Development of an Engineered Bioluminescent Reporter Phage for Detection of Bacterial Blight of Crucifers

Bacterial blight, caused by the phytopathogenPseudomonas cannabina pv.alisalensis , is an emerging disease afflicting important members of theBrassicaceae family. The disease is often misdiagnosed as pepper spot, a much less severe disease caused by the related pathogenPseudomonas syringae pv.maculicola . We have developed a phage-based diagnostic that can both identify and detect the causative agent of bacterial blight and differentiate the two pathogens. A recombinant “light”-tagged reporter phage was generated by integrating bacterialluxAB genes encoding luciferase into the genome ofP. cannabina pv.alisalensis phage PBSPCA1. The PBSPCA1::luxAB reporter phage is viable and stable and retains properties similar to those of the wild-type phage. PBSPCA1::luxAB rapidly and sensitively detectsP. cannabina pv.alisalensis by conferring a bioluminescent signal response to cultured cells. Detection is dependent on cell viability. Other bacterial pathogens ofBrassica species such asP. syringae pv.maculicola ,Pseudomonas marginalis ,Pectobacterium carotovorum ,Xanthomonas campestris pv.campestris , andX. campestris pv.raphani either do not produce a response or produce significantly attenuated signals with the reporter phage. Importantly, the reporter phage detectsP. cannabina pv.alisalensis on diseased plant specimens, indicating its potential for disease diagnosis.