First Report of Xanthomonas campestris pv. campestris Causing Black Rot on Cauliflower in Ecuador

In February 2024, cauliflower plants (Brassica oleracea var. botrytis) showing symptoms of black rot were observed at the experimental farm of the Faculty of Agricultural Sciences, Central University of Ecuador, in Quito, Ecuador. Affected plants exhibited characteristic V-shaped chlorotic lesions on leaves. Disease incidence was approximately 15% across the field. Symptomatic leaf tissue (5 mm² fragments) was surface-sterilized with 0.5% sodium hypochlorite, rinsed with sterile distilled water, and plated on nutrient agar. After 48 h at 28°C, yellow, mucoid, convex colonies with defined borders were observed. Two colonies were identified as Xanthomonas campestris using the Biolog system with 94 phenotypic tests. Genomic DNA from strain XccEC_1 was extracted using the CTAB protocol (Doyle and Doyle 1987) and sequenced on an Illumina NovaSeq platform (PE150, 100× coverage) at Biomarker Technologies (Hong Kong). The genome assembly (SPAdes v3.15; Bankevich et al. 2012) yielded 5,147,842 bp in 230 contigs with an N50 of 75,866 bp and 64.87% GC content. Phylogenomic analysis based on 100 conserved orthologous genes was performed using the BV-BRC codon tree pipeline (Olson et al. 2023). Protein sequences were aligned using MAFFT, and a maximum-likelihood tree was inferred using RAxML v8.2.12 under the GTRCAT model with 100 rapid bootstrap replicates. The resulting tree grouped XccEC_1 with X. campestris pv. campestris str. ATCC 33913 (100% bootstrap support) (Fig. S1). The complete genome of strain XccEC_1 was deposited in GenBank (accession no. JBUBWA000000000). For pathogenicity tests, five healthy five-week-old cauliflower seedlings were infiltrated on four leaves each with a bacterial suspension of XccEC_1 (10⁸ CFU/ml in PBS) using a needleless syringe, and one control plant was infiltrated with sterile distilled water. Plants were incubated at 25°C and 68% relative humidity. At 7 days post-inoculation, all inoculated plants developed chlorotic to necrotic foliar lesions, and by 12 days, vascular tissue blackening was evident (Fig. S2). Control plants remained asymptomatic, and no suspected X. campestris colonies were isolated from these plants. The bacterium was reisolated from symptomatic plants, and species identity was confirmed by 16S rRNA and GyrB sequencing (GenBank accession no. PX498286, PZ417655). The pathogenicity assay was repeated once with identical results. The XccEC_1 isolate was stored with code MB_59 at the Pontificia Universidad Católica del Ecuador Culture Collection. To our knowledge, this is the first report of X. campestris pv. campestris causing black rot on cauliflower in Ecuador. This pathogen poses a significant threat to Brassica production in the region (Vicente and Holub 2013) and warrants implementation of integrated disease management strategies.