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.

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.

Datathons: fostering equitability in data reuse in ecology.

Approaches to rapidly collecting global biodiversity data are increasingly important, but biodiversity blind spots persist. We organized a three-day Datathon event to improve the openness of local biodiversity data and facilitate data reuse by local researchers. The first Datathon, organized among microbial ecologists in Uruguay and Argentina assembled the largest microbiome dataset in the region to date and formed collaborative consortia for microbiome data synthesis.

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.

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).

Microbial community analysis of soils under different soybean cropping regimes in the Argentinean south-eastern Humid Pampas.

Soil microbial communities are key players of ecosystem processes and important for crop and soil health. The Humid Pampas region in Argentina concentrates 75% of the national soybean production, which is based on intensive use of agrochemicals, monocropping and no-till. A long-term field experiment under no-till management in the southeast of the Argentinean Pampas provides a unique opportunity to compare soybean under monocropping with cultivation including alternating cover crops or in a three-phase rotation. We hypothesized that cropping regimes and season affect soil microbial community composition and diversity. Amplicon sequencing of 16S rRNA genes and internal transcribed spacer fragments showed a stronger microbial seasonal dynamic in conservation regimes compared to monocropping. In addition, several bacterial (e.g. Catenulispora, Streptomyces and Bacillus) and fungal genera (e.g. Exophiala) with cropping regime-dependent differential relative abundances were identified. Despite a temporal shift in microbial and chemical parameters, this study shows that long-term cropping regimes shaped the soil microbiota. This might have important implications for soil quality and soybean performance and should therefore be considered in the development of sustainable agricultural managements.

Tillage shapes the soil and rhizosphere microbiome of barley-but not its susceptibility towards Blumeria graminis f. sp. hordei.

Long-term agricultural practices are assumed to shape the rhizosphere microbiome of crops with implications for plant health. In a long-term field experiment, we investigated the effect of different tillage and fertilization practices on soil and barley rhizosphere microbial communities by means of amplicon sequencing of 16S rRNA gene fragments from total community DNA. Differences in the microbial community composition depending on the tillage practice, but not the fertilization intensity were revealed. To examine whether these soil and rhizosphere microbiome differences influence the plant defense response, barley (cultivar Golden Promise) was grown in field or standard potting soil under greenhouse conditions and challenged with Blumeria graminis f. sp. hordei (Bgh). Amplicon sequence analysis showed that preceding tillage practice, but also aboveground Bgh challenge significantly influenced the microbial community composition. Expression of plant defense-related genes PR1b and PR17b was higher in challenged compared to unchallenged plants. The Bgh infection rates were strikingly lower for barley grown in field soil compared to potting soil. Although previous agricultural management shaped the rhizosphere microbiome, no differences in plant health were observed. We propose therefore that the management-independent higher microbial diversity of field soils compared to potting soils contributed to the low infection rates of barley.

Reduced tillage, cover crops and organic amendments affect soil microbiota and improve soil health in Uruguayan vegetable farming systems.

Conventional tillage and mineral fertilization (CTMF) jeopardize soil health in conventional vegetable production systems. Using a field experiment established in Uruguay in 2012, we aimed to compare the soil restoration potential of organic fertilization (compost and poultry manure) combined with conventional tillage and cover crop incorporated into the soil (CTOF) or with reduced tillage and the use of cover crop as mulch (RTOF). In 2017, table beet was cultivated under CTMF, CTOF and RTOF, and yields, soil aggregate composition and nutrients, as well as soil and table beet rhizosphere microbiota (here: bacteria and archaea) were evaluated. Microbiota was studied by high-throughput sequencing of 16S rRNA gene fragments amplified from total community DNA. RTOF exhibited higher soil aggregation, soil organic C, nutrient availability and microbial alpha-diversity than CTMF, and became more similar to an adjacent natural undisturbed site. The soil microbiota was strongly shaped by the fertilization source which was conveyed to the rhizosphere and resulted in differentially abundant taxa. However, 229 amplicon sequencing variants were found to form the core table beet rhizosphere microbiota shared among managements. In conclusion, our study shows that after only 5 years of implementation, RTOF improves soil health under intensive vegetable farming systems.

Impacts of switching tillage to no-tillage and vice versa on soil structure, enzyme activities and prokaryotic community profiles in Argentinean semi-arid soils.

The effects of tillage on soil structure, physiology and microbiota structure were studied in a long-term field experiment, with side-to-side plots, established to compare effects of conventional tillage (CT) vs no-till (NT) agriculture. After 27 years, part of the field under CT was switched to NT and vice versa. Soil texture, soil enzymatic profiles and the prokaryotic community structure (16S rRNA genes amplicon sequencing) were analyzed at two soil depths (0-5 and 5-10 cm) in samples taken 6, 18 and 30 months after switching tillage practices. Soil enzymatic activities were higher in NT than CT, and enzymatic profiles responded to the changes much earlier than the overall prokaryotic community structure. Beta diversity measurements of the prokaryotic community indicated that the levels of stratification observed in long-term NT soils were already recovered in the new NT soils 30 months after switching from CT to NT. Bacteria and Archaea OTUs that responded to NT were associated with coarse soil fraction, soil organic carbon and C cycle enzymes, while CT responders were related to fine soil fractions and S cycle enzymes. This study showed the potential of managing the soil prokaryotic community and soil health through changes in agricultural management practices.

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.

Impact of Long-Term Organic and Mineral Fertilization on Rhizosphere Metabolites, Root-Microbial Interactions and Plant Health of Lettuce.

Fertilization management can affect plant performance and soil microbiota, involving still poorly understood rhizosphere interactions. We hypothesized that fertilization practice exerts specific effects on rhizodeposition with consequences for recruitment of rhizosphere microbiota and plant performance. To address this hypothesis, we conducted a minirhizotron experiment using lettuce as model plant and field soils with contrasting properties from two long-term field experiments (HUB-LTE: loamy sand, DOK-LTE: silty loam) with organic and mineral fertilization history. Increased relative abundance of plant-beneficial arbuscular mycorrhizal fungi and fungal pathotrophs were characteristic of the rhizospheres in the organically managed soils (HU-org; BIODYN2). Accordingly, defense-related genes were systemically expressed in shoot tissues of the respective plants. As a site-specific effect, high relative occurrence of the fungal lettuce pathogen Olpidium sp. (76-90%) was recorded in the rhizosphere, both under long-term organic and mineral fertilization at the DOK-LTE site, likely supporting Olpidium infection due to a lower water drainage potential compared to the sandy HUB-LTE soils. However, plant growth depressions and Olpidium infection were exclusively recorded in the BIODYN2 soil with organic fertilization history. This was associated with a drastic (87-97%) reduction in rhizosphere abundance of potentially plant-beneficial microbiota (Pseudomonadaceae, Mortierella elongata) and reduced concentrations of the antifungal root exudate benzoate, known to be increased in presence of Pseudomonas spp. In contrast, high relative abundance of Pseudomonadaceae (Gammaproteobacteria) in the rhizosphere of plants grown in soils with long-term mineral fertilization (61-74%) coincided with high rhizosphere concentrations of chemotactic dicarboxylates (succinate, malate) and a high C (sugar)/N (amino acid) ratio, known to support the growth of Gammaproteobacteria. This was related with generally lower systemic expression of plant defense genes as compared with organic fertilization history. Our results suggest a complex network of belowground interactions among root exudates, site-specific factors and rhizosphere microbiota, modulating the impact of fertilization management with consequences for plant health and performance.

Rhizosphere Competence and Biocontrol Effect of Pseudomonas sp. RU47 Independent from Plant Species and Soil Type at the Field Scale.

Biocontrol inoculants often show inconsistency in their efficacy at field scale and the reason for this remains often unclear. A high rhizosphere competence of inoculant strains is assumed to be a key factor for successful biocontrol effects as the biocontrol strain has to compete with the indigenous microbial community in the rhizosphere. It is known that many factors, among them plant species and soil type shape the rhizosphere microbial community composition. However, microbial community composition in the rhizosphere can also be influenced by the presence of a pathogen. We hypothesized that plant species, soil type, and a pathogen affect the rhizosphere competence of a biocontrol strain and its biocontrol effect against a soil-borne pathogen. To test the hypothesis, we used an experimental plot system with three soil types (diluvial sand, alluvial loam, loess loam) kept under similar agricultural management at the same field site for 12 years. We investigate the rhizosphere competence of Pseudomonas sp. RU47 in two plant species (potato and lettuce) and its biocontrol effect against Rhizoctonia diseases. The colonization density of a rifampicin resistant mutant of RU47 in the rhizosphere of both crops was evaluated by plate counts. Bacterial community compositions were analyzed by denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments amplified from total community DNA. The inoculant RU47 was able to colonize the rhizosphere of both model crops in a sufficient density and to reduce disease severity of black scurf on potato and bottom rot on lettuce in all three soils. DGGE indicated that RU47 affected the bacterial community composition stronger in the rhizosphere of lettuce than in the potato rhizosphere. In contrast, the effect of the pathogen Rhizoctonia solani on the bacterial community was much stronger in the rhizosphere of potato than in the lettuce rhizosphere. A significant effect of RU47 on the Pseudomonas-specific gacA fingerprints of the rhizosphere was only observed in lettuce in alluvial soil. The soil type and plant species independent biocontrol effects of RU47 and its minor influence on the indigenous bacterial community composition might be important criteria for the registration and use of RU47 as biocontrol strain.

Fungal community profiles in agricultural soils of a long-term field trial under different tillage, fertilization and crop rotation conditions analyzed by high-throughput ITS-amplicon sequencing.

Fungal communities in agricultural soils are assumed to be affected by soil and crop management. Our intention was to investigate the impact of different tillage and fertilization practices on fungal communities in a long-term crop rotation field trial established in 1992 in Central Germany. Two winter wheat fields in replicated strip-tillage design, comprising conventional vs. conservation tillage, intensive vs. extensive fertilization and different pre-crops (maize vs. rapeseed) were analyzed by a metabarcoding approach applying Illumina paired-end sequencing of amplicons generated by two recently developed primer pairs targeting the two fungal Internal Transcribed Spacer regions (ITS1, ITS2). Analysis of 5.1 million high-quality sequence reads uncovered a diverse fungal community in the two fields, composed of 296 fungal genera including 3,398 Operational Taxonomic Units (OTUs) at the 97% sequence similarity threshold. Both primer pairs detected the same fungal phyla (Basidio-, Asco-, Zygo-, Glomero- and Chytridiomycota), but in different relative abundances. OTU richness was higher in the ITS1 dataset, while ITS2 data were more diverse and of higher evenness. Effects of farming practice on fungal community structures were revealed. Almost two-thirds of the fungal genera were represented in all different soil treatments, whereas the remaining genera clearly responded to farming practice. Principal Component Analysis revealed four distinct clusters according to tillage practice and pre-crop. Analysis of Variance (ANOVA) substantiated the results and proved significant influences of tillage and pre-crop, while fertilization had the smallest and non-significant effect. In-depth analysis of putative phytopathogenic and plant beneficial fungal groups indicated distinct responses; for example Fusarium was significantly enriched in the intensively fertilized conservation tillage variants with the pre-crop maize, while Phoma displayed significant association with conventional tillage and pre-crop rapeseed. Many putative plant beneficial fungi also reacted differentially to farming practice with the most distinct responders identified among the Glomeromycota (arbuscular mycorrhizal fungi, AMF).

Soil mineral composition matters: response of microbial communities to phenanthrene and plant litter addition in long-term matured artificial soils.

The fate of polycyclic aromatic hydrocarbons (PAHs) in soil is determined by a suite of biotic and abiotic factors, and disentangling their role in the complex soil interaction network remains challenging. Here, we investigate the influence of soil composition on the microbial community structure and its response to the spiked model PAH compound phenanthrene and plant litter. We used long-term matured artificial soils differing in type of clay mineral (illite, montmorillonite) and presence of charcoal or ferrihydrite. The soils received an identical soil microbial fraction and were incubated for more than two years with two sterile manure additions. The matured artificial soils and a natural soil were subjected to the following spiking treatments: (I) phenanthrene, (II) litter, (III) litter + phenanthrene, (IV) unspiked control. Total community DNA was extracted from soil sampled on the day of spiking, 7, 21, and 63 days after spiking. Bacterial 16S rRNA gene and fungal internal transcribed spacer amplicons were quantified by qPCR and subjected to denaturing gradient gel electrophoresis (DGGE). DGGE analysis revealed that the bacterial community composition, which was strongly shaped by clay minerals after more than two years of incubation, changed in response to spiked phenanthrene and added litter. DGGE and qPCR showed that soil composition significantly influenced the microbial response to spiking. While fungal communities responded only in presence of litter to phenanthrene spiking, the response of the bacterial communities to phenanthrene was less pronounced when litter was present. Interestingly, microbial communities in all artificial soils were more strongly affected by spiking than in the natural soil, which might indicate the importance of higher microbial diversity to compensate perturbations. This study showed the influence of soil composition on the microbiota and their response to phenanthrene and litter, which may increase our understanding of complex interactions in soils for bioremediation applications.

Metal oxides, clay minerals and charcoal determine the composition of microbial communities in matured artificial soils and their response to phenanthrene.

Microbial communities in soil reside in a highly heterogeneous habitat where diverse mineral surfaces, complex organic matter and microorganisms interact with each other. This study aimed to elucidate the long-term effect of the soil mineral composition and charcoal on the microbial community composition established in matured artificial soils and their response to phenanthrene. One year after adding sterile manure to different artificial soils and inoculating microorganisms from a Cambisol, the matured soils were spiked with phenanthrene or not and incubated for another 70 days. 16S rRNA gene and internal transcribed spacer fragments amplified from total community DNA were analyzed by denaturing gradient gel electrophoresis. Metal oxides and clay minerals and to a lesser extent charcoal influenced the microbial community composition. Changes in the bacterial community composition in response to phenanthrene differed depending on the mineral composition and presence of charcoal, while no shifts in the fungal community composition were observed. The abundance of ring-hydroxylating dioxygenase genes was increased in phenanthrene-spiked soils except for charcoal-containing soils. Here we show that the formation of biogeochemical interfaces in soil is an ongoing process and that different properties present in artificial soils influenced the bacterial response to the phenanthrene spike.

Mineral composition and charcoal determine the bacterial community structure in artificial soils.

To study the influence of the clay minerals montmorillonite (M) and illite (I), the metal oxides ferrihydrite (F) and aluminum hydroxide (A), and charcoal (C) on soil bacterial communities, seven artificial soils with identical texture provided by quartz (Q) were mixed with sterilized manure as organic carbon source before adding a microbial inoculant derived from a Cambisol. Bacterial communities established in artificial soils after 90 days of incubation were compared by DGGE analysis of bacterial and taxon-specific 16S rRNA gene amplicons. The bacterial community structure of charcoal-containing soils highly differed from the other soils at all taxonomic levels studied. Effects of montmorillonite and illite were observed for Bacteria and Betaproteobacteria, but not for Actinobacteria or Alphaproteobacteria. A weak influence of metal oxides on Betaproteobacteria was found. Barcoded pyrosequencing of 16S rRNA gene amplicons done for QM, QI, QIF, and QMC revealed a high bacterial diversity in the artificial soils. The composition of the artificial soils was different from the inoculant, and the structure of the bacterial communities established in QMC soil was most different from the other soils, suggesting that charcoal provided distinct microenvironments and biogeochemical interfaces formed. Several populations with discriminative relative abundance between artificial soils were identified.

Succession of bacterial and fungal 4-chloro-2-methylphenoxyacetic acid degraders at the soil-litter interface.

Phenoxyacetic acids can be degraded by diverse soil microorganisms. Nevertheless, we miss information about the succession of 4-chloro-2-methylphenoxyacetic acid (MCPA) degraders in micro-environments of soils as well as specific functions of different microbial groups during MCPA degradation. We studied MCPA degradation at the soil-litter interface in a microcosm experiment and followed the succession of different degrader populations by quantifying the abundance of 16S rRNA genes as well as, the fungal ITS fragment and the functional genes tfdA (in total and divided into three classes) and cadA. Adjacent to the litter layer, a dynamic depletion zone of MCPA indicated that the litter effect on MCPA degradation depends on substrate availability and the affected soil volume. The increase of the tfdA class III and cadA genes was linked to MCPA mineralisation. Total abundance of tfdA genes was dominated by class I MCPA degraders and did not reflect MCPA degradation potential of the soil. Litter addition induced the development of pioneer and late-stage fungal communities, which were probably both involved in MCPA degradation. The results underline the importance of the ecological behaviour of different degrader populations for the understanding of herbicide degradation in soils.