**4. Biology of** *P. brassicae*

*brassicae* consists of two phases; in a primary phase, under favorable conditions, resting spores germinate and produce primary zoospores that penetrate in root hairs and mass production of secondary zoospores occurs in the root hairs. The resting spores are about 3 μm in size and subspherical to spherical in shape and the surface of each resting spore is covered with spines [21]. Mass of primary zoospores is released from each resting spore, spindle-shaped or pyriform, 2.8–5.9 μm long, and biflagellate. When the zoospores come in contact with the surface of a root hair, it penetrates in the cell wall and it is also called root hair infection. Secondary phase of life cycle occurs in the root cortex as a result secondary plasmodia and gall formation occurs as a result restriction in water and nutrient uptake by plants (Figure 1) [21, 22]. The life cycle study of *P. brassicae* in *A. thaliana* reported uninucleate and binucleate myxamoeboid structure production within host cytoplasm that caused cell wall burst and production of secondary plasmodia [23]. During pathogen infection, secondary plasmodia proliferate in roots and plant hormone, especially auxin and cytokinin, biosynthesis altered in the root tissues that causes gall formation (Figure 2) [21]. Infected plants become stunted, yellowish in color, and eventually wilt, which causes severe reduction in yield and quality of crops [24]. Mature secondary plasmodia subsequently develop into resting spores that can survive for 20 years or more [25]. Clubroot disease pressure can significantly increase in those fields where crop rotation frequently includes canola/rapeseed or other Brassica crops. Acidic soil with high soil moisture is the most favorable condition for resting spore germination and

**Figure 1.** Life cycle of *P. brassicae* and club like gall formation on the roots of Brassica host plant.

subsequent secondary infection.

6 Plants for the Future

The genus *Plasmodiophora* is a monophyletic group with uncertain systematic affinities. The species belonging to this genus possess unique features such as cruciform nuclear division, parasitism, obligate nature, biflagella, heterocont zoospores, and environmentally resistance and long-living resting spores [26]. In this genus, the economically significant member is *P. brassicae*, which hosts Brassica species to cause clubroot disease. The pathogen shows a wide biological range and its populations usually consist of a mixture of different pathotypes [27– 30]. Soil environmental factors such as physical, chemical, and biological properties of soil may differentially influence the survival of some physiological races of the pathogen [21, 31]. In European, field isolates of *P. brassicae* display great variation and show a tendency to overcome different resistance sources from either *B. rapa* or *B. oleracea*.

To enhance our understanding of the pathogenicity factors of *P. brassicae* causing clubroot disease on different Brassica hosts, several molecular techniques and tools are employed to determine *P. brassicae* genome size, structure, and number of possible functional genes in the whole genome. Several studies reported use of pulse-field gel electrophoresis (PFGE) to determine the karyotypes for *P. brassicae*. Ito et al. [32] used sheroplasts and differentiated 13 chromosomal bands in the range of 1.9 Mb to 750 kb. Bryan et al. [33] used isolated plasmodia and differentiated six chromosomal bands in the range of 1.7 Mb to 680 kb. Similarly, Graf et al. [34] distinguished 16 chromosomal bands in the range of 2.2 Mb to 680 kb. Based on these studies, it is estimated that the *P. brassicae* total genome size can be 18–20.3 Mb [35].

On the other hand, several molecular marker techniques were employed to investigate virulent pattern of the *P. brassicae* population derived from single-spore isolate or field isolates [36–38]; however, the number of distinguishing patterns were very low and that could not correlate with virulence patterns. In continuous efforts, two RAPD markers [39] and one SCAR marker [40] were identified, which correlate to isolates of pathotype 1. Yet there are no sets of molecular markers that can distinguish other pathotypes from field isolates which make clubrootresistance breeding intriguing.
