MiR1885 Regulates Disease Tolerance Genes in Brassica rapa during Early Infection with Plasmodiophora brassicae.

Clubroot caused by Plasmodiophora brassicae is a severe disease of cruciferous crops that decreases crop quality and productivity. Several clubroot resistance-related quantitative trait loci and candidate genes have been identified. However, the underlying regulatory mechanism, the interrelationships among genes, and how genes are regulated remain unexplored. MicroRNAs (miRNAs) are attracting attention as regulators of gene expression, including during biotic stress responses. The main objective of this study was to understand how miRNAs regulate clubroot resistance-related genes in P. brassicae-infected Brassica rapa. Two Brassica miRNAs, Bra-miR1885a and Bra-miR1885b, were revealed to target TIR-NBS genes. In non-infected plants, both miRNAs were expressed at low levels to maintain the balance between plant development and basal immunity. However, their expression levels increased in P. brassicae-infected plants. Both miRNAs down-regulated the expression of the TIR-NBS genes Bra019412 and Bra019410, which are located at a clubroot resistance-related quantitative trait locus. The Bra-miR1885-mediated down-regulation of both genes was detected for up to 15 days post-inoculation in the clubroot-resistant line CR Shinki and in the clubroot-susceptible line 94SK. A qRT-PCR analysis revealed Bra019412 expression was negatively regulated by miR1885. Both Bra019412 and Bra019410 were more highly expressed in CR Shinki than in 94SK; the same expression pattern was detected in multiple clubroot-resistant and clubroot-susceptible inbred lines. A 5' rapid amplification of cDNA ends analysis confirmed the cleavage of Bra019412 by Bra-miR1885b. Thus, miR1885s potentially regulate TIR-NBS gene expression during P. brassicae infections of B. rapa.

Quantitative Trait Locus Mapping of Clubroot Resistance and Plasmodiophora brassicae Pathotype Banglim-Specific Marker Development in Brassica rapa.

Clubroot resistance is an economically important trait in Brassicaceae crops. Although many quantitative trait loci (QTLs) for clubroot resistance have been identified in Brassica, disease-related damage continues to occur owing to differences in host variety and constant pathogen variation. Here, we investigated the inheritance of clubroot resistance in a double haploid population developed by crossing clubroot resistant and susceptible lines "09CR500" and "09CR501", respectively. The resistance of "09CR500" to Plasmodiophora brassicae pathotype "Banglim" was controlled as a single dominant gene, with the segregation of resistance and susceptibility being nearly 1:1. PbBrA08Banglim was identified as having a logarithm of odds value of 7.9-74.8, and a phenotypic variance of 26.0-97.1% with flanking marker "09CR.11390652" in A08. After aligning QTL regions to the B. rapa reference genome, 11 genes were selected as candidates. PbBrA08Banglim was located near Crr1, CRs, and Rcr9 loci, but differences were validated by marker analysis, gene structural variations, and gene expression levels, as well as phenotypic responses to the pathotype. Genotyping using the "09CR.11390652" marker accurately distinguished the Banglim-resistance phenotypes in the double haploid population. Thus, the developed marker will be useful in Brassica breeding programs, marker-assisted selection, and gene pyramiding to identify and develop resistant cultivars.

Examining population structure of a bertha armyworm, Mamestra configurata (Lepidoptera: Noctuidae), outbreak in western North America: Implications for gene flow and dispersal.

The bertha armyworm (BAW), Mamestra configurata, is a significant pest of canola (Brassica napus L. and B. rapa L.) in western North America that undergoes cyclical outbreaks every 6-8 years. During peak outbreaks millions of dollars are spent on insecticidal control and, even with control efforts, subsequent damage can result in losses worth millions of dollars. Despite the importance of this pest insect, information is lacking on the dispersal ability of BAW and the genetic variation of populations from across its geographic range which may underlie potential differences in their susceptibility to insecticides or pathogens. Here, we examined the genetic diversity of BAW populations during an outbreak across its geographic range in western North America. First, mitochondrial cytochrome oxidase 1 (CO1) barcode sequences were used to confirm species identification of insects captured in a network of pheromone traps across the range, followed by haplotype analyses. We then sequenced the BAW genome and used double-digest restriction site associated DNA sequencing, mapped to the genome, to identify 1000s of single nucleotide polymorphisms (SNP) markers. CO1 haplotype analysis identified 9 haplotypes distributed across 28 sample locations and three laboratory-reared colonies. Analysis of genotypic data from both the CO1 and SNP markers revealed little population structure across BAW's vast range. The CO1 haplotype pattern showed a star-like phylogeny which is often associated with species whose population abundance and range has recently expanded and combined with pheromone trap data, indicates the outbreak may have originated from a single focal point in central Saskatchewan. The relatively recent introduction of canola and rapid expansion of the canola growing region across western North America, combined with the cyclical outbreaks of BAW caused by precipitous population crashes, has likely selected for a genetically homogenous BAW population adapted to this crop.

Temporal dynamics of bacterial and fungal communities during the infection of Brassica rapa roots by the protist Plasmodiophora brassicae.

The temporal dynamics of rhizosphere and root microbiota composition was compared between healthy and infected Chinese cabbage plants by the pathogen Plasmodiophora brassicae. When inoculated with P. brassicae, disease was measured at five sampling dates from early root hair infection to late gall development. The first symptoms of clubroot disease appeared 14 days after inoculation (DAI) and increased drastically between 14 and 35 DAI. The structure of microbial communities associated to rhizosphere soil and root from healthy and inoculated plants was characterized through high-throughput DNA sequencing of bacterial (16S) and fungal (18S) molecular markers and compared at each sampling date. In healthy plants, Proteobacteria and Bacteroidetes bacterial phyla dominated the rhizosphere and root microbiota of Chinese cabbage. Rhizosphere bacterial communities contained higher abundances of Actinobacteria and Firmicutes compared to the roots. Moreover, a drastic shift of fungal communities of healthy plants occurred between the two last sampling dates, especially in plant roots, where most of Ascomycota fungi dominated until they were replaced by a fungus assigned to the Chytridiomycota phylum. Parasitic invasion by P. brassicae disrupted the rhizosphere and root-associated community assembly at a late step during the root secondary cortical infection stage of clubroot disease. At this stage, Flavisolibacter and Streptomyces in the rhizosphere, and Bacillus in the roots, were drastically less abundant upon parasite invasion. Rhizosphere of plants colonized by P. brassicae was significantly more invaded by the Chytridiomycota fungus, which could reflect a mutualistic relationship in this compartment between these two microorganisms.

Mapping of a novel clubroot resistance QTL using ddRAD-seq in Chinese cabbage (Brassica rapa L.).

BACKGROUND: Plasmodiophora brassicae is a soil-borne plant pathogen that causes clubroot disease, which results in crop yield loss in cultivated Brassica species. Here, we investigated whether a quantitative trait locus (QTL) in B. rapa might confer resistance to a Korean P. brassicae pathotype isolate, Seosan. We crossed resistant and susceptible parental lines and analyzed the segregation pattern in a F2 population of 348 lines. We identified and mapped a novel clubroot resistance QTL using the same mapping population that included susceptible Chinese cabbage and resistant turnip lines. Forty-five resistant and 45 susceptible F2 lines along with their parental lines were used for double digest restriction site-associated DNA sequencing (ddRAD-seq). High resolution melting (HRM)-based validation of SNP positions was conducted to confirm the novel locus. RESULTS: A 3:1 ratio was observed for resistant: susceptible genotypes, which is in accordance with Mendelian segregation. ddRAD-seq identified a new locus, CRs, on chromosome A08 that was different from the clubroot resistance (CR) locus, Crr1. HRM analysis validated SNP positions and constricted CRs region. Four out of seventeen single nucleotide polymorphisms (SNPs) positions were within a 0.8-Mb region that included three NBS-LRR candidate genes but not Crr1. CONCLUSION: The newly identified CRs locus is a novel clubroot resistance locus, as the cultivar Akimeki bears the previously known Crr1 locus but remains susceptible to the Seosan isolate. These results could be exploited to develop molecular markers to detect Seosan-resistant genotypes and develop resistant Chinese cabbage cultivars.

Effects of land use on infestation and parasitism rates of cabbage seed weevil in oilseed rape.

BACKGROUND: This study investigated how infestation rates of an important oilseed rape pest, the cabbage seed weevil (Ceutorhynchus obstrictus) and rates of parasitization by its parasitoids are affected by land use, up to 1000 m from 18 focal fields. RESULTS: The mean proportion of C. obstrictus-infested pods per plant was 8% (2-19.5%). Infestation rates were higher if the adjacent habitat was a herbaceous semi-natural habitat than if it was either another crop or a woody habitat. Infestation rates were positively related to the area of herbaceous semi-natural vegetation, permanent grassland and wheat (which followed oilseed rape in the crop rotation) at a spatial scale of at least 1 km. The mean parasitism rate of C. obstrictus larvae was 55% (8.3-87%), sufficient to provide efficient biocontrol. Parasitism rates were unrelated to adjacent habitats, however, they were positively related to the presence of herbaceous linear elements in the landscape and negatively related to permanent grasslands at a spatial scale of 200 m. CONCLUSION: Proximity of herbaceous elements increased both infestation rates and parasitism, while infestation was also related to landscape factors at larger distances. The findings provide an empirical basis for designing landscapes that suppress C. obstrictus, at both field and landscape scales. © 2018 Society of Chemical Industry.

Mining of Brassica-Specific Genes (BSGs) and Their Induction in Different Developmental Stages and under Plasmodiophora brassicae Stress in Brassica rapa.

Orphan genes, also called lineage-specific genes (LSGs), are important for responses to biotic and abiotic stresses, and are associated with lineage-specific structures and biological functions. To date, there have been no studies investigating gene number, gene features, or gene expression patterns of orphan genes in Brassica rapa. In this study, 1540 Brassica-specific genes (BSGs) and 1824 Cruciferae-specific genes (CSGs) were identified based on the genome of Brassica rapa. The genic features analysis indicated that BSGs and CSGs possessed a lower percentage of multi-exon genes, higher GC content, and shorter gene length than evolutionary-conserved genes (ECGs). In addition, five types of BSGs were obtained and 145 out of 529 real A subgenome-specific BSGs were verified by PCR in 51 species. In silico and semi-qPCR, gene expression analysis of BSGs suggested that BSGs are expressed in various tissue and can be induced by Plasmodiophora brassicae. Moreover, an A/C subgenome-specific BSG, BSGs1, was specifically expressed during the heading stage, indicating that the gene might be associated with leafy head formation. Our results provide valuable biological information for studying the molecular function of BSGs for Brassica-specific phenotypes and biotic stress in B. rapa.

Effects of litter feeders on the transfer of 137Cs to plants.

The effects of the Japanese horned beetle larvae on the transfer of 137Cs from a contaminated leaf litter to the leaf vegetable, komatsuna (Brassica rapa var. perviridis) was studied. Feces of the larvae which were fed 137Cs-contaminated leaf litter were added to a potting mix in which komatsuna plants were cultivated. The presence of feces increased the harvest yield of komatsuna, suggesting that feces provided nutrients for the plant growth. In addition, the amount of exchangeable 137Cs in leaf litter was experimentally confirmed to be enhanced by the presence of feces which were excreted by larvae feeding. However, there was no difference in the soil-to-plant transfer factor of 137Cs for the presence and absence of feces. Interactions between clay minerals and exchangeable 137Cs in the soil beneath the litter layer may diminish the root uptake of 137Cs. From these results, it was concluded that the effect of exchangeable 137Cs released from feces was limited for the transfer of 137Cs to plants if plant roots were not present in litter layers.

Genome-wide identification and expression analysis of chitinase gene family in Brassica rapa reveals its role in clubroot resistance.

Chitinases, a category of pathogenesis-related proteins, are responsible for catalyzing the hydrolysis of chitin into the N-acetyl-d-glucosamine. Therefore, chitinases are believed to function as a guardian against chitin-containing pathogens. Here, we examined the role of the Brassica rapa chitinase family genes in clubroot disease. A total of 33 chitinase genes were identified and grouped into five classes based on their conserved domain. They were distributed unevenly across eight chromosomes in B. rapa, and 31 of them contained few introns (≤2). In addition, the expression of these genes was organ-specific, and 14 genes were expressed differentially in response to Plasmodiophora brassicae challenge of clubroot-susceptible (CS NIL) and resistant (CR NIL) lines. Furthermore, reduced pathogen DNA content and clubroot symptoms were observed in the CS NILs after their treatment with chitin oligosaccharides 24 h prior to inoculation with P. brassicae. The findings indicate that chitinases play a crucial role in pathogen resistance of the host plants. The results offer an insight into the role of chitinase in B. rapa-P. brassicae interaction.

Trans-generational inheritance of herbivory-induced phenotypic changes in Brassica rapa.

Biotic stress can induce plastic changes in fitness-relevant plant traits. Recently, it has been shown that such changes can be transmitted to subsequent generations. However, the occurrence and extent of transmission across different types of traits is still unexplored. Here, we assessed the emergence and transmission of herbivory-induced changes in Brassica rapa and their impact on interactions with insects. We analysed changes in morphology and reproductive traits as well as in flower and leaf volatile emission during two generations with leaf herbivory by Mamestra brassicae and Pieris brassicae and two subsequent generations without herbivory. Herbivory induced changes in all trait types, increasing attractiveness of the plants to the parasitoid wasp Cotesia glomerata and decreasing visitation by the pollinator Bombus terrestris, a potential trade-off. While changes in floral and leaf volatiles disappeared in the first generation after herbivory, some changes in morphology and reproductive traits were still measurable two generations after herbivory. However, neither parasitoids nor pollinators further discriminated between groups with different past treatments. Our results suggest that transmission of herbivore-induced changes occurs preferentially in resource-limited traits connected to plant growth and reproduction. The lack of alterations in plant-insect interactions was likely due to the transient nature of volatile changes.

Genotyping-by-sequencing reveals three QTL for clubroot resistance to six pathotypes of Plasmodiophora brassicae in Brassica rapa.

Clubroot, caused by Plasmodiophora brassicae, is an important disease of Brassica crops worldwide. F1 progeny from the Brassica rapa lines T19 (resistant) × ACDC (susceptible) were backcrossed with ACDC, then self-pollinated to produce BC1S1 lines, From genotyping-by-sequencing (GBS) of the parental lines and BC1 plants, about 1.32 M sequences from T19 were aligned into the reference genome of B. rapa with 0.4-fold coverage, and 1.77 M sequences with 0.5-fold coverage in ACDC. The number of aligned short reads per plant in the BC1 ranged from 0.07 to 1.41 M sequences with 0.1-fold coverage. A total of 1584 high quality SNP loci were obtained, distributed on 10 chromosomes. A single co-localized QTL, designated as Rcr4 on chromosome A03, conferred resistance to pathotypes 2, 3, 5, 6 and 8. The peak was at SNP locus A03_23710236, where LOD values were 30.3 to 38.8, with phenotypic variation explained (PVE) of 85-95%. Two QTLs for resistance to a novel P. brassicae pathotype 5x, designated Rcr8 on chromosome A02 and Rcr9 on A08, were detected with 15.0 LOD and 15.8 LOD, and PVE of 36% and 39%, respectively. Bulked segregant analysis was performed to examine TIR-NBS-LRR proteins in the regions harboring the QTL.

Detection of Ribosomal DNA Sequence Polymorphisms in the Protist Plasmodiophora brassicae for the Identification of Geographical Isolates.

Clubroot is a soil-borne disease caused by the protist Plasmodiophora brassicae (P. brassicae). It is one of the most economically important diseases of Brassica rapa and other cruciferous crops as it can cause remarkable yield reductions. Understanding P. brassicae genetics, and developing efficient molecular markers, is essential for effective detection of harmful races of this pathogen. Samples from 11 Korean field populations of P. brassicae (geographic isolates), collected from nine different locations in South Korea, were used in this study. Genomic DNA was extracted from the clubroot-infected samples to sequence the ribosomal DNA. Primers and probes for P. brassicae were designed using a ribosomal DNA gene sequence from a Japanese strain available in GenBank (accession number AB526843; isolate NGY). The nuclear ribosomal DNA (rDNA) sequence of P. brassicae, comprising 6932 base pairs (bp), was cloned and sequenced and found to include the small subunits (SSUs) and a large subunit (LSU), internal transcribed spacers (ITS1 and ITS2), and a 5.8s. Sequence variation was observed in both the SSU and LSU. Four markers showed useful differences in high-resolution melting analysis to identify nucleotide polymorphisms including single- nucleotide polymorphisms (SNPs), oligonucleotide polymorphisms, and insertions/deletions (InDels). A combination of three markers was able to distinguish the geographical isolates into two groups.

Use of an individual-based simulation model to explore and evaluate potential insecticide resistance management strategies.

BACKGROUND: Tools with the potential to predict risks of insecticide resistance and aid the evaluation and design of resistance management tactics are of value to all sectors of the pest management community. Here we describe use of a versatile individual-based model of resistance evolution to simulate how strategies employing single and multiple insecticides influence resistance development in the pollen beetle, Meligethes aeneus. RESULTS: Under repeated exposure to a single insecticide, resistance evolved faster to a pyrethroid (lambda-cyhalothrin) than to a pyridine azomethane (pymetrozine), due to difference in initial efficacy. A mixture of these compounds delayed resistance compared to use of single products. The effectiveness of rotations depended on the sequence in which compounds were applied in response to pest density thresholds. Effectiveness of a mixture strategy declined with reductions in grower compliance. At least 50% compliance was needed to cause some delay in resistance development. CONCLUSION: No single strategy meets all requirements for managing resistance. It is important to evaluate factors that prevail under particular pest management scenarios. The model used here provides operators with a valuable means for evaluating and extending sound resistance management advice, as well as understanding needs and opportunities offered by new control techniques. © 2016 Society of Chemical Industry.

The tandem repeated organization of NB-LRR genes in the clubroot-resistant CRb locus in Brassica rapa L.

To facilitate prevention of clubroot disease, a major threat to the successful cultivation of Chinese cabbage (Brassica rapa L.), we bred clubroot-resistant (CR) cultivars by introducing resistance genes from CR turnips via conventional breeding. Among 11 CR loci found in B. rapa, we identified CRb in Chinese cabbage cultivar 'CR Shinki' as a single dominant gene for resistance against Plasmodiophora brassicae pathotype group 3, against which the stacking of Crr1 and Crr2 loci was not effective. However, the precise location and pathotype specificity of CRb have been controversial, because CRa and Rcr1 also map near this locus. Previously, our fine-mapping study revealed that CRb is located in a 140-kb genomic region on chromosome A03. Here, we determined the nucleotide sequence of an approximately 64-kb candidate region in the resistant line; this region contains six open reading frames (ORFs) similar to NB-LRR encoding genes that are predicted to occur in tandem with the same orientation. Among the six ORFs present, only four on the genome of the resistant line showed a strong DNA sequence identity with each other, and only one of those four could confer resistance to P. brassicae isolate No. 14 of the pathotype group 3. These results suggest that these genes evolved through recent gene duplication and uneven crossover events that could lead to the acquisition of clubroot resistance. The DNA sequence of the functional ORF was identical to that of the previously cloned CRa gene; thus, we showed that the independently identified CRb and CRa are one and the same clubroot-resistance gene.

Impacts of neonicotinoid use on long-term population changes in wild bees in England.

Wild bee declines have been ascribed in part to neonicotinoid insecticides. While short-term laboratory studies on commercially bred species (principally honeybees and bumblebees) have identified sub-lethal effects, there is no strong evidence linking these insecticides to losses of the majority of wild bee species. We relate 18 years of UK national wild bee distribution data for 62 species to amounts of neonicotinoid use in oilseed rape. Using a multi-species dynamic Bayesian occupancy analysis, we find evidence of increased population extinction rates in response to neonicotinoid seed treatment use on oilseed rape. Species foraging on oilseed rape benefit from the cover of this crop, but were on average three times more negatively affected by exposure to neonicotinoids than non-crop foragers. Our results suggest that sub-lethal effects of neonicotinoids could scale up to cause losses of bee biodiversity. Restrictions on neonicotinoid use may reduce population declines.

Identification of Genome-Wide Variants and Discovery of Variants Associated with Brassica rapa Clubroot Resistance Gene Rcr1 through Bulked Segregant RNA Sequencing.

Clubroot, caused by Plasmodiophora brassicae, is an important disease on Brassica species worldwide. A clubroot resistance gene, Rcr1, with efficacy against pathotype 3 of P. brassicae, was previously mapped to chromosome A03 of B. rapa in pak choy cultivar "Flower Nabana". In the current study, resistance to pathotypes 2, 5 and 6 was shown to be associated with Rcr1 region on chromosome A03. Bulked segregant RNA sequencing was performed and short read sequences were assembled into 10 chromosomes of the B. rapa reference genome v1.5. For the resistant (R) bulks, a total of 351.8 million (M) sequences, 30,836.5 million bases (Mb) in length, produced 120-fold coverage of the reference genome. For the susceptible (S) bulks, 322.9 M sequences, 28,216.6 Mb in length, produced 109-fold coverage. In total, 776.2 K single nucleotide polymorphisms (SNPs) and 122.2 K insertion / deletion (InDels) in R bulks and 762.8 K SNPs and 118.7 K InDels in S bulks were identified; each chromosome had about 87% SNPs and 13% InDels, with 78% monomorphic and 22% polymorphic variants between the R and S bulks. Polymorphic variants on each chromosome were usually below 23%, but made up 34% of the variants on chromosome A03. There were 35 genes annotated in the Rcr1 target region and variants were identified in 21 genes. The numbers of poly variants differed significantly among the genes. Four out of them encode Toll-Interleukin-1 receptor / nucleotide-binding site / leucine-rich-repeat proteins; Bra019409 and Bra019410 harbored the higher numbers of polymorphic variants, which indicates that they are more likely candidates of Rcr1. Fourteen SNP markers in the target region were genotyped using the Kompetitive Allele Specific PCR method and were confirmed to associate with Rcr1. Selected SNP markers were analyzed with 26 recombinants obtained from a segregating population consisting of 1587 plants, indicating that they were completely linked to Rcr1. Nine SNP markers were used for marker-assisted introgression of Rcr1 into B. napus canola from B. rapa, with 100% accuracy in this study.

Evidence for pollinator cost and farming benefits of neonicotinoid seed coatings on oilseed rape.

Chronic exposure to neonicotinoid insecticides has been linked to reduced survival of pollinating insects at both the individual and colony level, but so far only experimentally. Analyses of large-scale datasets to investigate the real-world links between the use of neonicotinoids and pollinator mortality are lacking. Moreover, the impacts of neonicotinoid seed coatings in reducing subsequent applications of foliar insecticide sprays and increasing crop yield are not known, despite the supposed benefits of this practice driving widespread use. Here, we combine large-scale pesticide usage and yield observations from oilseed rape with those detailing honey bee colony losses over an 11 year period, and reveal a correlation between honey bee colony losses and national-scale imidacloprid (a neonicotinoid) usage patterns across England and Wales. We also provide the first evidence that farmers who use neonicotinoid seed coatings reduce the number of subsequent applications of foliar insecticide sprays and may derive an economic return. Our results inform the societal discussion on the pollinator costs and farming benefits of prophylactic neonicotinoid usage on a mass flowering crop.

Assessing climate change impacts on the rape stem weevil, Ceutorhynchus napi Gyll., based on bias- and non-bias-corrected regional climate change projections.

Agricultural production is directly affected by projected increases in air temperature and changes in precipitation. A multi-model ensemble of regional climate change projections indicated shifts towards higher air temperatures and changing precipitation patterns during the summer and winter seasons up to the year 2100 for the region of Goettingen (Lower Saxony, Germany). A second major controlling factor of the agricultural production is the infestation level by pests. Based on long-term field surveys and meteorological observations, a calibration of an existing model describing the migration of the pest insect Ceutorhynchus napi was possible. To assess the impacts of climate on pests under projected changing environmental conditions, we combined the results of regional climate models with the phenological model to describe the crop invasion of this species. In order to reduce systematic differences between the output of the regional climate models and observational data sets, two different bias correction methods were applied: a linear correction for air temperature and a quantile mapping approach for precipitation. Only the results derived from the bias-corrected output of the regional climate models showed satisfying results. An earlier onset, as well as a prolongation of the possible time window for the immigration of Ceutorhynchus napi, was projected by the majority of the ensemble members.

Heteroconium chaetospira induces resistance to clubroot via upregulation of host genes involved in jasmonic acid, ethylene, and auxin biosynthesis.

An endophytic fungus, Heteroconium chaetospira isolate BC2HB1 (Hc), suppressed clubroot (Plasmodiophora brassicae -Pb) on canola in growth-cabinet trials. Confocal microscopy demonstrated that Hc penetrated canola roots and colonized cortical tissues. Based on qPCR analysis, the amount of Hc DNA found in canola roots at 14 days after treatment was negatively correlated (r = 0.92, P<0.001) with the severity of clubroot at 5 weeks after treatment at a low (2×10(5) spores pot(-1)) but not high (2×10(5) spores pot(-1)) dose of pathogen inoculum. Transcript levels of nine B. napus (Bn) genes in roots treated with Hc plus Pb, Pb alone and a nontreated control were analyzed using qPCR supplemented with biochemical analysis for the activity of phenylalanine ammonia lyases (PAL). These genes encode enzymes involved in several biosynthetic pathways related potentially to plant defence. Hc plus Pb increased the activity of PAL but not that of the other two genes (BnCCR and BnOPCL) involved also in phenylpropanoid biosynthesis, relative to Pb inoculation alone. In contrast, expression of several genes involved in the jasmonic acid (BnOPR2), ethylene (BnACO), auxin (BnAAO1), and PR-2 protein (BnPR-2) biosynthesis were upregulated by 63, 48, 3, and 3 fold, respectively, by Hc plus Pb over Pb alone. This indicates that these genes may be involved in inducing resistance in canola by Hc against clubroot. The upregulation of BnAAO1 appears to be related to both pathogenesis of clubroot and induced defence mechanisms in canola roots. This is the first report on regulation of specific host genes involved in induced plant resistance by a non-mycorrhizal endophyte.

Disarming the jasmonate-dependent plant defense makes nonhost Arabidopsis plants accessible to the American serpentine leafminer.

Here, we analyzed the interaction between Arabidopsis (Arabidopsis thaliana) and the American serpentine leafminer (Liriomyza trifolii), an important and intractable herbivore of many cultivated plants. We examined the role of the immunity-related plant hormone jasmonate (JA) in the plant response and resistance to leafminer feeding to determine whether JA affects host suitability for leafminers. The expression of marker genes for the JA-dependent plant defense was induced by leafminer feeding on Arabidopsis wild-type plants. Analyses of JA-insensitive coi1-1 mutants suggested the importance of JA in the plant response to leafminer feeding. The JA content of wild-type plants significantly increased after leafminer feeding. Moreover, coi1-1 mutants showed lower feeding resistance against leafminer attack than did wild-type plants. The number of feeding scars caused by inoculated adult leafminers in JA-insensitive coi1-1 mutants was higher than that in wild-type plants. In addition, adults of the following generation appeared only from coi1-1 mutants and not from wild-type plants, suggesting that the loss of the JA-dependent plant defense converted nonhost plants to accessible host plants. Interestingly, the glucosinolate-myrosinase defense system may play at most a minor role in this conversion, indicating that this major antiherbivore defense of Brassica species plants probably does not have a major function in plant resistance to leafminer. Application of JA to wild-type plants before leafminer feeding enhanced feeding resistance in Chinese cabbage (Brassica rapa), tomato (Solanum lycopersicum), and garland chrysanthemum (Chrysanthemum coronarium). Our results indicate that JA plays an important role in the plant response and resistance to leafminers and, in so doing, affects host plant suitability for leafminers.