Multiplex real-time PCR for the detection of Clavibacter michiganensis subsp. michiganensis, Pseudomonas syringae pv. tomato and pathogenic Xanthomonas species on tomato plants

Abstract

A multiplex real-time PCR method based on fluorescent TaqMan® probes was developed for the simultaneous detection of the tomato pathogenic bacteria Clavibacter michiganensis subsp. michiganensis, Pseudomonas syringae pv. tomato and bacterial spot-causing xanthomonads. The specificity of the multiplex assay was validated on 44 bacterial strains, including 32 target pathogen strains as well as closely related species and nontarget tomato pathogenic bacteria. The designed multiplex real-time PCR showed high sensitivity when positive amplification was observed for one pg of bacterial DNA in the cases of Clavibacter michiganensis subsp. michiganensis and Pseudomonas syringae pv. tomato bacteria and 100 pg for bacterial spot-causing xanthomonads. The reliability of the developed multiplex real-time PCR assay for in planta detection was verified by recognition of the target pathogens in 18 tomato plants artificially inoculated by each of the target bacteria and tomato samples from production greenhouses.

Xanthomonas hortorum – beyond gardens: Current taxonomy, genomics, and virulence repertoires

“…michiganensis and Pseudomonas syringae pv. tomato , has been recently developed by targeting lepA (Peňázová et al, 2020).…”

Antibacterial activity of electrolyzed water on Pseudomonas syringae and Clavibactermichiganensis and its effect on seed germination

“…Diseases affecting this crop include bacterial freckling caused by Pseudomonas syringae pv. tomato (PEŇÁZOVÁ et al, 2020) and bacterial cancer caused by Clavibacter michiganensis subsp. michiganensis (NANDI et al, 2018).…”

“…This disease causes necrotic lesions surrounded by chlorotic halos on leaves, stems and fruits. The disease is spread by contaminated tomato seeds and infected weeds in which the bacteria can survive within the root system (PEŇÁZOVÁ et al, 2020).…”

“…This is considered one of the main bacterial pathogens in tomato production worldwide. It enters the plant through stomata and spreads within the xylem vessels where it colonizes the entire plant and forms extensive biofilm-like structures (PEŇÁZOVÁ et al, 2020). This interferes with water transport thus leading to wilting during the early stages of infection (NANDI et al, 2018).…”

“…This interferes with water transport thus leading to wilting during the early stages of infection (NANDI et al, 2018). Symptoms of the disease appear on aerial parts and include wilting of leaves and discoloration of vascular tissues (PEŇÁZOVÁ et al, 2020). If an infection occurs at a late stage of the plant's growth, the plants may survive and generate fruits that may show black spots with white halos, that are often referred to as "bird's-eye" spots (PEŇÁZOVÁ et al, 2020).…”