Transcriptional Changes in Potato Sprouts upon Interaction with Rhizoctonia solani Indicate Pathogen-Induced Interference in the Defence Pathways of Potato.

Rhizoctonia solani is the causer of black scurf disease on potatoes and is responsible for high economical losses in global agriculture. In order to increase the limited knowledge of the plants' molecular response to this pathogen, we inoculated potatoes with R. solani AG3-PT isolate Ben3 and carried out RNA sequencing with total RNA extracted from potato sprouts at three and eight days post inoculation (dpi). In this dual RNA-sequencing experiment, the necrotrophic lifestyle of R. solani AG3-PT during early phases of interaction with its host has already been characterised. Here the potato plants' comprehensive transcriptional response to inoculation with R. solani AG3 was evaluated for the first time based on significantly different expressed plant genes extracted with DESeq analysis. Overall, 1640 genes were differentially expressed, comparing control (-Rs) and with R. solani AG3-PT isolate Ben3 inoculated plants (+Rs). Genes involved in the production of anti-fungal proteins and secondary metabolites with antifungal properties were significantly up regulated upon inoculation with R. solani. Gene ontology (GO) terms involved in the regulation of hormone levels (i.e., ethylene (ET) and jasmonic acid (JA) at 3 dpi and salicylic acid (SA) and JA response pathways at 8 dpi) were significantly enriched. Contrastingly, the GO term "response to abiotic stimulus" was down regulated at both time points analysed. These results may support future breeding efforts toward the development of cultivars with higher resistance level to black scurf disease or the development of new control strategies.

Salmonella persistence in soil depends on reciprocal interactions with indigenous microorganisms.

Fresh fruits and vegetables have numerous benefits to human health. Unfortunately, their consumption is increasingly associated with food-borne diseases, Salmonella enterica being their most frequent cause in Europe. Agricultural soils were postulated as reservoir of human pathogens, contributing to the contamination of crops during the growing period. Since the competition with the indigenous soil microbiota for colonization sites plays a major role in the success of invading species, we hypothesized that reduced diversity will enhance the chance of Salmonella to successfully establish in agricultural environments. We demonstrated that the abundance of Salmonella drastically decreased in soil with highly diverse indigenous prokaryotic community, while in soil with reduced prokaryotic diversity, Salmonella persisted for a long period. Furthermore, in communities with low diversity, Salmonella had an impact on the abundance of other taxa. The high physiological plasticity allows Salmonella to use agricultural soils as alternative habitat which might provide a route of animal/human infections. In addition, adjusted transcriptional profile with amino acid biosynthesis and the glyoxylate cycle most prominently regulated, suggests an adaptation to the soil environment. Our results underline the importance of the maintenance of diverse soil microbiome as a part of strategy aiming at reduced risk of food-borne salmonellosis outbreaks.

Effect of long-term organic and mineral fertilization strategies on rhizosphere microbiota assemblage and performance of lettuce.

Long-term agricultural fertilization strategies gradually change soil properties including the associated microbial communities. Cultivated crops recruit beneficial microbes from the surrounding soil environment via root exudates. In this study, we aimed to investigate the effects of long-term fertilization strategies across field sites on the rhizosphere prokaryotic (Bacteria and Archaea) community composition and plant performance. We conducted growth chamber experiments with lettuce (Lactuca sativa L.) cultivated in soils from two long-term field experiments, each of which compared organic versus mineral fertilization strategies. 16S rRNA gene amplicon sequencing revealed the assemblage of a rhizosphere core microbiota shared in all lettuce plants across soils, going beyond differences in community composition depending on field site and fertilization strategies. The enhanced expression of several plant genes with roles in oxidative and biotic stress signalling pathways in lettuce grown in soils with organic indicates an induced physiological status in plants. Lettuce plants grown in soils with different fertilization histories were visibly free of stress symptoms and achieved comparable biomass. This suggests a positive aboveground plant response to belowground plant-microbe interactions in the rhizosphere. Besides effects of fertilization strategy and field site, our results demonstrate the crucial role of the plant in driving rhizosphere microbiota assemblage.

The Use of Features from Fluorescence, Thermography, and NDVI Imaging to Detect Biotic Stress in Lettuce.

Fluorescence, normalized difference vegetation index, and thermal imaging are three frequently used nondestructive methods to detect biotic stress in plants. Due, in part, to the inconsistent results reported in the literature and the lack of measurements on the whole-plant scale, we tested the suitability of a wide variety of variables obtained using these three imaging methods to classify young plants into biotically stressed and nonstressed plants. To this end, we applied the model plant-pathogen system lettuce-Rhizoctonia solani. The relevant data from each image and plant (healthy and diseased) was extracted semiautomatically using sophisticated image processing algorithms. This method enabled us to identify the most appropriate variables via discriminant function and logistic regression analysis: photosystem II maximum quantum yield (Fv/Fm) and fluorescence decline ratio can be used to classify variables with an error ≤0.052. Lettuce seedlings with an Fv/Fm ratio > 0.73 were consistently healthy. In some cases, it was possible to detect infection prior to the appearance of symptoms. Possibilities to transfer the method to horticultural practice are discussed.

Draft genome sequence of the potato pathogen Rhizoctonia solani AG3-PT isolate Ben3.

The basidiomycetes fungus Rhizoctonia solani AG3 is responsible for black scurf disease on potato and occurs in each potato growing area world-wide. In this study, the draft genome sequence of the black scurf pathogen R. solani AG3-PT isolate Ben3 is presented. The genome sequence of R. solani AG3-PT isolate Ben3 consists of 1385 scaffolds. These scaffolds amount to a size of approx. 51 Mb. Considering coverage analyses of contigs, the size of the diploid genome was estimated to correspond to 116 Mb. Gene prediction by applying AUGUSTUS (3.2.1.) resulted in 12,567 identified genes. Based on automatic annotation using GenDBE, genes potentially encoding cellulases and enzymes involved in secondary metabolite synthesis were identified in the R. solani AG3-PT isolate Ben3 genome. Comparative analyses including the R. solani AG3 isolate Rhs1AP, also originating from potato, revealed first insights into core genes shared by both isolates and unique determinants of each isolate.

Cyclic Lipopeptides of Bacillus amyloliquefaciens subsp. plantarum Colonizing the Lettuce Rhizosphere Enhance Plant Defense Responses Toward the Bottom Rot Pathogen Rhizoctonia solani.

The commercially available inoculant Bacillus amyloliquefaciens FZB42 is able to considerably reduce lettuce bottom rot caused by Rhizoctonia solani. To understand the interaction between FZB42 and R. solani in the rhizosphere of lettuce, we used an axenic system with lettuce bacterized with FZB42 and inoculated with R. solani. Confocal laser scanning microscopy showed that FZB42 could delay the initial establishment of R. solani on the plants. To show which secondary metabolites of FZB42 are produced under these in-situ conditions, we developed an ultra-high performance liquid chromatography coupled to time of flight mass spectrometry-based method and identified surfactin, fengycin, and bacillomycin D in the lettuce rhizosphere. We hypothesized that lipopeptides and polyketides play a role in enhancing the plant defense responses in addition to the direct antagonistic effect toward R. solani and used a quantitative real-time polymerase chain reaction-based assay for marker genes involved in defense signaling pathways in lettuce. A significant higher expression of PDF 1.2 observed in the bacterized plants in response to subsequent pathogen challenge showed that FZB42 could enhance the lettuce defense response toward the fungal pathogen. To identify if surfactin or other nonribosomally synthesized secondary metabolites could elicit the observed enhanced defense gene expression, we examined two mutants of FZB42 deficient in production of surfactin and the lipopetides and polyketides, by expression analysis and pot experiments. In the absence of surfactin and other nonribosomally synthesized secondary metabolites, there was no enhanced PDF 1.2-mediated response to the pathogen challenge. Pot experiment results showed that the mutants failed to reduce disease incidence in lettuce as compared with the FZB42 wild type, indicating, that surfactin as well as other nonribosomally synthesized secondary metabolites play a role in the actual disease suppression and on lettuce health. In conclusion, our study showed that nonribosomally synthesized secondary metabolites of FZB42 are actually produced in the lettuce rhizosphere and contribute to the disease suppression by mediating plant defense gene expression toward the pathogen R. solani.

Long-term conservation tillage with reduced nitrogen fertilization intensity can improve winter wheat health via positive plant-microorganism feedback in the rhizosphere.

Microbiome-based solutions are regarded key for sustainable agroecosystems. However, it is unclear how agricultural practices affect the rhizosphere microbiome, plant-microorganism interactions and crop performance under field conditions. Therefore, we installed root observation windows in a winter wheat field cultivated either under long-term mouldboard plough (MP) or cultivator tillage (CT). Each tillage practice was also compared at two nitrogen (N) fertilization intensities, intensive (recommended N-supply with pesticides/growth regulators) or extensive (reduced N-supply, no fungicides/growth regulators). Shoot biomass, root exudates and rhizosphere metabolites, physiological stress indicators, and gene expression were analyzed together with the rhizosphere microbiome (bacterial/archaeal 16S rRNA gene, fungal ITS amplicon, and shotgun metagenome sequencing) shortly before flowering. Compared to MP, the rhizosphere of CT winter wheat contained more primary and secondary metabolites, especially benzoxazinoid derivatives. Potential copiotrophic and plant-beneficial taxa (e.g. Bacillus, Devosia, and Trichoderma) as well as functional genes (e.g. siderophore production, trehalose synthase, and ACC deaminase) were enriched in the CT rhizosphere, suggesting that tillage affected belowground plant-microorganism interactions. In addition, physiological stress markers were suppressed in CT winter wheat compared to MP. In summary, tillage practice was a major driver of crop performance, root deposits, and rhizosphere microbiome interactions, while the N-fertilization intensity was also relevant, but less important.

Does Constitutive Expression of Defense-Related Genes and Salicylic Acid Concentrations Correlate with Field Resistance of Potato to Black Scurf Disease?

Black scurf disease on potato caused by Rhizoctonia solani AG3 occurs worldwide and is difficult to control. The use of potato cultivars resistant to black scurf disease could be part of an integrated control strategy. Currently, the degree of resistance is based on symptom assessment in the field, but molecular measures could provide a more efficient screening method. We hypothesized that the degree of field resistance to black scurf disease in potato cultivars is associated with defense-related gene expression levels and salicylic acid (SA) concentration. Cultivars with a moderate and severe appearance of disease symptoms on tubers were selected and cultivated in the same field. In addition, experiments were conducted under controlled conditions in an axenic in vitro culture and in a sand culture to analyze the constitutive expression of defense-related genes and SA concentration. The more resistant cultivars did not show significantly higher constitutive expression levels of defense-related genes. Moreover, the level of free SA was increased in the more resistant cultivars only in the roots of the plantlets grown in the sand culture. These results indicate that neither expression levels of defense-related genes nor the amount of SA in potato plants can be used as reliable predictors of the field resistance of potato genotypes to black scurf disease.

Monitoring of an Applied Beneficial Trichoderma Strain in Root-Associated Soil of Field-Grown Maize by MALDI-TOF MS.

The persistence of beneficial microorganisms in the rhizosphere or surrounding soil following their application is a prerequisite for the effective interaction with the plant or indigenous microbial communities in the respective habitats. The goal of the study was to analyze the establishment and persistence of the applied beneficial Trichoderma harzianum (OMG16) strain in the maize root-associated soil depending on agricultural practice (soil management practice, N-fertilizer intensity) in a field experiment. A rapid identification of the inoculated strain OMG16 is essential for its monitoring. We used a culture-based approach coupled to matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis for the rapid identification of the inoculated Trichoderma strain as part of the beneficial microbe consortium (BMc). We isolated 428 fungal isolates from eight treatments of the field experiment. Forty eight percent of the isolated fungi equivalent to 205 fungal isolates were identified as Trichoderma, of which 87% (=179 isolates) were obtained from the fields inoculated with BMc. Gene sequence analysis showed a high similarity of the MALDI-TOF MS-identified Trichoderma, with that of the inoculated Trichoderma harzianum OMG16 confirming the re-isolation of the added beneficial fungus. This study highlighted the use of MALDI-TOF MS analysis as a quick, cost-effective detection and efficient monitoring tool for microbial-based bioinoculants in the field.

Beneficial microbial consortium improves winter rye performance by modulating bacterial communities in the rhizosphere and enhancing plant nutrient acquisition.

The beneficial effect of microbial consortium application on plants is strongly affected by soil conditions, which are influenced by farming practices. The establishment of microbial inoculants in the rhizosphere is a prerequisite for successful plant-microorganism interactions. This study investigated whether a consortium of beneficial microorganisms establishes in the rhizosphere of a winter crop during the vegetation period, including the winter growing season. In addition, we aimed for a better understanding of its effect on plant performance under different farming practices. Winter rye plants grown in a long-time field trial under conventional or organic farming practices were inoculated after plant emergence in autumn with a microbial consortium containing Pseudomonas sp. (RU47), Bacillus atrophaeus (ABi03) and Trichoderma harzianum (OMG16). The density of the microbial inoculants in the rhizosphere and root-associated soil was quantified in autumn and the following spring. Furthermore, the influence of the consortium on plant performance and on the rhizosphere bacterial community assembly was investigated using a multidisciplinary approach. Selective plating showed a high colonization density of individual microorganisms of the consortium in the rhizosphere and root-associated soil of winter rye throughout its early growth cycle. 16S rRNA gene amplicon sequencing showed that the farming practice affected mainly the rhizosphere bacterial communities in autumn and spring. However, the microbial consortium inoculated altered also the bacterial community composition at each sampling time point, especially at the beginning of the new growing season in spring. Inoculation of winter rye with the microbial consortium significantly improved the plant nutrient status and performance especially under organic farming. In summary, the microbial consortium showed sufficient efficacy throughout vegetation dormancy when inoculated in autumn and contributed to better plant performance, indicating the potential of microbe-based solutions in organic farming where nutrient availability is limited.

Influence of sewage sludge stabilization method on microbial community and the abundance of antibiotic resistance genes.

Municipal sewage sludge (MSS) and other biosolids are of high interest for agriculture. These nutrient-rich organic materials can potentially serve as organic fertilizers. Besides an increase of organic matter in soil, other positive effects were shown after their application. Especially the positive influence on circular economy increased the attention paid to management of MSS in recent years. Unfortunately, the use of sewage sludge has some drawbacks. Biosolids are frequently polluted with heavy metals, xenobiotic organic compounds and industrial chemicals, which may be hazardous for the environment and humans. Here, we investigated the influence of stabilization method and the size of wastewater treatment plant on the structure of microbial communities as well as the abundance of antibiotic resistance genes (ARG) and mobile genetic elements (MGE). All tested ARG and MGE were detectable in almost all of the samples. Interestingly, the presence of MGE as well as particular heavy metals correlated positively with the presence of several ARG. We conclude that the distribution of ARG and MGE in biosolids originated from municipal wastewater treatment plants, cannot be explained by the size of the facility or the applied stabilization method. Moreover, we postulate that the presence of pollutants and long-term impacts should be assessed prior to a possible use of sewage sludge as fertilizer.

What Drives the Assembly of Plant-associated Protist Microbiomes? Investigating the Effects of Crop Species, Soil Type and Bacterial Microbiomes.

In a field experiment we investigated the influence of the environmental filters soil type (i.e. three contrasting soils) and plant species (i.e. lettuce and potato) identity on rhizosphere community assembly of Cercozoa, a dominant group of mostly bacterivorous soil protists. Plant species (14%) and rhizosphere origin (vs bulk soil) with 13%, together explained four times more variation in cercozoan beta diversity than the three soil types (7% explained variation). Our results clearly confirm the existence of plant species-specific protist communities. Network analyses of bacteria-Cercozoa rhizosphere communities identified scale-free small world topologies, indicating mechanisms of self-organization. While the assembly of rhizosphere bacterial communities is bottom-up controlled through the resource supply from root (secondary) metabolites, our results support the hypothesis that the net effect may depend on the strength of top-down control by protist grazers. Since grazing of protists has a strong impact on the composition and functioning of bacteria communities, protists expand the repertoire of plant genes by functional traits, and should be considered as 'protist microbiomes' in analogy to 'bacterial microbiomes'.

Long-Term Fertilization Strategy Impacts Rhizoctonia solani-Microbe Interactions in Soil and Rhizosphere and Defense Responses in Lettuce.

The long-term effects of agricultural management such as different fertilization strategies on soil microbiota and soil suppressiveness against plant pathogens are crucial. Therefore, the suppressiveness of soils differing in fertilization history was assessed using two Rhizoctonia solani isolates and their respective host plants (lettuce, sugar beet) in pot experiments. Further, the effects of fertilization history and the pathogen R. solani AG1-IB on the bulk soil, root-associated soil and rhizosphere microbiota of lettuce were analyzed based on amplicon sequencing of the 16S rRNA gene and ITS2 region. Organic fertilization history supported the spread of the soil-borne pathogens compared to long-term mineral fertilization. The fertilization strategy affected bacterial and fungal community composition in the root-associated soil and rhizosphere, respectively, but only the fungal community shifted in response to the inoculated pathogen. The potential plant-beneficial genus Talaromyces was enriched in the rhizosphere by organic fertilization and presence of the pathogen. Moreover, increased expression levels of defense-related genes in shoots of lettuce were observed in the soil with organic fertilization history, both in the absence and presence of the pathogen. This may reflect the enrichment of potential plant-beneficial microorganisms in the rhizosphere, but also pathogen infestation. However, enhanced defense responses resulted in retarded plant growth in the presence of R. solani (plant growth/defense tradeoff).

Colonization of roots of cultivated Solanum lycopersicum by dark septate and other ascomycetous endophytes.

Tomato (Solanum lycopersicum L.) roots from four different crop sites in Colombia were surface sterilized and 51 fungal isolates were obtained and conserved for further analysis. Based on microscopical observations and growth characteristics, 20 fungal isolates corresponded to genus Fusarium, six presented asexual conidia different from Fusarium, eight were sterile mycelia, seven of which had dark septate hyphae and 17 did not continue to grow on plates after being recovered from conservation. Growth on different media, detailed morphological characterization and ITS region sequencing of the six sporulating and eight sterile isolates revealed that they belonged to different orders of Ascomycota and that the sterile dark septate endophytes did not correspond to the well known Phialocephala group. Interactions of nine isolates with tomato plantlets were assessed in vitro. No effect on shoot development was revealed, but three isolates caused brown spots in roots. Colonization patterns as analyzed by confocal microscopy differed among the isolates and ranged from epidermal to cortical penetration. Altogether 11 new isolates from root endophytic fungi were obtained, seven of which showed features of dark septate endophytes. Four known morphotypes were represented by five isolates, while six isolates belonged to five morphotypes of putative new unknown species.

Effects of dark septate endophytes on tomato plant performance.

Non-mycorrhizal fungal root endophytes can be found in all natural and cultivated ecosystems, but little is known about their impact on plant performance. The impact of three mitosporic dark septate endophytes (DSE48, DSE49 and Leptodontidium orchidicola) on tomato plant characteristics was studied. Their effects on root and shoot growth, their influence on fruit yield and fruit quality parameters and their ability to diminish the impact of the pathogen Verticillium dahliae were investigated. While shoot biomass of young plants was enhanced between 10% and 20% by the endophytes DSE48 and L. orchidicola in one of two experiments and by DSE49 in both experiments, vegetative growth parameters of 24-week-old plants were not affected except a reproducible increase of root diameter by the isolate DSE49. Concerning fruit yield and quality, L. orchidicola could double the biomass of tomatoes and increased glucose content by 17%, but this was dependent on date of harvest and on root colonisation density. Additionally, the endophytes DSE49 and L. orchidicola decreased the negative effect of V. dahliae on tomato, but only at a low dosage of the pathogen. This indicates that the three dark septate endophytes can have a significant impact on tomato characters, but that the effects are only obvious at early stages of vegetative and generative development and currently too inconsistent to recommend the application of these DSEs in horticultural practice.

Impact of biotic and a-biotic parameters on structure and function of microbial communities living on sclerotia of the soil-borne pathogenic fungus Rhizoctonia solani.

The plant pathogen Rhizoctonia solani is very difficult to control due to its persistent, long-living sclerotial structures in soil. Sclerotia are the main source of infection for Rhizoctonia diseases, which cause high yield losses on a broad host range world-wide. Little is known about micro-organisms associated with sclerotia in soil. Therefore, microbial communities of greenhouse and field incubated Rhizoctonia sclerotia were analysed by a multiphasic approach. Using microbial fingerprints performed by PCR-SSCP, sclerotia-associated bacterial communities showed a high diversity, whereas only a few fungi could be detected. Statistical analysis of fingerprints revealed the influence of soil types, incubation conditions (greenhouse, field), and incubation time (5 and 12 weeks) on the bacterial as well as fungal community. No significant differences were found for the microbial community associated with different Rhizoctonia anastomosis sub-groups (AG 1-IB and AG 1-IC). Rhizoctonia sclerotia are an interesting bio-resource: high proportions of fungal cell-wall degrading isolates as well as those with antagonistic activity towards R. solani were found. While a fraction of 28.4% of sclerotia-associated bacteria (=40 isolates) with antagonistic properties was determined, only 4.4% (=6 isolates) of the fungal isolates were antagonistic. We identified strong antagonists of the genera Bacillus, Enterobacter, Pseudomonas, and Stenotrophomonas, which can be used as biological control agents incorporated in soil or applied to Rhizoctonia host plants.

Potato plant spheres and to a lesser extent the soil type influence the proportion and diversity of bacterial isolates with in vitro antagonistic activity towards Ralstonia solanacearum.

Ralstonia solanacearum biovar2-race3 (Rs r3b2) is an epidemic soil-borne bacterial phytopathogen causing brown rot disease in potato. In this study, we assessed how three soil types stored at the same field site influenced the proportion and diversity of bacterial isolates with in vitro antagonistic activity towards Rs in bulk soil and different potato plant spheres (rhizosphere, endorhiza and endocaulosphere; ecto- and endosphere of seed and yield tubers). In general, the plate counts observed for each sample type were not significantly different. A total of 96 colonies per sample type was picked and screened for in vitro antagonistic activity against Rs. Antagonists were obtained from all bulk soils and plant spheres with the highest proportion obtained from the endorhiza and endocaulosphere of potato plants. BOX-PCR fingerprints of antagonists showed that some were specific for particular plant spheres independent of the soil type, while others originated from different plant spheres of a particular soil type. The majority of antagonists belonged to Pseudomonas. A high proportion of antagonists produced siderophores, and interestingly antagonists from potato tubers frequently carried multiple antibiotic production genes. Our data showed an enrichment of bacteria with genes or traits potentially involved in biocontrol in the rhizosphere and in endophytic compartments. We report that the proportion and diversity of in vitro antagonists towards Rs isolated from bulk soil and different spheres of potato plants grown under field conditions in three different soil types was mainly shaped by the plant sphere and to a lesser extent by the soil type. Bacteria with antagonistic activity towards Ralstonia solanacearum were isolated from all plant spheres and bulk soils but their proportion was highest in endophytic compartments.

Genome Analyses of the Less Aggressive Rhizoctonia solani AG1-IB Isolates 1/2/21 and O8/2 Compared to the Reference AG1-IB Isolate 7/3/14.

Rhizoctonia solani AG1-IB of the phylum Basidiomycota is known as phytopathogenic fungus affecting various economically important crops, such as bean, rice, soybean, figs, cabbage and lettuce. The isolates 1/2/21 and O8/2 of the anastomosis group AG1-IB originating from lettuce plants with bottom rot symptoms represent two less aggressive R. solani isolates, as confirmed in a pathogenicity test on lettuce. They were deeply sequenced on the Illumina MiSeq system applying the mate-pair and paired-end mode to establish their genome sequences. Assemblies of obtained sequences resulted in 2092 and 1492 scaffolds, respectively, for isolate 1/2/21 and O8/2, amounting to a size of approximately 43 Mb for each isolate. Gene prediction by applying AUGUSTUS (v. 3.2.1.) yielded 12,827 and 12,973 identified genes, respectively. Based on automatic functional annotation, genes potentially encoding cellulases and enzymes involved in secondary metabolite synthesis were identified in the AG1-IB genomes. The annotated genome sequences of the less aggressive AG1-IB isolates were compared with the isolate 7/3/14, which is highly aggressive on lettuce and other vegetable crops such as bean, cabbage and carrot. This analysis revealed the first insights into core genes of AG1-IB isolates and unique determinants of each genome that may explain the different aggressiveness levels of the strains.

Role of Benzoic Acid and Lettucenin A in the Defense Response of Lettuce against Soil-Borne Pathogens.

Soil-borne pathogens can severely limit plant productivity. Induced defense responses are plant strategies to counteract pathogen-related damage and yield loss. In this study, we hypothesized that benzoic acid and lettucenin A are involved as defense compounds against Rhizoctonia solani and Olpidium virulentus in lettuce. To address this hypothesis, we conducted growth chamber experiments using hydroponics, peat culture substrate and soil culture in pots and minirhizotrons. Benzoic acid was identified as root exudate released from lettuce plants upon pathogen infection, with pre-accumulation of benzoic acid esters in the root tissue. The amounts were sufficient to inhibit hyphal growth of R. solani in vitro (30%), to mitigate growth retardation (51%) and damage of fine roots (130%) in lettuce plants caused by R. solani, but were not able to overcome plant growth suppression induced by Olpidium infection. Additionally, lettucenin A was identified as major phytoalexin, with local accumulation in affected plant tissues upon infection with pathogens or chemical elicitation (CuSO4) and detected in trace amounts in root exudates. The results suggest a two-stage defense mechanism with pathogen-induced benzoic acid exudation initially located in the rhizosphere followed by accumulation of lettucenin A locally restricted to affected root and leaf tissues.

Distinct rhizomicrobiota assemblages and plant performance in lettuce grown in soils with different agricultural management histories.

A better understanding of factors shaping the rhizosphere microbiota is important for sustainable crop production. We hypothesized that the effect of agricultural management on the soil microbiota is reflected in the assemblage of the rhizosphere microbiota with implications for plant performance. We designed a growth chamber experiment growing the model plant lettuce under controlled conditions in soils of a long-term field experiment with contrasting histories of tillage (mouldboard plough vs cultivator tillage), fertilization intensity (intensive standard nitrogen (N) + pesticides/growth regulators vs extensive reduced N without fungicides/growth regulators), and last standing field crop (rapeseed vs winter wheat). High-throughput sequencing of bacterial and archaeal 16S rRNA genes and fungal ITS2 regions amplified from total community DNA showed that these factors shaped the soil and rhizosphere microbiota of lettuce, however, to different extents among the microbial domains. Pseudomonas and Olpidium were identified as major indicators for agricultural management in the rhizosphere of lettuce. Long-term extensive fertilization history of soils resulted in higher lettuce growth and increased expression of genes involved in plant stress responses compared to intensive fertilization. Our work adds to the increasing knowledge on how soil microbiota can be manipulated by agricultural management practices which could be harnessed for sustainable crop production.