Background: Influenza B virus (IBV) contributes to seasonal epidemics, but its molecular evolution is less defined than influenza A. We analyzed IBVs collected in Japan and Myanmar (2016-2020) to investigate lineage dynamics, reassortment, and genetic mismatch with vaccine strains.

Methods: Respiratory specimens from patients with influenza-like illness were collected in Japan (January 2016-March 2020) and Myanmar (June 2016-October 2019). Lineages were determined by RT-PCR, isolates cultured in MDCK-based cells, and whole-genome sequencing performed on the Illumina iSeq 100. Phylogenetic analyses of all eight segments and hemagglutinin (HA) comparisons with WHO-recommended vaccine strains were conducted.

Results: Of 4703 specimens, 1164 were IBV-positive: 632 B/Victoria (322 Japan and 310 Myanmar) and 532 B/Yamagata (452 Japan and 80 Myanmar). Whole-genome sequences were obtained for 148 isolates. Despite differing seasonality, both countries showed parallel transitions: B/Yamagata predominated in 2017-2018 but disappeared after 2019, while B/Victoria shifted from clade V1A to emerging clade V1A.3 with a three-amino acid deletion in HA (Δ162-164). Two 2018 B/Victoria reassortants in Japan contained four internal genes from B/Yamagata. Multiple HA substitutions were observed among V1A.3 viruses, differing from contemporaneous vaccine strains (clade V1A.1), indicating potential antigenic mismatch. Mutations related to antiviral resistance in polymerase acidic and neuraminidase genes were also assessed.

Conclusions: This study reveals synchronized lineage shifts, inter-lineage reassortment, and vaccine mismatches of IBVs in two distinct countries. The disappearance of B/Yamagata and emergence of divergent B/Victoria clades highlight the need for sustained molecular surveillance to guide vaccine strain selection.