Susceptibility of Seasonal Influenza Viruses to Neuraminidase Inhibitors and Baloxavir in Qinghai Province in 2021–2025

ABSTRACT Background This study aimed to analyze whether seasonal influenza viruses circulating in Qinghai Province from 2021 to 2025 are susceptible to neuraminidase inhibitors (oseltamivir and zanamivir) and baloxavir. Methods Based on routine surveillance, 214 isolates were randomly selected for phenotypic neuraminidase inhibition testing against oseltamivir and zanamivir. The half-maximal inhibitory concentration was calculated and compared with subtype/lineage-specific median reference values. Neuraminidase genes of isolates showing reduced inhibition (RI) or highly reduced inhibition (HRI) were sequenced. Additionally, polymerase acidic gene sequencing used 141 randomly selected isolates to screen for known baloxavir resistance–associated substitutions. Results In total, 212 (99.07%) isolates were sensitive to oseltamivir; two B/Victoria (0.93%) showed RI. For zanamivir, 209 (97.66%) were sensitive; 4 (1.87%) showed RI (1 A[H1N1]pdm09, 3 B/Victoria), while 1 (0.47%) A(H1N1)pdm09 showed HRI. Among the seven isolates with abnormal neuraminidase inhibition testing results, only one B/Victoria (RI to zanamivir) carried a well-known resistance mutation (D197N). One A(H1N1)pdm09 (HRI to zanamivir) carried K331R, and one B/Victoria (RI to oseltamivir) carried Y214H. However, evidence for these two substitutions conferring resistance remains insufficient, requiring functional validation. The four remaining isolates had no amino acid changes in neuraminidase compared with the consensus sequences of contemporaneous sensitive isolates. Furthermore, no isolates harbored any known baloxavir resistance–associated mutations in the polymerase acidic gene. Conclusion Seasonal influenza viruses circulating in Qinghai in 2021–2025 remained largely susceptible to neuraminidase inhibitors, without known baloxavir resistance–associated substitutions detected by genotypic screening. Thus, continuous surveillance integrating phenotypic and genotypic methods is essential.