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SUMMARY Plasmodiophora brassicae, a soil‐borne biotroph, establishes galls as strong physiological sinks on Brassicaceae plants including Brassica napus and Arabidopsis thaliana. We compare transcriptional profiles of phloem dissected from leaf petioles and hypocotyls of healthy and infected B. napus plants. Our results highlight how pathogenesis accompanies phloem‐mediated defence responses whilst exerting a strong influence on carbon–nitrogen (C–N) economy. We observe transcriptional changes indicating decreased aliphatic glucosinolate biosynthesis, fluctuating jasmonic acid responses, altered amino acid (AA) and nitrate transport, carbohydrate metabolism and modified cytokinin responses. Changes observed in phloem‐dissected from upper versus lower plant organs point to phloem as a conduit in mediating C–N repartitioning, nutrition‐related signalling and cytokinin dynamics over long distances during clubroot disease. To assess changes in physiology, we measured AAs, sugars and cytokinins, in phloem exudates from B. napus plants. Despite the decrease in most AA and sucrose levels, isopentyl‐type cytokinins increased within infected plants. Furthermore, we employed Arabidopsis for visualising promoter activities of B. napus AA and N transporter orthologues and tested the impact of disrupted cytokinin transport during P. brassicae‐induced gall formation using Atabcg14 mutants. Our physiological and microscopy studies show that the host developmental reaction to P. brassicae relies on cytokinin and is accompanied by intense nitrogen and carbon repartitioning. Overall, our work highlights the systemic aspects of host responses that should be taken into account when studying clubroot disease. [ABSTRACT FROM AUTHOR]

The effect of crop rotation on soil-borne diseases is a representative case of plant–soil feedback in the sense that plant disease resistance is influenced by soils with different cultivation histories. This study examined the microbial mechanisms inducing the differences in the clubroot (caused by Plasmodiophora brassicae pathogen) damage of Chinese cabbage (Brassica rapa subsp. pekinensis) after the cultivation of different preceding crops. It addresses two key questions in crop rotation: changes in the soil bacterial community induced by the cultivation of different plants and the microbial mechanisms responsible for the disease-suppressive capacity of Chinese cabbage. Twenty preceding crops from different plant families showed significant differences in the disease damage, pathogen density, and bacterial community composition of the host plant. Structural equation modelling revealed that the relative abundance of four key bacterial orders in Chinese cabbage roots can explain 85% and 70% of the total variation in pathogen density and disease damage, respectively. Notably, the relative dominance of Bacillales and Rhizobiales, which have a trade-off relationship, exhibited predominant effects on pathogen density and disease damage. The disease-suppressive soil legacy effects of preceding crops are reflected in compositional changes in key bacterial orders, which are intensified by the bacterial community network. [ABSTRACT FROM AUTHOR]

Abstract Aims: Straw turnover plays an important role in reducing soil diseases, improving the ecological environment of plowland and realizing the effective ecological utilization of straw. Methods: Pot and field experiments were carried out to investigate the effects of maize, rice and wheat straws on the growth, clubroot disease of Chinese cabbage and soil nutrients. Undecomposed and decomposed maize, rice and wheat straws were quantitatively added to the monocultural soil of Chinese cabbage, and the crops without straw were taken as the control. Results: The results showed that the addition of maize, wheat and rice straws could promote the growth of monocultural Chinese cabbage, inhibit the occurrence of clubroot disease, increase soil pH value, the content of soil organic matter, alkaline hydrolyzable nitrogen and available potassium in pot experiment. Exogenous straw application could reduce the incidence rate by 22.54 ~ 47.85%, increase the plot yield of field 95.15 ~ 365.81%. Conclusions: In terms of inhibiting clubroot disease and improving soil properties, undecomposed rice straw is superior to maize and wheat straw, while decomposed maize straw is superior to rice and wheat straw.

Clubroot, a significant soil-borne disease, severely impacts the productivity of cruciferous crops. The identification and development of clubroot resistance (CR) genes are crucial for mitigating this disease. This study investigated the genetic inheritance of clubroot resistance within an F2 progeny derived from the cross of a resistant parent, designated "377", and a susceptible parent, designated "12A". Notably, "377" exhibited robust resistance to the "KEL-23" strain of Plasmodiophora brassicae, the causative agent of clubroot. Genetic analyses suggested that the observed resistance is controlled by a single dominant gene. Through Bulked Segregant Analysis sequencing (BSA-seq) and preliminary gene mapping, we localized the CR gene locus, designated as BraPb8.3, to a 1.30 Mb genomic segment on chromosome A08, flanked by the markers "333" and "sau332-1". Further fine mapping precisely narrowed down the position of BraPb8.3 to a 173.8 kb region between the markers "srt8-65" and "srt8-25", where we identified 22 genes, including Bra020861 with a TIR-NBS-LRR domain and Bra020876 with an LRR domain. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analyses confirmed that both Bra020861 and Bra020876 exhibit increased expression levels in the resistant parent "377" following inoculation with P. brassicae, thereby underscoring their potential as key genes implicated in BraPb8.3-mediated clubroot resistance. This study not only identifies molecular markers associated with BraPb8.3 but also enriches the genetic resources available for breeding programs aimed at enhancing resistance to clubroot. [ABSTRACT FROM AUTHOR]

Introduction: Clubroot disease is a devastating soil borne disease caused by infection with Plasmodiophora brassicae, which primarily affects cruciferous plants. The microbial diversity of the soil is an essential indicator of its quality. Methods: This study measured the physicochemical properties of the soil to study the effect of its microbial diversity on the infection of oilseed rape with P. brassicae. High-throughput sequences of the soil bacteria and fungi in the interroot soils of P. brassicae were analyzed under different treatment conditions. Results: In the study, it was found that the efficiency of strain X216 in preventing and controlling the root disease of rapeseed was positively correlated with the amount of solution used to irrigate the root system. The results of the greenhouse and field trials showed that the efficiency of strain X216 against the root disease of rapeseed was 43.16% in the field and 62.14% in the greenhouse. Proteobacteria, Chloroflexi, Rozellomycota, and Basidiomycota are critical phylum in the development of clubroot disease. The application of biocontrol increased the relative abundance of Actinobacteria, Bacillus, Mesorhizobium, Mycobacterium, Streptomyces and Filobasidium, which affected the structure and abundance of microbial communities. A principal coordinate analysis showed that the microbial structure in the soil varied substantially in the bacterial community, and there was no significant difference in soil structure in the fungal community. Discussion: The occurrence of clubroot disease affected the structure of interroot microbial community composition in the soil, which resulted in a decrease in its community diversity. The application of the biocontrol bacterium X216 increased the soil microbial diversity. It effectively reduced the occurrence of P. brassicae, and this study provides a basis to study the microbial diversity in cruciferous crops. [ABSTRACT FROM AUTHOR]

Infection by the mustard clubroot disease pathogen Plasmodiophora brassicae has a significant negative impact on the quality and yield of Chinese mustard (Brassica juncea). At present, screening resistant resources for breeding programs is the most economical and effective method available to control this disease. In this study, we isolated P. brassicae physiological race 4 from Chinese cabbage and examined 483 mustard germplasm resources (193 leaf mustard, 96 stem mustard, and 194 root mustard) from China and abroad to identify resistance to clubroot disease at the seedling stage through irrigation inoculation with the isolated pathogen. The results showed that there were no immune varieties among the tested mustard germplasm, but that there were differences in resistance to clubroot disease among the three mustard types. More than 90% of leaf and stem mustard resources were susceptible to clubroot disease, whereas 38.66% of root mustard resources showed resistance. In total, we detected 4 highly resistant, 9 resistant, and 83 moderately resistant varieties, of which 4 highly resistant, 8 resistant, and 63 moderately resistant varieties were root mustard resources, whereas only 1 resistant and 5 moderately resistant varieties were stem mustard resources, and 15 moderately resistant varieties were leaf mustard resources. In addition, we used seven molecular markers for clubroot disease resistance in Chinese cabbage to detect stem and root mustard resources. The results showed that the marker CRk was detected in 97.87% of stem mustard and 92.49% of root mustard resources. Six markers (Crr1, Crr2, Crr3, CRa, CRb, and CRc) were detected in 18.09%, 7.45%, 2.13%, 6.38%, 12.77%, and 12.77% of stem mustard germplasms, and four markers (Crr1, Crr2, Crr3, and CRc) were detected in 8.09%, 8.67%, 10.40%, and 8.67% of root mustard germplasms, respectively, suggesting that these markers are not suitable for detecting mustard germplasm resistance to clubroot disease. This study provides a technical reference and material support for the breeding of mustard varieties resistant to clubroot disease. [ABSTRACT FROM AUTHOR]

Clubroot, a soil-borne disease caused by Plasmodiophora brassicae, is one of the most destructive diseases of Brassica oleracea all over the world. However, the mechanism of clubroot resistance remains unclear. In this research, transcriptome sequencing was conducted on root samples from both resistant (R) and susceptible (S) B. oleracea plants infected by P. brassicae. Then the comparative analysis was carried out between the R and S samples at different time points during the infection stages to reveal clubroot resistance related pathways and candidate genes. Compared with 0 days after inoculation, a total of 4991 differential expressed genes were detected from the S pool, while only 2133 were found from the R pool. Gene function enrichment analysis found that the effector-triggered immunity played a major role in the R pool, while the pathogen-associated molecular pattern triggered immune response was stronger in the S pool. Simultaneously, candidate genes were identified through weighted gene co-expression network analysis, with Bol010786 (CNGC13) and Bol017921 (SD2-5) showing potential for conferring resistance to clubroot. The findings of this research provide valuable insights into the molecular mechanisms underlying clubroot resistance and present new avenues for further research aimed at enhancing the clubroot resistance of B. oleracea through breeding. [ABSTRACT FROM AUTHOR]

The priority in oilseed rape (Brassica napus L.) research and breeding programs worldwide is to combine different features to develop cultivars tailored to specific applications of this crop. In this study, forms with a modified fatty acid composition of seed oil were successfully combined with a source of resistance to Plasmodiophora brassicae Wor., a harmful protist-causing clubroot. Three HO-type recombinants in F6–F12 generations with oleic acid content of 80.2–82.1% and one HOLL-type F6 inbred mutant recombinant (HOmut × LLmut), with a high oleic acid content (80.9%) and reduced linolenic acid content (2.3%), were crossed with the cultivar Tosca, resistant to several pathotypes of P. brassicae. The work involved genotyping with the use of DNA markers specific for allelic variants of desaturase genes responsible for the synthesis of oleic and linolenic fatty acids, CAPS (FAD2 desaturase, C18:1), and SNaPshot (FAD3 desaturase, C18:3), respectively. Of 350 progenies in the F3 generation, 192 (55%) were selected for further studies. Among them, 80 HO (≥ 72%) lines were identified, 10 of which showed resistance to at least one up to four P. brassicae pathotypes. Thirty lines in the selected progeny contained high oleic acid and less than 5% linolenic acid; eight of them belonged to the HOLL type conferring resistance to at least one pathotype. Two HO lines and two HOLL lines were resistant to four pathotypes. The resulting HO-CR and HOLL-CR inbred lines with altered seed oil fatty acid composition and resistance to P. brassicae represent unique oilseed rape material with the desired combination of valuable traits. [ABSTRACT FROM AUTHOR]

Plasmodiophora brassicae is a serious threat to Brassica crops worldwide, resulting in substantial economic losses for growers. Biocontrol with chitinolytic bacteria producing chitinase is gaining attention as a natural alternative to chemicals. This approach is favored due to the essential role chitinases play in protecting against chitin-containing pathogens. Given that chitin is a major component in the resting spores of P. brassicae and plays a crucial role during pathogenesis, it is probable that Paenibacillus chitinolyticus, producing a high level of chitinase, could suppress P. brassicae by targeting chitin in a critical stage of this pathogen's life cycle. Our research aimed to evaluate the effect of various applications of P. chitinolyticus on clubroot suppression in two economically important Brassica species: Chinese cabbage and rapeseed. The effectiveness of the cell-free supernatant (CFS) of an endemic strain of P. chitinolyticus from the Czech Republic at five different time points was studied in the greenhouse by measuring the disease severity index. Early application of P. chitinolyticus decreased the disease index significantly within both plant species resulting in increased shoot dry weight. In conclusion, the CFS of P.chitinolyticus has significant antagonistic activity against clubroot in Chinese cabbage and rapeseed in the early developmental stages of clubroot occurrence and holds the potential as a biofertilizer as well as bioprotectant agent in clubroot management of P. brassicae. [ABSTRACT FROM AUTHOR]

Oilseed radish (OR; Raphanus sativus var. oleiferus) is grown as a cover crop and develops a unique taproot, absorbing nitrogen left by the previous crop. The aim of this project was to investigate the resistance of OR cultivars (cvs.) to Plasmodiophora brassicae, the causal agent of clubroot disease. Twelve market cvs. were compared with cvs. of clubroot-resistant (CR) winter oilseed rape (OSR; Brassica napus) and other selected species of the Brassicaceae family. The study was performed as a replicated bioassay in a growth chamber using a specially composed mixture of field soils holding the natural inoculum of P. brassicae. The results show that the OR cultivars were infected, which implies that OR multiplies the pathogen. The susceptibility of the OR cultivars was not significantly different from that of the CR OSR cultivars Alister and Archimedes, but it was significantly different from that of the OSR cv. Mendel. The disease severity index (DSI) for OR cultivars ranged from 2.3 to 9.3, and disease incidence was 3–17%. The best performance was shown by black radish (Raphanus sativus var. niger) with a DSI of 0.3. For sustainable brassica crop production, we suggest avoiding OR as a cover crop in crop rotations, including OSR or other brassica crops, since there is a risk of increasing inoculum in the soil. [ABSTRACT FROM AUTHOR]

Clubroot, caused by Plasmodiophora brassicae, is a significant disease affecting brassica crops worldwide and poses a threat to canola (Brassica napus) production in western Canada. Management of this disease heavily relies on the use of resistant cultivars, but resistance erosion is a serious concern due to the highly diverse pathogen populations. Understanding resistance mechanisms may aid in better deployment/rotation of clubroot resistance (CR) genes and improve resistance resilience. In this study, we conducted a comparative analysis using resistant canola varieties carrying either a single (Rcr1) or double CR genes (Rcr1+Crr1rutb) to decipher the resistance modes associated with these genes. Cell wall (CW) biopolymeric compounds in different root layers were mapped and quantified using Fourier-transform mid-infrared microspectroscopy for changes in CW elements associated with clubroot resistance. Transmission electron and confocal microscopy were used to assess root infection details and relative transcript abundance was analyzed to determine the activation of the lignin-related pathway in relation to resistance. Neither resistant variety affected the primary infection of root hairs/epidermal cells compared to the susceptible "Westar", but both exhibited strong inhibition of cortical infection, effectively 'trapping' the pathogen in the exodermis. The most prominent change observed was increased lignin accumulation associated with resistance. In Westar, the pathogen was able to degrade CW lignin, facilitating access to the root cortex by secondary plasmodia of P. brassicae. In contrast, resistant varieties showed clear lignin accumulation around the penetration site on the exodermis, accompanied by elevated expression of genes involved in the phenylpropanoid pathway. These results suggest that induced lignin accumulation plays a role in clubroot resistance mediated by the CR genes Rcr1 and Crr1rutb in canola, providing cellular and structural evidence that supports the data from earlier transcriptomic studies. [ABSTRACT FROM AUTHOR]

In the genome of the obligate biotrophic protist Plasmodiophora brassicae, which causes clubroot disease in the Brassicaceae family, a single gene was identified with homology to the group of GH3 proteins synthesizing amino acid conjugates with carboxylic acids. In comparison to plant members the P. brassicae GH3 protein seems to be very promiscuous with respect to the substrates converted. In vitro assays showed conversion of three different auxins (indole-3-acetic, IAA; indole-propionic, IPA; and indole-3-butyric acids, IBA), jasmonic acid (JA) and also 12-oxo-phytodienoic acid (OPDA). Auxin and Methyl-JA responsive elements were found in the promoter sequence of PbGH3. For further analysis Arabidopsis thaliana was transformed with PbGH3. Seedlings with confirmed constitutive PbGH3 protein production were grown to adult plants that were shown to produce fewer inflorescences per plant. Growth on 10 µM IAA or JA levels also altered the phenotype of the seedlings, but not so much between wild types and transgenic lines. The hormone profiles in seedlings and adult plants were examined for IAA, JA, OPDA, and respective conjugates. Transgenic seedlings displayed changes that could be partially attributed to the overexpression of a GH3 gene like lower levels of free IAA and the JA precursor cis-OPDA under control conditions, and higher levels of some auxin conjugates accumulated after IAA treatment compared to the corresponding wild types. However, the hormone patterns in adult plants were more complex and varied based on different tissue types analyzed. After inoculation with P. brassicae the transgenic plants did not show any changes in infection rate and disease index, but had higher shoot and lower root weight in infected plants compared to controls. Based on the above findings we discuss a role for PbGH3 during pathogenesis. [ABSTRACT FROM AUTHOR]

The sustainable cultivation of canola is under threat from clubroot disease (Plasmodiophora brassicae). The pathogen's resting spores can survive in the soil for extended periods, complicating disease management. Therefore, effective clubroot control requires a combination of tactics that provide multiple layers of protection. Management strategies have focused on pathogen avoidance and reducing disease levels in infested fields. The sanitation of machinery and field equipment remains the most effective method for preventing the pathogen's introduction into non-infested fields. For disease reduction, crop rotation, liming, chemical control, and host resistance are commonly employed, with the use of clubroot-resistant cultivars being the most effective to date. However, resistance breakdown has been observed within four years of the introduction of new cultivars, jeopardizing the long-term effectiveness of this approach. A promising yet underexplored strategy is the use of cultivar mixtures. This approach leverages mechanisms such as the dilution effect, the barrier effect, induced resistance, disruptive selection, and the compensatory effect to control the disease. Cultivar mixtures have the potential to reduce the impact of clubroot on canola production while preserving pathogen population structure, thereby minimizing the likelihood of resistance breakdown. Given its potential, further research into cultivar mixtures as a management strategy for clubroot disease is warranted. [ABSTRACT FROM AUTHOR]

Many economically important crops and vegetables belonging to the cruciferous family are heavily endangered by clubroot disease caused by Plasmodiophora brassicae infection. Breeding of clubroot resistant cultivars based on mapping and cloning of resistant genes is commonly regarded as the most cost-effective and efficient way to fight against this disease. The traditional way of R gene functional validation requires stable transformation that is both time- and labor-consuming. In this study, a rapid and efficient hairy-root transgenic protocol mediated by Agrobacterium rhizogenes was developed. The transformation positive rate was over 80% in Brassica napus showed by GUS reporter gene and this transformation only took 1/6 of the time compared with stable transformation. The system was applicable to different B. napus varieties and other cruciferous crops including Brassica rapa and Brassica oleracea. In particular, two known CR genes, CRA3.7.1 and CRA8.2.4 were used respectively, as example to show that the system works well for CR gene study combined with subsequent P. brassicae infection in B. napus. Most importantly, it works both in over-expression that led to disease resistance, as well as in RNAi which led to disease susceptible phenotype. Therefore, this system can be used in batch-wise identification of CR genes, and also offered the possibility of manipulating key genes within the P. brassicae genome that could improve our knowledge on host–pathogen interaction.

Clubroot disease is a severe threat to Brassica crops globally, particularly in western Canada. Genetic resistance, achieved through pyramiding clubroot resistance (CR) genes with different modes of action, is the most important strategy for managing the disease. However, studies on the CR gene functions are quite limited. In this study, we have conducted investigations into the temporal, structural, and interacting features of a newly cloned CR gene, Rcr1, using CRISPR/Cas9 technology. For temporal functionality, we developed a novel CRISPR/Cas9- based binary vector, pHHIGR-Hsp18.2, to deliver Rcr1 into a susceptible canola line (DH12075) and observed that early expression of Rcr1 is critical for conferring resistance. For structural functionality, several independent mutations in specific domains of Rcr1 resulted in loss-offunction, highlighting their importance for CR phenotype. In the study of the interacting features of Rcr1, a cysteine protease gene and its homologous allele in canola were successfully disrupted via CRISPR/Cas9 as an interacting component with Rcr1 protein, resulting in the conversion from clubroot resistant to susceptible in plants carrying intact Rcr1. These results indicated an indispensable role of these two cysteine proteases in Rcr1-mediated resistance response. This study, the first of its kind, provides valuable insights into the functionality of Rcr1. Further, the new vector pHHIGR-Hsp18.2 demonstrated an inducible feature on the removal of add-on traits, which should be useful for functional genomics and other similar research in brassica crops. [ABSTRACT FROM AUTHOR]

Clubroot disease caused by Plasmodiophora brassicae is becoming a serious threat to rapeseed (Brassica napus) production worldwide. Breeding resistant varieties using CR (clubroot resistance) loci is the most promising solution. Using marker‐assisted selection and speed‐breeding technologies, we generated Brassica napus materials in homozygous or heterozygous states using CRA3.7, CRA08.1, and CRA3.2 loci in the elite parental line of the Zhongshuang11 background. We developed three elite lines with two CR loci in different combinations and one line with three CR loci at the homozygous state. In our study, we used six different clubroot strains (Xinmin, Lincang, Yuxi, Chengdu, Chongqing, and Jixi) which are categorized into three groups based on our screening results. The newly pyramided lines with two or more CR loci displayed better disease resistance than the parental lines carrying single CR loci. There is an obvious gene dosage effect between CR loci and disease resistance levels. For example, pyramided lines with triple CR loci in the homozygous state showed superior resistance for all pathogens tested. Moreover, CR loci in the homozygous state are better on disease resistance than the heterozygous state. More importantly, no negative effect was observed on agronomic traits for the presence of multiple CR loci in the same background. Overall, these data suggest that the pyramiding of triple clubroot resistance loci conferred superior resistance with no negative effects on agronomic traits in Brassica napus. [ABSTRACT FROM AUTHOR]

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Clubroot disease, which is caused by the obligate biotrophic protist Plasmodiophora brassicae, leads to the formation of galls, commonly known as pathogen-induced tumors, on the roots of infected plants. The identification of crucial regulators of host tumor formation is essential to unravel the mechanisms underlying the proliferation and differentiation of P. brassicae within plant cells. To gain insight into this process, transcriptomic analysis was conducted to identify key genes associated with both primary and secondary infection of P. brassicae in Chinese cabbage. Our results demonstrate that the k-means clustering of subclass 1, which exhibited specific trends, was closely linked to the infection process of P. brassicae. Of the 1610 differentially expressed genes (DEGs) annotated in subclass 1, 782 were identified as transcription factors belonging to 49 transcription factor families, including bHLH, B3, NAC, MYB_related, WRKY, bZIP, C2H2, and ERF. In the primary infection, several genes, including the predicted Brassica rapa probable pectate lyase, RPM1-interacting protein 4-like, L-type lectin-domain-containing receptor kinase, G-type lectin S-receptor-like serine, B. rapa photosystem II 22 kDa protein, and MLP-like protein, showed significant upregulation. In the secondary infection stage, 45 of 50 overlapping DEGs were upregulated. These upregulated DEGs included the predicted B. rapa endoglucanase, long-chain acyl-CoA synthetase, WRKY transcription factor, NAC domain-containing protein, cell division control protein, auxin-induced protein, and protein variation in compound-triggered root growth response-like and xyloglucan glycosyltransferases. In both the primary and secondary infection stages, the DEGs were predicted to be Brassica rapa putative disease resistance proteins, Ltype lectin domain-containing receptor kinases, ferredoxin-NADP reductases, 1-aminocyclopropane-1-carboxylate synthases, histone deacetylases, UDPglycosyltransferases, putative glycerol-3-phosphate transporters, and chlorophyll a-binding proteins, which are closely associated with plant defense responses, biosynthetic processes, carbohydrate transport, and photosynthesis. This study revealed the pivotal role of transcription factors in the initiation of infection and establishment of intracellular parasitic relationships during the primary infection stage, as well as the proliferation and differentiation of the pathogenwithin the host cell during the secondary infection stage. [ABSTRACT FROM AUTHOR]

The resistance of Arabidopsis thaliana to clubroot, a major disease of Brassicaceae caused by the obligate protist Plasmodiophora brassicae, is controlled in part by epigenetic factors. The detection of some of these epigenetic quantitative trait loci (QTLepi) has been shown to depend on experimental conditions. The aim of the present study was to assess whether and how temperature and/or soil water availability influenced both the detection and the extent of the effect of response QTLepi. The epigenetic recombinant inbred line (epiRIL) population, derived from the cross between ddm1-2 and Col-0 (partially resistant and susceptible to clubroot, respectively), was phenotyped for response to P. brassicae under four abiotic conditions including standard conditions, a 5°C temperature increase, drought, and flooding. The abiotic constraints tested had a significant impact on both the leaf growth of the epiRIL population and the outcome of the epiRIL-pathogen interaction. Linkage analysis led to the detection of a total of 31 QTLepi, 18 of which were specific to one abiotic condition and 13 common to at least two environments. EpiRIL showed significant plasticity under epigenetic control, which appeared to be specific to the traits evaluated and to the abiotic conditions. These results highlight that the environment can affect the epigenetic architecture of plant growth and immune responses and advance our understanding of the epigenetic factors underlying plasticity in response to climate change. [ABSTRACT FROM AUTHOR]

The clubroot disease has become a worldwide threat for crucifer crop production, due to its soil-borne nature and difficulty to eradicate completely from contaminated field. In this study we used an elite resistant European fodder turnip ECD04 and investigated its resistance mechanism using transcriptome, sRNA-seq, degradome and gene editing. A total of 1751 DEGs were identified from three time points after infection, among which 7 hub genes including XTH23 for cell wall assembly and two CPK28 genes in PTI pathways. On microRNA, we identified 17 DEMs and predicted 15 miRNA-target pairs (DEM-DEG). We validated two pairs (miR395-APS4 and miR160-ARF) by degradome sequencing. We investigated the miR395-APS4 pair by CRISPR-Cas9 mediated gene editing, the result showed that knocking-out APS4 could lead to elevated clubroot resistance in B. napus. In summary, the data acquired on transcriptional response and microRNA as well as target genes provide future direction especially gene candidates for genetic improvement of clubroot resistance on Brassica species. [ABSTRACT FROM AUTHOR]

Plasmodiophora brassicae (Woronin, 1877), a biotrophic, obligate parasite, is the causal agent of clubroot disease in brassicas. The clubroot pathogen has been reported in more than 80 countries worldwide, causing economic losses of hundreds of millions every year. Despite its widespread impact, very little is known about the molecular strategies it employs to induce the characteristic clubs in the roots of susceptible hosts during infection, nor about the mechanisms it uses to overcome genetic resistance. Here, we provide the first telomere-to-telomere complete genome of P. brassicae. We generated ∼27 Gb of Illumina, Oxford Nanopore, and PacBio HiFi data from resting spores of strain Pb3A and produced a 25.3 Mb assembly comprising 20 chromosomes, with an N50 of 1.37 Mb. The BUSCO score, the highest reported for any member of the group Rhizaria (Eukaryota: 88.2%), highlights the limitations within the Eukaryota database for members of this lineage. Using available transcriptomic data and protein evidence, we annotated the Pb3A genome, identifying 10,521 protein-coding gene models. This high-quality, complete genome of P. brassicae will serve as a crucial resource for the plant pathology community to advance the much-needed understanding of the evolution of the clubroot pathogen. [ABSTRACT FROM AUTHOR]

Pyramiding resistance genes may expand the efficacy and scope of a canola variety against clubroot (Plasmodiophora brassicae), a serious threat to canola production in western Canada. However, the mechanism(s) of multigenic resistance, especially the potential interaction among clubroot resistance (CR) genes, are not well understood. In this study, transcriptome was compared over three canola (Brassica napus L.) inbred/hybrid lines carrying a single CR gene in chromosome A03 (CRaM, Line 16) or A08 (Crr1rutb, Line 20), and both genes (CRaM+Crr1rutb, Line 15) inoculated with a field population (L-G2) of P. brassicae pathotype X, a new variant found in western Canada recently. The line16 was susceptible, while lines 15 and 20 were partially resistant. Functional annotation identified differential expression of genes (DEGs) involved in biosynthetic processes responsive to stress and regulation of cellular process; The Venn diagram showed that the partially resistant lines 15 and 20 shared 1,896 differentially expressed genes relative to the susceptible line 16, and many of these DEGs are involved in defense responses, activation of innate immunity, hormone biosynthesis and programmed cell death. The transcription of genes involved in Pathogen-Associated Molecular Pattern (PAMP)-Triggered and Effector-Triggered Immunity (PTI and ETI) was particularly up-regulated, and the transcription level was higher in line 15 (CRaM + Crr1rutb) than in line 20 (Crr1rutb only) for most of the DEGs. These results indicated that the partial resistance to the pathotype X was likely conferred by the CR gene Crr1rutb for both lines 15 and 20 that functioned via the activation of both PTI and ETI signaling pathways. Additionally, these two CR genes might have synergistic effects against the pathotype X, based on the higher transcription levels of defense-related DEGs expressed by inoculated line 15, highlighting the benefit of gene stacking for improved canola resistance as opposed to a single CR gene alone. [ABSTRACT FROM AUTHOR]

In this study, we explored the resilience of a clubroot resistance (CR) stacking model against a field population of Plasmodiophora brassicae pathotype 3H. This contrasts with our earlier work, where stacking CRaM and Crr1rutb proved only moderately resistant to pathotype X. Canola varieties carrying Rcr1/Crr1rutb and Rcr1 + Crr1rutb were repeatedly exposed to 3H at low (1 × 104/g soil) and high (1 × 107/g soil) initial resting spore concentrations over five planting cycles under controlled environments to mimic intensive canola production. Initially, all resistant varieties showed strong resistance. However, there was a gradual decline in resistance over time for varieties carrying only a single CR gene, particularly with Crr1rutb alone and at the high inoculum level, where the disease severity index (DSI) increased from 9% to 39% over five planting cycles. This suggests the presence of virulent pathotypes at initially low levels in the 3H inoculum. In contrast, the variety with stacked CR genes remained resilient, with DSI staying below 3% throughout, even at the high inoculum level. Furthermore, the use of resistant varieties, carrying either a single or stacked CR genes, reduced the total resting spore numbers in soil over time, while the inoculum level either increased or remained high in soils where susceptible Westar was continuously grown. Our study demonstrates greater resistance resilience for stacking Rcr1 and Crr1rutb against the field population of 3H. Additionally, the results suggest that resistance may persist even longer in fields with lower levels of inoculum, highlighting the value of extended crop rotation (reducing inoculum) alongside strategic CR-gene deployment to maximize resistance resilience. [ABSTRACT FROM AUTHOR]

Clubroot disease poses a significant threat to Brassica crops, necessitating ongoing updates on resistance gene sources. In F2 segregants of the clubrootresistant inbred line BrT18-6-4-3 and susceptible DH line Y510, the genetic analysis identified a single dominant gene responsible for clubroot resistance. Through bulk segregant sequencing analysis and kompetitive allele-specific polymerase chain reaction assays, CRA8.1.6 was mapped within 110 kb (12,255-12,365 Mb) between markers L-CR11 and L-CR12 on chromosome A08. We identified BraA08g015220.3.5C as the candidate gene of CRA8.1.6. Upon comparison with the sequence of disease-resistant material BrT18-6-4-3, we found 249 single-nucleotide polymorphisms, seven insertions, six deletions, and a long terminal repeat (LTR) retrotransposon (5,310 bp) at 909 bp of the first intron. However, the LTR retrotransposon was absent in the coding sequence of the susceptible DH line Y510. Given the presence of a non-functional LTR insertion in other materials, it showed that the LTR insertion might not be associated with susceptibility. Sequence alignment analysis revealed that the fourth exon of the susceptible line harbored two deletions and an insertion, resulting in a frameshift mutation at 8,551 bp, leading to translation termination at the leucine-rich repeat domain's C-terminal in susceptible material. Sequence alignment of the CDS revealed a 99.4% similarity to Crr1a, which indicate that CRA8.1.6 is likely an allele of the Crr1a gene. Two functional markers, CRA08-InDel and CRA08-KASP1, have been developed for marker-assisted selection in CR turnip cultivars. Our findings could facilitate the development of clubrootresistance turnip cultivars through marker-assisted selection. [ABSTRACT FROM AUTHOR]

Clubroot caused by Plasmodiophora brassicae is a devastating disease of Cruciferous crops. Developing cultivars with clubroot resistance (CR) is the most effective control measure. For the two major Brassica vegetable species B. rapa and B. oleracea, several commercial cultivars with unclear CR pedigrees have been intensively used as CR donors in breeding. However, the continuous occurrence of CR-breaking makes the CR pedigree underlying these cultivars one of the breeders' most urgent concerns. The complex intraspecific diversity of these two major Brassica vegetables has also limited the applicability of CR markers in different breeding programs. Here we first traced the pedigree underlying two kinds of CR that have been widely applied in breeding by linkage and introgression analyses based on public resequencing data. In B. rapa, a major locus CRzi8 underlying the CR of the commercial CR donor 'DegaoCR117' was identified. CRzi8 was further shown to have been introgressed from turnip (B. rapa ssp. rapifera) and that it carried a potential functional allele of Crr1a. The turnip introgression carried CRbc, sharing the same coding sequence with the CRb that was also identified from chromosome C07 of B. oleracea CR cultivars with different morphotypes. Within natural populations, variation analysis of linkage intervals of CRzi8, PbBa8.1, CRb, and CRbc yielded easily resolved InDel markers (> 20 bp) for these fundamental CR genes. The specificity of these markers was tested in diverse cultivars panels, and each exhibited high reliability in breeding. Our research demonstrates the value of the practice of applying resequencing big data to solve urgent concerns in breeding programs. [ABSTRACT FROM AUTHOR]

Plasmodiophora brassicae Woronin is responsible for an infectious disease called clubroot, which poses a significant threat to cruciferous vegetables. This study aimed to determine the efficacy of biological agents including Trichoderma harzianum and Sophora flavescens in reducing clubroot severity in Pak choi. For this purpose, an in vitro pot experiment was conducted. For the application of T. harzianum (MD), 10 g powder was mixed in each pot with the substrate, while 5 mL (from the diluted solution 5 mL/L) of S. flavescens (RD), and 20 mL (from the diluted solution 500 µL/L) of fluazinam (ZD) was applied around the root of every plant along with clubroot pathogen P. brassicae. The results revealed that the clubroot disease index (DI) efficiently reduced by the application of T. harzianum and S. flavescens up to 36.02%, 45.43%, as compared to positive control fluazinam 49.05%, respectively. Moreover, under clubroot stress T. harzianum increased total chlorophyll contents, total carotenoid contents, shoot diameter, leaf number, fresh weight of shoot by 15.33%, 21.21%, 21.83%, 25%, 14.24%, and enzymatic activities of peroxidase (POD), catalase (CAT), superoxide dismutase (SOD) increased by 14.85%, 21.76% and 19.22%, respectively, as compared to fluazinam. Additionally, the 16S rRNA gene sequence of endophytic bacterial communities under clubroot stress revealed that plants treated with P. brassicae only (D) increased the relative abundance of phylum Proteobacteria by 82% as compared to fluazinam (67%). Beta diversity distance matrix analysis based on the principal coordinate algorithm (PCoA) and Non-metric multidimensional scaling method showed a significant distance between T. harzianum and other applied treatments. Overall, T. harzianum and S. flavescens affected and structured the bacterial community in Pak choi and efficiently reduced the clubroot disease. This study will facilitate researchers to biologically control cruciferous clubroot and their ecological functions with endophytic bacteria in brassica species. [ABSTRACT FROM AUTHOR]

The effect of crop rotation on soil-borne diseases is a representative case of plant–soil feedback in the sense that plant disease resistance is influenced by soils with different cultivation histories. This study examined the microbial mechanisms inducing the differences in the clubroot (caused by Plasmodiophora brassicae pathogen) damage of Chinese cabbage (Brassica rapa subsp. pekinensis) after the cultivation of different preceding crops. It addresses two key questions in crop rotation: changes in the soil bacterial community induced by the cultivation of different plants and the microbial mechanisms responsible for the disease-suppressive capacity of Chinese cabbage. Twenty preceding crops from different plant families showed significant differences in the disease damage, pathogen density, and bacterial community composition of the host plant. Structural equation modelling revealed that the relative abundance of four key bacterial orders in Chinese cabbage roots can explain 85% and 70% of the total variation in pathogen density and disease damage, respectively. Notably, the relative dominance of Bacillales and Rhizobiales, which have a trade-off relationship, exhibited predominant effects on pathogen density and disease damage. The disease-suppressive soil legacy effects of preceding crops are reflected in compositional changes in key bacterial orders, which are intensified by the bacterial community network.

Clubroot is one of the important diseases affecting members of the Cruciferae family. This disease is caused by soil-borne pathogen, called Plasmodiophora brassicae. The pathogen produces motile spores called zoospores. This pathogen results in a drop of cruciferous plant productivity. The objectives of this research were to identify specific Trichoderma species in three districts in Sumatra Utara and to evaluate the potency of T. harzianum (both local species from Berastagi and species developed by Indonesian Vegetable Research Institute (IVEGRI)) to control the disease and then comparing them with the application of synthetic fungicide. The research was conducted in a greenhouse of Research Installation and Application of Agricultural Technology (IP2TP), Berastagi, North Sumatera from April-September 2018, using Completely Randomized Design (CRD) with eight treatments: C1 (control-without Trichoderma), C2 (T. harzianum IVEGRI obtained from corn substrate 2 g/polybag), C3 (T. harzianum IVEGRI obtained from rice substrate 2 g/polybag), C4 (T. harzianum IVEGRI obtained from corn substrate 4 g/polybag, C5 (T. harzianum IVEGRI obtained from rice substrate 4 g/polybag), C6 (local T. harzianum obtained from corn substrate 2 g/polybag), C7 (local T. harzianum obtained from corn substrate 4 g/polybag) and C8 (synthetic fungicide Nebijin). Each treatment contained 10 polybags of plants. These treatments were replicated four times. The results exhibited there were 3 Trichoderma species found in Berastagi: T. harzianum, T. viride and T. koningii. Also, 4 g of local T. harzianum (corn substrate) has better performance (0% disease incidence and 0% disease severity) compared to other treatments. [ABSTRACT FROM AUTHOR]

Straw return is an effective agricultural management practice for alleviating soil sickness, but only a few studies have focused on the incorporation of straw with deep plowing and rotary tillage practices in vegetable production. To determine the effects of rice straw return on Chinese cabbage clubroot, a field experiment for three consecutive years in the same area was performed. Soil microbial high-throughput sequencing, quantitative real-time polymerase chain reaction (PCR) and other methods were used to detect Chinese cabbage plant growth, clubroot occurrence, soil chemical properties and soil microbial diversity and abundance. The results showed that straw addition could significantly reduce the clubroot disease incidence. Through Illumina Miseq sequencing, the diversity of the fungi decreased obviously. The relative abundance of the phyla Proteobacteria and Firmicutes was strikingly reduced, while that of Chloroflexi was significantly increased. Redundancy analysis suggests that soil properties may also affect the soil microbial composition; changes in the microbial structure of bacteria and fungi were associated with the available phosphorus. In conclusion, the continuous addition of rice straw can promote the growth and control the occurrence of clubroot, which is closely related to the microbial composition, and the inhibition effect is proportional to the age of addition. [ABSTRACT FROM AUTHOR]

Plasmodiophora brassicae Woronin, an obligate biotrophic soil-borne pathogen, poses a significant threat to cruciferous crops worldwide by causing the devastating disease known as clubroot. Pathogenic variability in P. brassicae populations has been recognized since the 1930s based on its interactions with Brassica species. Over time, numerous sets of differential hosts have been developed and used worldwide to explore the pathogenic variability within P. brassicae populations. These sets encompass a range of systems, including the Williams system, the European Clubroot Differential set (ECD), the Brassica napus set, the Japanese Clubroot Differential Set, the Canadian Clubroot Differential Set (CCS), the Korean Clubroot Differential Set, and the Chinese Sinitic Clubroot Differential set (SCD). However, all existing systems possess both advantages as well as limitations regarding the detection of pathotypes from various Brassica species and their corresponding virulence pattern on Brassica genotypes. This comprehensive review aims to compare the main differential systems utilized in classifying P. brassicae pathotypes worldwide. Their strengths, limitations, and implications are evaluated, thereby enhancing our understanding of pathogenic variability. [ABSTRACT FROM AUTHOR]

The soil-borne pathogen Plasmodiophora brassicae is the causal agent of clubroot, a major disease in Chinese cabbage (Brassica rapa ssp. pekinensis). The host's resistance genes often confer immunity to only specific pathotypes and may be rapidly overcome. Identification of novel clubroot resistance (CR) from germplasm sources is necessary. In this study, Bap246 was tested by being crossed with different highly susceptible B. rapa materials and showed recessive resistance to clubroot. An F2 population derived from Bap246 × Bac1344 was used to locate the resistance Quantitative Trait Loci (QTL) by Bulk Segregant Analysis Sequencing (BSA-Seq) and QTL mapping methods. Two QTL on chromosomes A01 (4.67–6.06 Mb) and A08 (10.42–11.43 Mb) were found and named Cr4Ba1.1 and Cr4Ba8.1, respectively. Fifteen and eleven SNP/InDel markers were used to narrow the target regions in the larger F2 population to 4.67–5.17 Mb (A01) and 10.70–10.84 Mb (A08), with 85 and 19 candidate genes, respectively. The phenotypic variation explained (PVE) of the two QTL were 30.97% and 8.65%, respectively. Combined with gene annotation, mutation site analysis, and real-time quantitative polymerase chain reaction (qRT-PCR) analysis, one candidate gene in A08 was identified, namely Bra020861. And an insertion and deletion (InDel) marker (co-segregated) named Crr1-196 was developed based on the gene sequence. Bra013275, Bra013299, Bra013336, Bra013339, Bra013341, and Bra013357 in A01 were the candidate genes that may confer clubroot resistance in Chinese cabbage. The resistance resource and the developed marker will be helpful in Brassica breeding programs. [ABSTRACT FROM AUTHOR]

Breeding for disease resistance in major crops is of crucial importance for global food security and sustainability. However, common biotechnologies such as traditional transgenesis or genome editing do not provide an ideal solution, whereas transgenic crops free of selection markers such as cisgenic/intragenic crops might be suitable. In this study, after cloning and functional verification of the Rcr1 gene for resistance to clubroot (Plasmodiophora brassicae), we confirmed that the genes Rcr1 , Rcr2 , Rcr4 , and CRa from Brassica rapa crops and the resistance gene from B. napus oilseed rape cv. 'Mendel' on chromosome A03 were identical in their coding regions. We also determined that Rcr1 has a wide distribution in Brassica breeding materials and renders potent resistance against multiple representative clubroot strains in Canada. We then modified a CRISPR/Cas9-based cisgenic vector system and found that it enabled the fast breeding of selection-marker-free transgenic crops with add-on traits, with selection-marker-free canola (B. napus) germplasms with Rcr1 -rendered stable resistance to clubroot disease being successfully developed within 2 years. In the B. napus background, the intragenic vector system was able to remove unwanted residue sequences from the final product with high editing efficiency, and off-target mutations were not detected. Our study demonstrates the potential of applying this breeding strategy to other crops that can be transformed by Agrobacterium. Following the streamlined working procedure, intragenic germplasms can be developed within two generations, which could significantly reduce the breeding time and labor compared to traditional introgression whilst still achieving comparable or even better breeding results. [ABSTRACT FROM AUTHOR]

Clubroot, caused by Plasmodiophora brassicae, is a destructive soil-borne disease significantly harming global Brassica crop production. This study employed the Williams and European Clubroot Differential (ECD) and Williams systems to identify the pathotypes of P. brassicae collected from Hangzhou City, Yuhang District and Quzhou City, Kaihua County in Zhejiang Province. Greenhouse and field trials were conducted to evaluate the effects of plastic film covering and four chemical agents on the growth parameters and clubroot severity of the Chinese cabbage cultivar 'Granaat'. Potential treatment mechanisms on clubroot were explored through a qPCR analysis of the resting spore density and pH measurement of the soil. Furthermore, treatment with 1-napthaleneacetic acid (NAA), a synthetic auxin, was also evaluated for its potential role in suppressing clubroot. The results indicate that the pathotypes of P. brassicae in the two districts were P1, ECD20/31/12, and P3, ECD20/15/4. While an individual application of plastic film covering could not effectively control clubroot, calcium cyanamid, dazomet and ammonium bicarbonate demonstrated significant efficacy in its management. These three agents significantly reduced the resting spore density in the soil, with calcium cyanamid and ammonium bicarbonate also increasing soil alkalinity. Additionally, ammonium bicarbonate promotes lateral root development in 'Granaat,' helping infected plants access adequate water and nutrients. However, NAA exhibited no efficacy in clubroot control. Therefore, sustained lateral root development is crucial for effectively resisting P. brassicae invasion. Considering application costs and environmental friendliness, we propose the field application of ammonium bicarbonate as the optimal method for clubroot disease management in Zhejiang Province. [ABSTRACT FROM AUTHOR]

The resistance of Arabidopsis thaliana to clubroot, a major disease of Brassicaceae caused by the obligate protist Plasmodiophora brassicae, is controlled in part by epigenetic factors. The detection of some of these epigenetic quantitative trait loci (QTLepi) has been shown to depend on experimental conditions. The aim of the present study was to assess whether and how temperature and/or soil water availability influenced both the detection and the extent of the effect of response QTLepi. The epigenetic recombinant inbred line (epiRIL) population, derived from the cross between ddm1-2 and Col-0 (partially resistant and susceptible to clubroot, respectively), was phenotyped for response to P. brassicae under four abiotic conditions including standard conditions, a 5°C temperature increase, drought, and flooding. The abiotic constraints tested had a significant impact on both the leaf growth of the epiRIL population and the outcome of the epiRIL–pathogen interaction. Linkage analysis led to the detection of a total of 31 QTLepi, 18 of which were specific to one abiotic condition and 13 common to at least two environments. EpiRIL showed significant plasticity under epigenetic control, which appeared to be specific to the traits evaluated and to the abiotic conditions. These results highlight that the environment can affect the epigenetic architecture of plant growth and immune responses and advance our understanding of the epigenetic factors underlying plasticity in response to climate change.

Clubroot caused by the protist Plasmodiophora brassicae is a major disease affecting cultivated Brassicaceae. Using a combination of quantitative trait locus (QTL) fine mapping, CRISPR-Cas9 validation, and extensive analyses of DNA sequence and methylation patterns, we revealed that the two adjacent neighboring NLR (nucleotide-binding and leucine-rich repeat) genes AT5G47260 and AT5G47280 cooperate in controlling broad-spectrum quantitative partial resistance to the root pathogen P. brassicae in Arabidopsis and that they are epigenetically regulated. The variation in DNA methylation is not associated with any nucleotide variation or any transposable element presence/absence variants and is stably inherited. Variations in DNA methylation at the Pb-At5.2 QTL are widespread across Arabidopsis accessions and correlate negatively with variations in expression of the two genes. Our study demonstrates that natural, stable, and transgenerationally inherited epigenetic variations can play an important role in shaping resistance to plant pathogens by modulating the expression of immune receptors.

Chinese cabbage is an economically important Brassica vegetable worldwide, and clubroot, which is caused by the soil-borne protist plant pathogen Plasmodiophora brassicae is regarded as a destructive disease to Brassica crops. Previous studies on the gene transcripts related to Chinese cabbage resistance to clubroot mainly employed RNA-seq technology, although it cannot provide accurate transcript assembly and structural information. In this study, PacBio RS II SMRT sequencing was used to generate full-length transcriptomes of mixed roots at 0, 2, 5, 8, 13, and 22 days after P. brassicae infection in the clubroot-resistant line DH40R. Overall, 39 376 high-quality isoforms and 26 270 open reading frames (ORFs) were identified from the SMRT sequencing data. Additionally, 426 annotated long noncoding RNAs (lncRNAs), 56 transcription factor (TF) families, 1 883 genes with poly(A) sites and 1 691 alternative splicing (AS) events were identified. Furthermore, 1 201 of the genes had at least one AS event in DH40R. A comparison with RNA-seq data revealed six differentially expressed AS genes (one for disease resistance and five for defensive response) that are potentially involved in P. brassicae resistance. The results of this study provide valuable resources for basic research on clubroot resistance in Chinese cabbage.

Infection by the mustard clubroot disease pathogen Plasmodiophora brassicae has a significant negative impact on the quality and yield of Chinese mustard (Brassica juncea). At present, screening resistant resources for breeding programs is the most economical and effective method available to control this disease. In this study, we isolated P. brassicae physiological race 4 from Chinese cabbage and examined 483 mustard germplasm resources (193 leaf mustard, 96 stem mustard, and 194 root mustard) from China and abroad to identify resistance to clubroot disease at the seedling stage through irrigation inoculation with the isolated pathogen. The results showed that there were no immune varieties among the tested mustard germplasm, but that there were differences in resistance to clubroot disease among the three mustard types. More than 90% of leaf and stem mustard resources were susceptible to clubroot disease, whereas 38.66% of root mustard resources showed resistance. In total, we detected 4 highly resistant, 9 resistant, and 83 moderately resistant varieties, of which 4 highly resistant, 8 resistant, and 63 moderately resistant varieties were root mustard resources, whereas only 1 resistant and 5 moderately resistant varieties were stem mustard resources, and 15 moderately resistant varieties were leaf mustard resources. In addition, we used seven molecular markers for clubroot disease resistance in Chinese cabbage to detect stem and root mustard resources. The results showed that the marker CRk was detected in 97.87% of stem mustard and 92.49% of root mustard resources. Six markers (Crr1, Crr2, Crr3, CRa, CRb, and CRc) were detected in 18.09%, 7.45%, 2.13%, 6.38%, 12.77%, and 12.77% of stem mustard germplasms, and four markers (Crr1, Crr2, Crr3, and CRc) were detected in 8.09%, 8.67%, 10.40%, and 8.67% of root mustard germplasms, respectively, suggesting that these markers are not suitable for detecting mustard germplasm resistance to clubroot disease. This study provides a technical reference and material support for the breeding of mustard varieties resistant to clubroot disease.

Oilseed radish (OR; Raphanus sativus var. oleiferus) is grown as a cover crop and develops a unique taproot, absorbing nitrogen left by the previous crop. The aim of this project was to investigate the resistance of OR cultivars (cvs.) to Plasmodiophora brassicae, the causal agent of clubroot disease. Twelve market cvs. were compared with cvs. of clubroot-resistant (CR) winter oilseed rape (OSR; Brassica napus) and other selected species of the Brassicaceae family. The study was performed as a replicated bioassay in a growth chamber using a specially composed mixture of field soils holding the natural inoculum of P. brassicae. The results show that the OR cultivars were infected, which implies that OR multiplies the pathogen. The susceptibility of the OR cultivars was not significantly different from that of the CR OSR cultivars Alister and Archimedes, but it was significantly different from that of the OSR cv. Mendel. The disease severity index (DSI) for OR cultivars ranged from 2.3 to 9.3, and disease incidence was 3–17%. The best performance was shown by black radish (Raphanus sativus var. niger) with a DSI of 0.3. For sustainable brassica crop production, we suggest avoiding OR as a cover crop in crop rotations, including OSR or other brassica crops, since there is a risk of increasing inoculum in the soil.

The objective of this research was to determine the potential use of eco-friendly technologies to reduce the clubroot disease caused by Plasmodiophora brassicae, the main constraint of cruciferous crops worldwide. Two commercial bioproducts were evaluated in susceptible broccoli, one based on the PGPR consortium (Bacillus amyloliquefaciens, Bacillus pumilus, and Agrobacterium radiobacter K84) and the other one based on Trichoderma koningiopsis Th003 (Tricotec® WG). Additionally, the resistant broccoli cv. Monclano® was tested under two concentrations of resting spores (RS) of P. brassicae, 1 x 10³ and 1 x 105 RS g-1 of soil. The first phase of evaluations with broccoli was carried out under a greenhouse, while susceptible broccoli, cauliflower, and red cabbage were included in a subsequent field phase. Tebuconazole + Trifloxystrobin mixture and Fluazinam were included as positive controls. The effectiveness of the bioproducts depended on the nature of the biocontrol agent, the concentration of P. brassicae, and the dose of treatment. Tricotec® showed consistent plant growth promotion but no biocontrol effect against clubroot, and the rhizobacteria-based bioproduct significantly reduced the disease in both greenhouse and field experiments. Higher disease severity was observed with the higher dose of Tricotec®. Under field conditions, the rhizobacteria reduced the incidence progress by 26%, 39%, and 57% under high, medium, and low pressure of the pathogen, respectively. However, no reduction of clubroot severity under high pressure of the pathogen was observed. Complete inhibition of club formation in roots was achieved via the fungicide, but a phytotoxic effect was observed under greenhouse conditions. Fungicides reduced the incidence progress of clubroot, but not the severity under high inoculum pressure in the field. The fungicides, the bacterial treatment, and the combination of bioproducts tended to delay the progress of the disease compared with the negative control and Tricotec alone. The resistant broccoli showed a low level of disease under high concentrations of P. brassicae (less than 10% incidence and up to 2% severity). These results suggested the overall potential of commercial tools based on the PGPR consortium and plant resistance to control P. brassicae. The integration of control measures, the role of Trichoderma spp. in P. brassicae--cruciferous pathosystems, and the need to recover highly infested soils will be discussed. [ABSTRACT FROM AUTHOR]

Clubroot disease, caused by Plasmodiophora brassicae, is a serious soil-borne disease in Brassica crops worldwide. It seriously occurs in conducive soils of southern China, while never happens in some areas of northern China with suppressive soils. To understanding the differences, we measured the soil suppressiveness, chemical properties, and microbial communities in suppressive and conducive soils by bioassay and sequencing of 16S and 18S rRNA amplicons. The biological basis of clubroot suppressiveness was supported by the ability to remove it by pasteurization. The pH value and calcium content in the suppressive soils were higher than those in the conducive soils. Suppressive soils were associated with higher fungal diversity and bacterial abundance. The fungal phyla Chytridiomycota, Olpidiomycota, and Mucoromycota and the bacterial phyla Acidobacteriota and Gemmatimonadota were enriched in suppressive soils. More abundant beneficial microbes, including Chaetomium and Lysobacter, were found in the suppressive soils than in the conducive soils. Molecular ecological network analysis revealed that the fungal network of suppressive soils was more complex than that of conducive soils. Our results indicate that plant health is closely related to soil physicochemical and biological properties. This study is of great significance for developing strategies for clubtroot disease prevention and control. [ABSTRACT FROM AUTHOR]

Flusulfamide inhibits germination of Plasmodiophora brassicae resting spores to suppress clubroot disease, but its mechanism of action on the germination of P. brassicae resting spores remains unclear. In this study, P. brassicae resting spores were treated with flusulfamide and visualized using transmission electron microscopy (TEM). The gene expression of P. brassicae resting spores was analyzed using RT-PCR, followed by immunoblotting analysis. TEM results revealed that flusulfamide suppressed the primary zoosporogenesis of P. brassicae resting spores during the early phase, and RT-PCR results revealed that flusulfamide affected the gene expression during the germination of the resting spores. Immunoblot and RT-qPCR analyses revealed that PbCyp3, an immunophilin (peptidyl-prolyl-isomerase) gene, was highly expressed, resulting in the unusual accumulation of PbCYP3 protein in P. brassicae resting spores immediately after treatment with flusulfamide. This suggests that flusulfamide may cause aberrant folding of proteins involved in primary zoosporogenesis, thereby inhibiting germination. [ABSTRACT FROM AUTHOR]

Clubroot disease, caused by Plasmodiophora brassicae, poses a significant global threat to cruciferous crops. The epidemic area of clubroot disease is expanding rapidly. In response to this pressing issue, there is a compelling need for the development of clubroot disease-resistant radish cultivars. China boasts an extensive array of radish varieties and germplasm resources. However, a comprehensive assessment of their resistance to clubroot has not yet been carried out, thereby impeding the effective utilization of germplasm and clubroot-resistant breeding. Therefore, it is urgent to systematically evaluate the clubroot resistance of the radish germplasm and identify resistant resources. In this study, clubroot resistance evaluations were conducted on 268 excellent radish varieties derived from 30 provinces in China, as well as seven accessions from Russia, North Korea, France, South Korea, and Germany. The resistance evaluation revealed a diverse range of resistance indices, with a mean disease index (DI) ranging from 0.6 to 58.5, showing significant disparities in clubroot resistance among these radish resources. A total of six accessions were characterized as highly resistant to clubroot, and a further 50 accessions were characterized as resistant. The disease-resistant radishes showed diversity in horticultural traits. Provinces in South China contributed significantly more resistance germplasm than those of North China. These materials are of great value for both genetic investigation and the crop breeding of clubroot resistance. Furthermore, we employed a previously established clubroot-resistance-linked SSR marker to analyze the clubroot-resistant resources. The accessions exhibited dissimilar genetic profiles from known clubroot-resistant germplasm, suggesting their potential status as novel sources of clubroot resistance. Conclusively, these newly identified accessions enriched the genetic diversity within the clubroot-resistant gene pool and may contribute to the future cloning of previously undiscovered clubroot-resistant genes. [ABSTRACT FROM AUTHOR]

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Clubroot, caused by Plasmodiophora brassicae, is an important disease of canola (Brassica napus). Amisulbrom, a quinone inside inhibitor (QiI), was evaluated for its effectiveness in clubroot management in Alberta, Canada. Resting spores of P. brassicae were treated in vitro with 0, 0.01, 0.1, 1, and 10% (w/v) amisulbrom to determine its effect on spore germination and viability. Amisulbrom inhibited resting spore germination by up to 79% and reduced viable spores by 31% relative to the control. Applications of a liquid solution (AL1000, 1000 g active ingredient (ai) ha−1) and granular formulations (AF700, 700 g ai ha−1; AF1000, 1000 g ai ha−1; AF1500, 1500 g ai ha−1) of amisulbrom were tested on the canola cultivars '45H31' (clubroot-susceptible) and 'CS2000' (moderately resistant) under greenhouse conditions and in field experiments in 2019 and 2020. In the greenhouse, the treatments were evaluated at inoculum concentrations of 1 × 105 or 1 × 107 resting spores g−1 soil. A trend of decreasing clubroot severity with an increasing amisulbrom rate was observed. At the lower spore concentration, treatment with AF1500 resulted in a clubroot disease severity index (DSI) <20% for both cultivars, while the lowest DSI under both low and high spore concentrations was obtained with AL1000. The field results indicated a significant reduction in DSI, with varied effects of rates and liquid vs. granular formulations. The greatest reductions (up to 58.3%) in DSI were obtained with AF1500 and AL1000 in 2020. These findings suggest that amisulbrom holds promise as part of an integrated clubroot management approach. [ABSTRACT FROM AUTHOR]

The commercial aspect of growing Brassica crops has always been influenced by the worldwide occurrence of the clubroot pathogen, Plasmodiophora brassicae. Clubroot symptoms reduce crop yield dramatically and the resilient protist is hard to eradicate from infested soils. Chemical treatments are not so efficient and their use is allowed only in a few areas of the world, none of them in the EU. The majority of clubroot control is mediated by using resistant crops, but not all species have good or durable resistance sources available, and these can be overcome by evolving or new Plasmodiophora pathotypes. Some commercially available biocontrol agents have been tested and found to reduce clubroot on crops such as rapeseed, cauliflower and Chinese cabbage to some extent. More biocontrol organisms have been isolated and described in recent decades but for many commercial application is still a long way off. In this review we summarize trends for bacterial and fungal endophytes for clubroot biocontrol as well as mechanisms behind the effects reported, such as antibiosis, defense induction or competition for space and nutrients. There are indeed plenty of studies on biocontrol of clubroot but not many have reached a point where the biocontrol agents are ready to be applied at field scale. The potential of endophytic microbes in pest management against clubroot disease is huge. [ABSTRACT FROM AUTHOR]

Clubroot disease has spread explosively in recent years, posing a serious threat to cruciferous crop production. Take rapeseed for example, according to the statistics, until year 2022 the incidence of the clubroot has exceeded 1.33 million hectares in China. Breeding for resistant cultivars is considered as the most efficient and economical way to prevent clubroot disease spreading. However, the current CR cultivars mostly used CR loci of PbBa8.1 or CRb. Due to the complexity and diversity of Plasmodiophora brassicae from different locations, resources for resistant germplasm need to be explored. In this study, 14 clubroot-resistant Chinese cabbages were characterized using different clubroot pathogens and molecular markers for known CR loci. We found that among them, five materials, DG117, ZS105, SG01, SR06 and JQ07 did not contain CRb locus, and their disease resistance were stronger than Huashuang5R/Huayouza5R. At last, KL02 containing the CRb locus showed stronger resistance to PbXM (strain from Xinmin) than 409R, therefore it may contain other novel loci leading to better resistance. These six Chinese cabbage resources can be considerd as new resistance resources and could be utilized in future breeding work for CR Brassica napus. [ABSTRACT FROM AUTHOR]

Currently, clubroot disease has become one of the main diseases of rapeseed in the Yangtze River basin, which seriously affects the development of rapeseed industry. To use molecular marker assisted selection technology to rapidly transfer parents with resistance to root nodules and high compatibility, 18NS (carrying with Pb-Ba8.1 and CRb locus) was used as the donor parent for transferring resistant genes into the pol CMS restorer line 37R. Combined with molecular marker-assisted selection, comprehensive selection of field phenotypic traits, quality analysis, combining ability determination, and resistance identification, a new parent 37RR with double resistance of brassica rapeseed to clubroot disease was developed. Through molecular marker-assisted selection, 8 lines carrying PbBa8.1 + CRb locus were screened in BC3F3 generation. The results of field resistance identification of clubroot disease demonstrated that 8 lines showed immune resistance in Nanchong and Guangyuan. Combining ability of 8 lines was 20S200-2 > 20S199-8 > 20S200-11 > 20S205-3 > 20S200-8 > 20S199-1 > 20S199-7 > 20S205-1 from high to low. The line 20S200-2 was finally identified and designated "37RR" in 2021. Its hybrid 1937R participated in the winter rape variety test in Southwest China, exhibited increased production at 8 points out of 9 test sites, with an average yield of 3204.00 kg•hm-2 per unit area. While gaining resistance to clubroot disease, its yield was also higher than that of the original hybrid Nanyou 1937. [ABSTRACT FROM AUTHOR]