biostudies-literatureKim SWselection of research-oriented professor of Jeonbuk National UniversityKorea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheriesresearch funds for newly appointed professors of Jeonbuk National University127https://www.ebi.ac.uk/biostudies/studies/S-EPMC12944848biostudies-literatureUnknown13(2)The widespread panzootic of clade 2.3.4.4b highly pathogenic avian influenza (HPAI) H5N1 necessitates the development of vaccine platforms capable of rapid adaptation to emerging antigenic variants. Although commercial recombinant turkey herpesvirus (rHVT) vaccines are available, they often utilize heterologous inserts that may fail to optimally limit viral shedding of novel field strains. Here, we report the rapid construction of a homologous rHVT-H5 vaccine expressing the hemagglutinin (HA) gene of a representative clade 2.3.4.4b isolate via CRISPR/Cas9-mediated non-homologous end joining (NHEJ). In vitro characterization confirmed stable HA surface expression and growth kinetics comparable to the parental virus. In specific-pathogen-free (SPF) chickens, rHVT-H5 elicited robust hemagglutVeterinary sciencesAntigenic Matching of rHVT-H5 via CRISPR/Cas9 Confers Complete Protection Against Novel H5N1 Clade 2.3.4.4b in Chicken.PMC129448482025RS-2025-023070592022RS-2024-00397877Kim SWJang HKJeon WBWei BPark JYKim KWSon JEKang MYu CDChoi YRfalseAntigenic Matching of rHVT-H5 via CRISPR/Cas9 Confers Complete Protection Against Novel H5N1 Clade 2.3.4.4b in Chicken.The widespread panzootic of clade 2.3.4.4b highly pathogenic avian influenza (HPAI) H5N1 necessitates the development of vaccine platforms capable of rapid adaptation to emerging antigenic variants. Although commercial recombinant turkey herpesvirus (rHVT) vaccines are available, they often utilize heterologous inserts that may fail to optimally limit viral shedding of novel field strains. Here, we report the rapid construction of a homologous rHVT-H5 vaccine expressing the hemagglutinin (HA) gene of a representative clade 2.3.4.4b isolate via CRISPR/Cas9-mediated non-homologous end joining (NHEJ). In vitro characterization confirmed stable HA surface expression and growth kinetics comparable to the parental virus. In specific-pathogen-free (SPF) chickens, rHVT-H5 elicited robust hemagglut2026-01-01T00:00:00Z2026 Jan2026-07-11T03:18:37.769Z2026-07-11T03:09:42.744ZS-EPMC129448484174592110.3390/vetsci13020127