Plasmodiophora brassicae
, a parasitic protist, induces club‐shaped tumor‐like growth of host Brassicas roots and hypocotyls after infection. Due to its soil‐borne nature and intracellular, biotrophic parasitism the infection biology and early pathogenesis remains in doubt. In this study, we have established a new protocol, based on a two‐step axenic culture of
P. brassicae
with its host tissues, for easy and
in planta
observation of cellular interactions between
P. brassicae
and host plants: first, coculture of
P. brassicae
with infected canola root tissues, on growth‐medium plates, enables the propagation of
P. brassicae
that serves as pure inoculum for pathogenicity assays, and second, the pure inoculum is subsequently used for pathogenicity tests on both canola and
Arabidopsis
seedlings grown on medium plates in Petri dishes. During the first axenic culture, we established a staining protocol by which the pathogen was fluorescently labeled with Nile red and calcofluor white, thus allowing
in planta
observation of pathogen development. In the pathogenicity assays, our results showed that axenic cultures of
P. brassicae
, in calli, remains fully virulent and completes its life cycle in both canola and
Arabidopsis
roots grown in Petri dishes. Combining visualization of fluorescent probe‐labeled
P. brassicae
structures with fluorescent protein tagging of
Arabidopsis
cellular components, further revealed dynamic responses of host cells at the early stages of
P. brassicae
infection. Thus, established protocols for
in planta
detection of
P. brassicae
structures and the live cell imaging of
P. brassicae
—
Arabidopsis
interactions provide a novel strategy for improving our detailed knowledge of
P. brassicae
infection in host tissues.