A gapA PCR-sequencing Assay for Identifying the Dickeya and Pectobacterium Potato Pathogens

Abstract

Several pectinolytic Pectobacterium and Dickeya species and subspecies are causative agents of blackleg and soft rot diseases on potato plants and tubers. Rapid and accurate identification of these taxa is a crucial issue for the production and international trade of potato seed-tubers. Here, we developed a PCR-sequencing tool to easily characterize the different Pectobacterium and Dickeya taxa. The gapA gene sequences from 53 published genomes were aligned and a phylogeny tree was constructed. A set of 35 signature nucleotides was discovered to distinguish the Pectobacterium and Dickeya genera, species, and subspecies. Then, a PCR-primer couple was designed for amplifying the gapA gene in pectinolytic enterobacteria. The primers were tested on 22 isolates recovered from blackleg symptoms in several potato fields. Amplicons were sequenced and signature-nucleotides were analyzed. A phylogeny that includes gapA sequence specimens confirmed the taxonomical identification of these environmental isolates.