Live Cell Imaging of Plasmodiophora brassicae Infection and Host Interactions
Plasmodiophora brassicae Woronin, is a soil-borne obligate biotrophic plant pathogen responsible for clubroot, one of the most devastating diseases of Brassicaceae. Previous studies into the lifecycle of P. brassicae have focused on fixed tissue samples for histological or transmission electron microscopic investigations due to the lack of a pathogen-specific stain for live cell/tissue study. Using the fluorophore Nile red to stain lipid droplets in all life stages of P. brassicae allows for live cell microscopic investigation of this pathogen. Nile red can be used to label P. brassicae ex planta in the soil during the resting spore and infective zoospore phases, and in planta during its obligate life cycle phases. This Nile red labelling technique combined with transgenic Arabidopsis thaliana plants expressing fluorescent protein-labelled organelle markers permits imaging of the zoospore penetration of the host cell wall, subsequent pathogen development within the intracellular environment, and further allows the investigation of pathogen-induced organelle recruitment and/or disruption during the P. brassicae-A. thaliana interaction. Specifically, the translocation of the nucleus to the penetration site and its subsequent envelopment by plasmodia, and the establishment of a cytoplasmic vacuole-derived encasement termed the parasitophorus vacuole. This staining technique has provided insight pertaining to the cellular interactions of P. brassicae and its hosts.
Plasmodiophora brassicae, Clubroot, Arabidopsis, Brassicae, Confocal, Nile red
Master of Science (M.Sc.)