Root Hair Infection Research Articles

New endophytic strains of Trichoderma promote growth and reduce clubroot severity of rapeseed (Brassica napus).

Rapeseed (Brassica napus L.) is the world's third most important edible oilseed crop after soybean and palm. The clubroot disease caused by Plasmodiophora brassicae poses a significant risk and causes substantial yield losses in rapeseed. In this study, 13 endophytic fungal strains were isolated from the healthy roots of rapeseed (B. napus) grown in a clubroot-infested field and molecularly identified. Based on germination inhibition of resting spores of P. brassicae, two endophytic fungal antagonists, Trichoderma spp. ReTk1 and ReTv2 were selected to evaluate their potential for plant growth promotion and biocontrol of P. brassicae. The Trichoderma isolates were applied as a soil drench (1×107 spore/g soil) to a planting mix and field soil, in which plants were grown under non-infested and P. brassicae-infested (2×106 spore/g soil) conditions. The endophytic fungi were able to promote plant growth, significantly increasing shoot and root length, leaf diameter, and biomass production (shoots and root weight) both in the absence or presence of P. brassicae. The single and dual treatments with the endophytes were equally effective in significantly decreasing the root-hair infection, root index, and clubroot severity index. Both ReTk1 and ReTv2 inhibited the germination of resting spores of P. brassicae in root exudates. Moreover, the endophytic fungi colonized the roots of rapeseed extensively and possibly induced host resistance by up-regulated expression of defense-related genes involved in jasmonate (BnOPR2), ethylene (BnACO and BnSAM3), phenylpropanoid (BnOPCL and BnCCR), auxin (BnAAO1) and salicylic acid (BnPR2) pathways. Based on these findings, it is evident that the rapeseed root endophytes Trichoderma spp. ReTk1 and ReTv2 could suppress the gall formation on rapeseed roots via antibiosis, induced systemic resistance (ISR), and/or systemic acquired resistance (SAR). According to our knowledge, this is the first report of the endophytic Trichoderma spp. isolated from root tissues of healthy rapeseed plants (B. napus.), promoting plant growth and reducing clubroot severity.

Role of phytohormones in legumes infected intercellularly by rhizobia without infection threads formation

Rhizobia grow in the soil as free-living organisms but they also can live as nitrogen-fixing symbionts inside root nodules of legumes. Two modes of rhizobial entry into roots are a) intracellularly, through root hairs, inside infection threads, and b) intercellularly, between epidermal cells, never involving root hair or transcellular infection thread formation. Intracellular entry is the best-known infection process, especially in the model legumes Medicago truncatula and Lotus japonicus. Intercellular entry of rhizobia never involving infection thread formation is found in some of the dalbergioid and genistoid legumes, such as Aeschynomene spp. and Arachis hypogaea. This infection mechanism has been scarcely investigated and little is known about molecular players involved in this infection mechanism. Here we focus our interest in comparing the role of phytohormones between legumes infected intracellular and intercellularly. To reach this aim we selected the legumes A. hypogaea and Aeschynomene spp., and the model legumes M. truncatula and L. japonicus.

Development and validation of a novel rapid in vitro assay for determining resistance of potato cultivars to root attachment by Spongospora subterranea zoospores

AbstractSpongospora subterranea f. sp. subterranea is a major pathogen of potatoes leading to losses in tuber quality and yield. Disease can be expressed as root infection, root galling and tuber lesions, the latter known as powdery scab. Attachment of zoospores to potato root hairs is the first step before infection of roots and disease development. Root hair infection results in root dysfunction leading to impaired plant productivity and yield. Varieties vary in their susceptibility to root and tuber disease; however, varietal screening is both time and resource intensive. Furthermore, traditional screens assess root galling or tuber disease and not root infection. In this study, we determined optimal conditions for zoospore release and attachment of zoospores to plant roots and used this information to develop an in vitro bioassay to assess resistance to zoospore root attachment among 153 potato lines and cultivars. Optimal zoospore release occurred at 20°C in Hoagland's solution in a rapid and synchronized manner over the first 2 days, followed by a steep decline. The extent of zoospore root attachment varied with cultivar (Iwa > Agria > Russet Burbank > Gladiator), region of the root maturation zone (lower > middle > upper) and temperature (greatest zoospore root attachment occurring at 15°C). Further comparisons suggested efficiency of zoospore root attachment was also generally associated with known variety resistance to powdery scab, zoosporangial infection and root galling, with a few notable exceptions. The bioassay proved to be a rapid and robust method for screening cultivar resistance to zoospore root attachment.

Molecular Mechanisms of Intercellular Rhizobial Infection: Novel Findings of an Ancient Process.

Establishment of the root-nodule symbiosis in legumes involves rhizobial infection of nodule primordia in the root cortex that is dependent on rhizobia crossing the root epidermal barrier. Two mechanisms have been described: either through root hair infection threads or through the intercellular passage of bacteria. Among the legume genera investigated, around 75% use root hair entry and around 25% the intercellular entry mode. Root-hair infection thread-mediated infection has been extensively studied in the model legumes Medicago truncatula and Lotus japonicus. In contrast, the molecular circuit recruited during intercellular infection, which is presumably an ancient and simpler pathway, remains poorly known. In recent years, important discoveries have been made to better understand the transcriptome response and the genetic components involved in legumes with obligate (Aeschynomene and Arachis spp.) and conditional (Lotus and Sesbania spp.) intercellular rhizobial infections. This review addresses these novel findings and briefly considers possible future research to shed light on the molecular players that orchestrate intercellular infection in legumes.

Spatiotemporal changes in gibberellin content are required for soybean nodulation.

The plant hormone gibberellin (GA) is required at different stages of legume nodule development, with its spatiotemporal distribution tightly regulated. Transcriptomic and bioinformatic analyses established that several key GA biosynthesis and catabolism enzyme encoding genes are critical to soybean (Glycine max) nodule formation. We examined the expression of several GA oxidase genes and used a Förster resonance energy transfer-based GA biosensor to determine the bioactive GA content of roots inoculated with DsRed-labelled Bradyrhizobium diazoefficiens. We manipulated the level of GA by genetically disrupting the expression of GA oxidase genes. Moreover, exogenous treatment of soybean roots with GA3 induced the expression of key nodulation genes and altered infection thread and nodule phenotypes. GmGA20ox1a, GmGA3ox1a, and GmGA2ox1a are upregulated in soybean roots inoculated with compatible B. diazoefficiens. GmGA20ox1a expression is predominately localized to the transient meristem of soybean nodules and coincides with the spatiotemporal distribution of bioactive GA occurring throughout nodule organogenesis. GmGA2ox1a exhibits a nodule vasculature-specific expression pattern, whereas GmGA3ox1a can be detected throughout the nodule and root. Disruptions in the level of GA resulted in aberrant rhizobia infection and reduced nodule numbers. Collectively, our results establish a central role for GAs in root hair infection by symbiotic rhizobia and in nodule organogenesis.

Transcriptome and Coexpression Network Analyses Reveal Hub Genes in Chinese Cabbage (Brassica rapa L. ssp. pekinensis) During Different Stages of Plasmodiophora brassicae Infection.

Clubroot, caused by the soil-borne protist Plasmodiophora brassicae, is one of the most destructive diseases of Chinese cabbage worldwide. However, the clubroot resistance mechanisms remain unclear. In this study, in both clubroot-resistant (DH40R) and clubroot-susceptible (DH199S) Chinese cabbage lines, the primary (root hair infection) and secondary (cortical infection) infection stages started 2 and 5 days after inoculation (dai), respectively. With the extension of the infection time, cortical infection was blocked and complete P. brassica resistance was observed in DH40R, while disease scales of 1, 2, and 3 were observed at 8, 13, and 22 dai in DH199S. Transcriptome analysis at 0, 2, 5, 8, 13, and 22 dai identified 5,750 relative DEGs (rDEGs) between DH40R and DH199S. The results indicated that genes associated with auxin, PR, disease resistance proteins, oxidative stress, and WRKY and MYB transcription factors were involved in clubroot resistance regulation. In addition, weighted gene coexpression network analysis (WGCNA) identified three of the modules whose functions were highly associated with clubroot-resistant, including ten hub genes related to clubroot resistance (ARF2, EDR1, LOX4, NHL3, NHL13, NAC29, two AOP1, EARLI 1, and POD56). These results provide valuable information for better understanding the molecular regulatory mechanism of Chinese cabbage clubroot resistance.

Microrna Regulation of Nodule Zone-Specific Gene Expression In Soybean

Microrna Regulation of Nodule Zone-Specific Gene Expression In Soybean

Exogenous inoculation of endophytic bacterium Bacillus cereus suppresses clubroot (Plasmodiophora brassicae) occurrence in pak choi (Brassica campestris sp. chinensis L.).

The presence of Bacillus cereus plays a key role in clubroot suppression and improves plant biomass in pak choi. B. cereus is reported for the first time as a novel biocontrol agent against clubroot. Plasmodiophora brassicae Woronin causes a devastating infectious disease known as clubroot that is damaging to cruciferous vegetables. This study aimed to isolate beneficial bacteria from the rhizosphere soil of pak choi (Brassica campestris sp. chinensis) and to evaluate the ability of the isolate to reduce the severity of clubroot. Strains obtained from the rhizosphere of symptomless pak choi were first selected on the basis of their germination inhibition rate and effects on the viability of P. brassicae resting spores. Eight bacterial isolates had inhibitory effects against the resting spores of clubroot causing pathogen. However, MZ-12 showed the highest inhibitory effect at 73.4%. Inoculation with MZ-12 enhanced the plant biomass relative to plants grown without MZ-12 as well as P. brassicae infected plants. Furthermore, enhanced antioxidant enzymatic activities were observed in clubroot-infected plants during bacterial association. Co-inoculation of the plant with both P. brassicae and MZ-12 resulted in a 64% reduction of gall formation in comparison to plants inoculated with P. brassicae only. Three applications of MZ-12 to plants infected with P. brassicae at 7, 14 and 21days after seeding (DAS) were more effective than one application and repressed root hair infection. According to 16S rDNA sequence analysis, strain MZ-12 was identified as had a 100% sequence similarity with type strain Bacillus cereus. The findings of the present study will facilitate further investigation into biological mechanisms of cruciferous clubroot control.

Glycine max NNL1 restricts symbiotic compatibility with widely distributed bradyrhizobia via root hair infection.

Symbiosis between soybean (Glycine max) and rhizobia is essential for efficient nitrogen fixation. Rhizobial effectors secreted through the type-III secretion system are key for mediating the interactions between plants and rhizobia, but the molecular mechanism remains largely unknown. Here, our genome-wide association study for nodule number identified G. max Nodule Number Locus 1 (GmNNL1), which encodes a new R protein. GmNNL1 directly interacts with the nodulation outer protein P (NopP) effector from Bradyrhizobium USDA110 to trigger immunity and inhibit nodulation through root hair infection. The insertion of a 179 bp short interspersed nuclear element (SINE)-like transposon into GmNNL1 leads to the loss of function of GmNNL1, enabling bradyrhizobia to successfully nodulate soybeans through the root hair infection route and enhancing nitrogen fixation. Our findings provide important insights into the coevolution of soybean-bradyrhizobia compatibility and offer a way to design new legume-rhizobia interactions for efficient symbiotic nitrogen fixation.

Brassinosteroids play multiple roles in nodulation of pea via interactions with ethylene and auxin.

A comprehensive analysis of the role of brassinosteroids in nodulation, including their interactions with auxin and ethylene revealed thatbrassinosteroids inhibit infection, promote nodule initiation but do not influence nodule organogenesis or function. Nodulation, the symbiosis between legumes and rhizobial bacteria, is regulated by a suite of hormones including brassinosteroids. Previous studies have found that brassinosteroids promote nodule number by inhibiting ethylene biosynthesis. In this study, we examined the influence of brassinosteroids on the various stages of infection and nodule development. We utilise pea mutants, including brassinosteroid mutants lk, lka and lkb, the ethylene insensitive ein2 mutant and the lk ein2 double mutant, along with transgenic lines expressing the DR5::GUS auxin activity marker to investigate how brassinosteroids interact with ethylene and auxin during nodulation. We show that brassinosteroids inhibit the early stages of nodulation, including auxin accumulation, root hair deformation and infection thread formation, and demonstrate that infection thread formation is regulated by brassinosteroids in an ethylene independent manner. In contrast, brassinosteroids appear to act as promoters of nodule initiation through both an ethylene dependent and independent pathway. Although brassinosteroids positively influence the ultimate number of nodules formed, we found that brassinosteroid-deficiency did not influence nodule structure including the vascular pattern of auxin activity or nitrogen-fixation capacity. These findings suggest that brassinosteroids are negative regulators of infection but positive regulators of nodule initiation. Furthermore, brassinosteroids do not appear to be essential for nodule organogenesis or function. Given the influence of brassinosteroids on discreet stages of nodulation but not nodule function, manipulation of brassinosteroids may be an interesting avenue for future research on the optimisation of nodulation.

A Methionine Sulfoxide Reductase B Is Required for the Establishment of Astragalus sinicus-Mesorhizobium Symbiosis.

Methionine sulfoxide reductase B (MsrB) is involved in oxidative stress or defense responses in plants. However, little is known about its role in legume-rhizobium symbiosis. In this study, an MsrB gene was identified from Astragalus sinicus and its function in symbiosis was characterized. AsMsrB was induced under phosphorus starvation and displayed different expression patterns under symbiotic and nonsymbiotic conditions. Hydrogen peroxide or methyl viologen treatment enhanced the transcript level of AsMsrB in roots and nodules. Subcellular localization showed that AsMsrB was localized in the cytoplasm of onion epidermal cells and co-localized with rhizobia in nodules. Plants with AsMsrB-RNAi hairy roots exhibited significant decreases in nodule number, nodule nitrogenase activity and fresh weight of the aerial part, as well as an abnormal nodule and symbiosome development. Statistical analysis of infection events showed that plants with AsMsrB-RNAi hairy roots had significant decreases in the number of root hair curling events, infection threads and nodule primordia compared with the control. The content of hydrogen peroxide increased in AsMsrB-RNAi roots but decreased in AsMsrB overexpression roots at the early stage of infection. The transcriptome analysis showed synergistic modulations of the expression of genes involved in reactive oxygen species generation and scavenging, defense and pathogenesis and early nodulation. In addition, a candidate protein interacting with AsMsrB was identified and confirmed by bimolecular fluorescence complementation. Taken together, our results indicate that AsMsrB plays an essential role in nodule development and symbiotic nitrogen fixation by affecting the redox homeostasis in roots and nodules.

Two New Biocontrol Agents Against Clubroot Caused by Plasmodiophora brassicae.

Clubroot disease caused by Plasmodiophora brassicae can lead to serious yield losses in crucifers such as Brassica napus. In this study, 323 bacterial strains were isolated from the rhizosphere of severely diseased B. napus in Dangyang county, Hubei province, China. Antagonistic strains were first identified based on dual culture inhibition zones with Fusarium oxysporum and Magnaporthe oryzae. These were then further screened in germination inhibition and viability assays of resting spores of P. brassicae. Finally, eight of the antagonistic strains were found to significantly reduce the disease severity of clubroot by more than 40% under greenhouse conditions, and two strains, F85 and T113, were found to have efficacy of more than 80%. Root hair infection experiments showed that F85 and T113 can inhibit early infection of root hairs, reduce the differentiation of primary plasmodia of P. brassicae, and inhibit formation of secondary zoosporangia. Based on sequence analysis of 16S rDNA gene, gyrA gene and 22 housekeeping genes as well as carbon source utilization analysis, the F85 was identified as Bacillus velezensis and T113 as Bacillus amyloliquefaciens. Genome analysis, PCR and RT-PCR detection revealed that both F85 and T113 harbor various antibiotic biosynthesis gene clusters required to form peptides with antimicrobial activity. To our knowledge, this is the first report of B. velezensis as a biocontrol agent against clubroot disease.

The NMN Module Conducts Nodule Number Orchestra.

Legumes control nodule number through nodulation and autoregulation of nodulation (AON) pathways. Nodule Inception (NIN) is essential for rhizobial infection and nodule organogenesis in legumes. The GmNINa-miR172c-NNC1 (NMN) module, which consists of two positive regulators, GmNINa and miR172c, and a suppressor, NNC1, integrates both pathways. GmNINa activates miR172c to downregulate NNC1, leading to nodulation, while NNC1 inhibits miR172c expression, forming a negative feedback loop. GmNINa and NNC1 interact with each other and antagonistically fine-tune GmRIC1/RIC2 expression, turning AON on and off. Conversely, activation of AON inhibits GmNINa and miR172c expression, thereby reducing their inhibitory effects on NNC1 to attenuate both nodulation signaling and AON. The NMN module functions not only as an “accelerator” of the nodulation signal to promote nodulation but also as a “brake” on the signal by activating AON to orchestrate nodule number.

A review on clubroot of crucifers: symptoms, life-cycle of pathogen, factors affecting severity, and management strategies

Clubroot is a devastating disease of crucifers throughout the world. It is caused by a soil-borne obligate phytoparasite, Plasmodiophora brassicae Wor. Plant affected by this disease shows flagging of leaves, unthrifty growth, and even premature death. When uprooted, root shows characteristic symptom of hypertrophied club-shaped root system. Several biotic and abiotic factors affect the disease severity. Biotic factors include spore load in soil and virulence of pathogen, whereas abiotic factors generally include soil environmental factors such as soil temperature, soil pH, soil moisture, and soil type. Pathogen survives, for substantial period of time in absence of host, through its double-walled resting spores in soil or crop debris. Temperature affects spore germination, occurrence, and pathogen proliferation. Acidic soil reaction is crucial for pathogen to proliferate, metabolize, secret enzymes, and to complete life-cycle. All type of soil textures favor disease; however, severity differs with type of soil and soil organic matter content. Soil moisture provides platform to move bi-flagellated zoospores to infect root hairs of crops. Root hair infection is commensurate with inoculum density or spore load in soil. Immediate management strategies entail cultural practices, use of biocontrol agents, and application of chemical as last resort. Trichoderma spp., Pseudomonas fluorescens, Bacillus subtilis, and Gliocladium catenulatum are potential biocontrol agents. Flusalfamide, Fluazinam, and Cyazofamid are some common chemicals used to control clubroot. Soil carried by farm implements, human body, irrigation water, and flood can be potential source of pathogen. The risk of clubroot can be reduced by ensuring phyto-sanitory measures, destroying host crop debris, regular scouting, growing resistant cultivars, avoiding acidic soil reaction, eliminating weedy hosts, and reducing soil movement.

The Rpf84 gene, encoding a ribosomal large subunit protein, RPL22, regulates symbiotic nodulation in Robinia pseudoacacia.

A homologue of the ribosomal protein L22e, Rpf84, regulates root nodule symbiosis by mediating the infection process of rhizobia and preventing bacteroids from degradation in Robinia pseudoacacia. Ribosomal proteins (RPs) are known to have extraribosomal functions, including developmental regulation and stress responses; however, the effects of RPs on symbiotic nodulation of legumes are still unclear. Ribosomal protein 22 of the large 60S subunit (RPL22), a non-typical RP that is only found in eukaryotes, has been shown to function as a tumour suppressor in animals. Here, a homologue of RPL22, Rpf84, was identified from the leguminous tree R. pseudoacacia. Subcellular localization assays showed that Rpf84 was expressed in the cytoplasm and nucleus. Knockdown of Rpf84 by RNA interference (RNAi) technology impaired the infection process and nodule development. Compared with the control, root and stem length, dry weight and nodule number per plant were drastically decreased in Rpf84-RNAi plants. The numbers of root hair curlings, infection threads and nodule primordia were also significantly reduced. Ultrastructure analyses showed that Rpf84-RNAi nodules contained fewer infected cells with fewer bacteria. In particular, remarkable deformation of bacteroids and fusion of multiple symbiosomes occurred in infected cells. By contrast, overexpression of Rpf84 promoted nodulation, and the overexpression nodules maintained a larger infection/differentiation region and had more infected cells filled with bacteroids than the control at 45days post inoculation, suggesting a retarded ageing process in nodules. These results indicate for the first time that RP regulates the symbiotic nodulation of legumes and that RPL22 may function in initiating the invasion of rhizobia and preventing bacteroids from degradation in R. pseudoacacia.

Endosymbiotic Sinorhizobium meliloti modulate Medicago root susceptibility to secondary infection via ethylene.

A complex network of pathways coordinates nodulation and epidermal root hair infection in the symbiotic interaction between rhizobia and legume plants. Whereas nodule formation was known to be autoregulated, it was so far unclear whether a similar control is exerted on the infection process. We assessed the capacity of Medicago plants nodulated by Sinorhizobium meliloti to modulate root susceptibility to secondary bacterial infection or to purified Nod factors in split-root and volatile assays using bacterial and plant mutant combinations. Ethylene implication in this process emerged from gas production measurements, use of a chemical inhibitor of ethylene biosynthesis and of a Medicago mutant affected in ethylene signal transduction. We identified a feedback mechanism that we named AOI (for Autoregulation Of Infection) by which endosymbiotic bacteria control secondary infection thread formation by their rhizospheric peers. AOI involves activation of a cyclic adenosine 3',5'-monophosphate (cAMP) cascade in endosymbiotic bacteria, which decreases both root infectiveness and root susceptibility to bacterial Nod factors. These latter two effects are mediated by ethylene. AOI is a novel component of the complex regulatory network controlling the interaction between Sinorhizobium meliloti and its host plants that emphasizes the implication of endosymbiotic bacteria in fine-tuning the interaction.

Understanding the Resistance Mechanism in Brassica napus to Clubroot Caused by Plasmodiophora brassicae

Exploring the mechanism of plant resistance has become the basis for selection of resistance varieties but reports on revealing resistant mechanism in Brassica napus against Plasmodiophora brassicae are rare. In this study, RNA-seq was conducted in the clubroot-resistant B. napus breeding line ZHE-226 and in the clubroot-susceptible rapeseed cultivar Zhongshuang 11 at 0, 3, 6, 9, and 12 days after inoculation. Strong alteration was detected specifically in ZHE-226 as soon as the root hair infection happened, and significant promotion was found in ZHE-226 on cell division or cell cycle, DNA repair and synthesis, protein synthesis, signaling, antioxidation, and secondary metabolites. Combining results from physiological, biochemical, and histochemical assays, our study highlights an effective signaling in ZHE-226 in response to P. brassicae. This response consists of a fast initiation of receptor kinases by P. brassicae; the possible activation of host intercellular G proteins which might, together with an enhanced Ca2+ signaling, promote the production of reactive oxygen species; and programmed cell death in the host. Meanwhile, a strong ability to maintain homeostasis of auxin and cytokinin in ZHE-226 might effectively limit the formation of clubs on host roots. Our study provides initial insights into resistance mechanism in rapeseed to P. brassicae.

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.

LACK OF SYMBIONT ACCOMMODATION controls intracellular symbiont accommodation in root nodule and arbuscular mycorrhizal symbiosis in Lotus japonicus.

Nitrogen-fixing rhizobia and arbuscular mycorrhizal fungi (AMF) form symbioses with plant roots and these are established by precise regulation of symbiont accommodation within host plant cells. In model legumes such as Lotus japonicus and Medicago truncatula, rhizobia enter into roots through an intracellular invasion system that depends on the formation of a root-hair infection thread (IT). While IT-mediated intracellular rhizobia invasion is thought to be the most evolutionarily derived invasion system, some studies have indicated that a basal intercellular invasion system can replace it when some nodulation-related factors are genetically modified. In addition, intracellular rhizobia accommodation is suggested to have a similar mechanism as AMF accommodation. Nevertheless, our understanding of the underlying genetic mechanisms is incomplete. Here we identify a L. japonicus nodulation-deficient mutant, with a mutation in the LACK OF SYMBIONT ACCOMMODATION (LAN) gene, in which root-hair IT formation is strongly reduced, but intercellular rhizobial invasion eventually results in functional nodule formation. LjLAN encodes a protein that is homologous to Arabidopsis MEDIATOR 2/29/32 possibly acting as a subunit of a Mediator complex, a multiprotein complex required for gene transcription. We also show that LjLAN acts in parallel with a signaling pathway including LjCYCLOPS. In addition, the lan mutation drastically reduces the colonization levels of AMF. Taken together, our data provide a new factor that has a common role in symbiont accommodation process during root nodule and AM symbiosis.

USING γ RAY-INDUCED SILVER NANOPARTICLES ON CONTROL OF CLUBROOT DISEASE OF CHINESE CABBAGE CAUSED BY PLASMODIOPHORA BRASSICAE

Plasmodiophora brassicae- the casual cause of clubroot disease of plants belonging to the family Brassicaceae. This soil-borne fungus infects plants through root hairs. Diseased roots become swollen and deformed. Thus, it’s difficult for plants to absorb water and nutrients. There are some methods used to control this disease including soil fumigation, soil pH control, fungicides, surfactants applications, calcium supplementation, but the results are still restricted. Plasmodiophora brassicae was isolated, identified and artificial inoculated on the Chinese cabbage. The rate of root hair infection and gall formation depended on the spore density and growth media. The increase of the pathogen density lead to the increase of disease symptoms. Silver nanoparticles (SNPs) are widely used as an agent for antifungal treatment. The antimicrobial activity of the nanoparticles varies on the size, shape, and the function of the surface area. The γ - irradiation method to synthesize SNPs is known as a method that can be controlled the out coming sizes, shapes and size distribution of particles. The average diameter of silver nanoparticles in this research was 9.5nm. In the liquid culture and soil, SNPs exhibited a controlling effect on P. brassicae. The fungicide activity depended on the SNP concentrations and the medium; no disease symptoms were found at the SNP concentrations of 1.06 ppm in the liquid culture, and 10 and 20 ppm in soil. Results of the study also showed that the biomass and height of infected plants were much lower than those of the healthy plants. Comparing to the plants that were grown in the sterilized soil, there was not significant difference between the biomass and the height of plants exposed to pathogen and treated with SNPs.