Targeted and Effective Phage-Based Biocontrol of Black Rot Disease in Broccoli

Xanthomonas species are Gram-negative bacterial pathogens responsible for diseases in over 400 plant hosts, including numerous economically important crops such as Brassica species. The limited efficacy and environmental concerns associated with chemical control strategies underscore the need for sustainable and targeted alternatives. In this study, we evaluated the suitability and biocontrol efficacy of phages Phi1 and Phi3 to combat Xanthomonas campestris pv. campestris (Xcc) in broccoli plants. Kill-curve assays demonstrated that both phages effectively suppressed Xcc growth across a range of multiplicities of infection. Transmission electron microscopy further confirmed their lytic activity, revealing pronounced structural damage to Xcc cells following phage treatment, accompanied by the subsequent release of phage progeny. To assess host specificity and biosafety, the phages were tested against 41 bacterial isolates that were isolated and taxonomically characterized from broccoli and cauliflower in this study. Neither Phi1 nor Phi3 exhibited lytic activity against any non-target isolate, indicating high host specificity and minimal risk to the native Brassica-associated microbiota. In planta assays demonstrated that the combined application of Phi1 and Phi3 reduced Xcc-induced symptom severity in broccoli plants by 80%. Collectively, these results demonstrate that phages Phi1 and Phi3 represent effective and biologically precise agents for the control of black rot disease in Brassica crops.