Genomic and biological characterization of a novel bacteriophage X1 infecting Xanthomonas campestris pv. campestris with biocontrol potential against cabbage black rot.

Bacteriophages have emerged as promising alternatives to pesticides for controlling bacterial pathogens in crops. Black rot, caused by Xanthomonas campestris pv. campestris (Xcc), is one of the most destructive bacterial diseases affecting cruciferous crops worldwide. In this study, a novel lytic bacteriophage, X1, infecting Xcc 8004, was isolated from sediment collected in an oligotrophic karst cave. Phage X1 formed clear plaques and exhibited myovirus-like morphology with an icosahedral head and a contractile tail. Biological characterization revealed productive infection of the tested Xcc strains but no detectable lytic activity against the non-Xcc strains included in the limited host panel. Phage X1 showed rapid adsorption to Xcc 8004 (> 99% within 10 min), a latent period of approximately 60 min, and a burst size of approximately 123 plaque-forming units per infected cell. Under laboratory conditions, the phage exhibited stable after incubation at 4-50 °C, pH 5-9, chloroform exposure, and the tested UV-A/UV-C irradiation regimes. Genome sequencing revealed that phage X1 harbors a large linear double-stranded DNA genome of 200,058 bp, encoding 311 predicted open reading frames and 34 tRNA genes. DRAM-v analysis identified 28 candidate auxiliary metabolic genes (AMGs), potentially associated with host metabolic regulation and environmental adaptation. No recognizable antibiotic resistance, virulence, integrase, or lysogeny-associated genes were identified in its genome. Comparative genomic and phylogenetic analyses indicated that X1 is most closely related to Xanthomonas phage BUDD within the class Caudoviricetes, but represents a distinct species-level genome based on intergenomic similarity. In controlled pot assays, preventive and therapeutic applications of X1 reduced lesion lengths by 88.27 and 73.25%, respectively, compared with Xcc-only treatment. Furthermore, preventive application treatment also reduced culturable Xcc populations to approximately one-fourth of the level observed after therapeutic treatment. These findings suggest that phage X1 has promising potential for further development as a phage-based biocontrol agent against cabbage black rot.