H3N2 and 51 samples of B/Victoria were screened. Subsequently, virus nucleic acid extraction, whole genome sequencing, amino acid and nucleotide homology analysis, phylogenetic analysis, and amino acid site mutation analysis were performed.</p> Results <p>Miniseq sequencing was performed on 117 influenza strains. Homology analysis showed that H1N1 and H3N2 strains had relatively low homology with vaccine representative strains. Amino acid mutation analysis of HA and NA showed that the H1N1 strains had receptor binding site and antigen site mutations, but no glycosylation site and resistance site mutations. However, H3N2 strains had mutations at receptor binding sites, antigenic sites, glycosylation sites, and resistance sites. The B/Victoria strains had antigenic site mutations, but no drug resistance site mutations. Moreover, only strains from 2021 to 2022 exhibit a glycosylation site mutation.</p> Conclusion <p>Genetic characteristics analysis was conducted for the first time on H1N1, H3N2, and B/Victoria viruses prevalent in Qingyang, which enables us to understand the molecular evolution and amino acid mutations of HA and NA.</p> Clinical trial number <p>Not applicable.</p>