Characterization of a new lytic bacteriophage vB_RanS_GDF21 and its endolysin LysGDF21 with antimicrobial activity against Riemerella anatipestifer

Introduction Riemerella anatipestifer ( R. anatipestifer ) presents as a major pathogen causing septicemia among poultry. The complex serotype diversity and multidrug resistance of R. anatipestifer severely compromise infection prevention and treatment strategies. Bacteriophages and their endolysins represent a promising alternative strategy against R. anatipestifer . Methods The phage vB_RanS_GDF21 (GDF21) was isolated from feces of a duck farm, and its biological properties were characterized. Whole-genome sequencing was performed using the Illumina NovaSeq 6000 platform, followed by a bioinformatic analysis of the genome. The derived endolysin, LysGDF21 (encoded by orf 65 ), was expressed in Escherichia coli BL21 (DE3), and its anti-biofilm and antimicrobial activities were assessed. Results Transmission electron microscopy showed that phage GDF21 had an icosahedral symmetric head, and a long, non-contractile tail. It exhibited a 20 min eclipse period, burst size of 74 PFU/infected cell, and stability at pH 5-11 and temperatures below 50 °C. Whole-genome sequencing revealed that GDF21 possesses a double-stranded DNA genome with length of 46,925 bp and G+C content of 35.25%. Among the 75 predicted ORFs, 27 were functionally annotated and no genes associated with virulence or antibiotic resistance were identified. Phylogeny and classification analysis indicated that GDF21 is a member of the class Caudoviricetes and is closely related to Riemerella phage vB_RanS_CRP19. LysGDF21 significantly inhibited biofilm formation and disrupted mature biofilms in R. anatipestifer . When combined with EDTA, LysGDF21 exhibited broad antibacterial activity against R. anatipestifer . Conclusion The characterization of phage GDF21 enriches the phage database, while the potent anti-biofilm and antibacterial activities of LysGDF21 highlight its therapeutic potential against R. anatipestifer infections.