Harnessing root-soil-microbiota interactions for drought-resilient cereals.

Cereal plants form complex networks with their associated microbiome in the soil environment. A complex system including variations of numerous parameters of soil properties and host traits shapes the dynamics of cereal microbiota under drought. These multifaceted interactions can greatly affect carbon and nutrient cycling in soil and offer the potential to increase plant growth and fitness under drought conditions. Despite growing recognition of the importance of plant microbiota to agroecosystem functioning, harnessing the cereal root microbiota remains a significant challenge due to interacting and synergistic effects between root traits, soil properties, agricultural practices, and drought-related features. A better mechanistic understanding of root-soil-microbiota associations could lead to the development of novel strategies to improve cereal production under drought. In this review, we discuss the root-soil-microbiota interactions for improving the soil environment and host fitness under drought and suggest a roadmap for harnessing the benefits of these interactions for drought-resilient cereals. These methods include conservative trait-based approaches for the selection and breeding of plant genetic resources and manipulation of the soil environments.

Early assessment of fungal and oomycete pathogens in greenhouse irrigation water using Oxford nanopore amplicon sequencing.

Water-borne plant pathogenic fungi and oomycetes are a major threat in greenhouse production systems. Early detection and quantification of these pathogens would enable us to ascertain both economic and biological thresholds required for a timely treatment, thus improving effective disease management. Here, we used Oxford nanopore MinION amplicon sequencing to analyze microbial communities in irrigation water collected from greenhouses used for growing tomato, cucumber and Aeschynanthus sp. Fungal and oomycete communities were characterized using primers that amplify the full internal transcribed spacer (ITS) region. To assess the sensitivity of the MinION sequencing, we spiked serially diluted mock DNA into the DNA isolated from greenhouse water samples prior to library preparation. Relative abundances of fungal and oomycete reads were distinct in the greenhouse irrigation water samples and in water samples from setups with tomato that was inoculated with Fusarium oxysporum. Sequence reads derived from fungal and oomycete mock communities were proportionate in the respective serial dilution samples, thus confirming the suitability of MinION amplicon sequencing for environmental monitoring. By using spike-ins as standards to test the reliability of quantification using the MinION, we found that the detection of spike-ins was highly affected by the background quantities of fungal or oomycete DNA in the sample. We observed that spike-ins having shorter length (538bp) produced reads across most of our dilutions compared to the longer spikes (>790bp). Moreover, the sequence reads were uneven with respect to dilution series and were least retrievable in the background samples having the highest DNA concentration, suggesting a narrow dynamic range of performance. We suggest continuous benchmarking of the MinION sequencing to improve quantitative metabarcoding efforts for rapid plant disease diagnostic and monitoring in the future.

Mechanisms Underlying the Overlooked Chiral Fungicide-Driven Enantioselective Proliferation of Antibiotic Resistance in Earthworm Intestinal Microbiome.

From a "One Health" perspective, the global threat of antibiotic resistance genes (ARGs) is associated with modern agriculture practices including agrochemicals application. Chiral fungicides account for a considerable proportion of wildly used agrochemicals; however, whether and how their enantiomers lead to differential proliferation of antibiotic resistance in agricultural environments remain overlooked. Focused on the soil-earthworm ecosystem, we for the first time deciphered the mechanisms underlying the enantioselective proliferation of antibiotic resistance driven by the enantiomers of a typical chiral fungicide mandipropamid (i.e., R-MDP and S-MDP) utilizing a multiomic approach. Time-series metagenomic analysis revealed that R-MDP led to a significant enhancement of ARGs with potential mobility (particularly the plasmid-borne ARGs) in the earthworm intestinal microbiome. We further demonstrated that R-MDP induced a concentration-dependent facilitation of plasmid-mediated ARG transfer among microbes. In addition, transcriptomic analysis with verification identified the key aspects involved, where R-MDP enhanced cell membrane permeability, transfer ability, biofilm formation and quorum sensing, rebalanced energy production, and decreased cell mobility versus S-MDP. Overall, the findings provide novel insights into the enantioselective disruption of microbiome and resistome in earthworm gut by chiral fungicides and offer significant contributions to the comprehensive risk assessment of chiral agrochemicals in agroecosystems.

Fungal Communities of Spring Barley from Seedling Emergence to Harvest During a Severe Puccinia hordei Epidemic.

All plant tissues from leaves, stems, and roots are hosting a wide diversity of fungal species. Our understanding of the assembly of this diversity of fungi during the plant growth cycle is limited. Here, we characterized the mycobiome of three spring barley cultivars grown in Zealand, Denmark, at weekly intervals during a growth season from seedling emergence to senescence and seed maturity. A notable proportion of members of the fungal communities were shared among different plant organs, but community dynamics were tissue-specific. A severe attack of Puccinia hordei occurring during the vegetative stage had profound effects on the mycobiome, and P. hordei biomass displaced that of other taxa. Plant tissue type was the most important factor determining the mycobiome, but also plant age was contributing significantly. Using a random forest model, we found that specific members of the mycobiome were responding differently to plant age, for instance, Olpidium and Articulospora in roots, Dioszegia and Sporobolomyces in leaves, Pyrenophora in stems, and Epicoccum in heads. A co-occurrence network analysis revealed complex interactions among fungal OTUs, and network connectivity was changing as per plant growth stage and plant tissue type. This study contributes to the understanding of assembly of fungal communities in cereals by providing a detailed description of fungal communities associated with barley. This knowledge will be vital for microbiome assisted plant health management and our study will serve as an important baseline for future efforts to harness microbiota in cereal health.

Distinct chemical resistance-inducing stimuli result in common transcriptional, metabolic, and nematode community signatures in rice root and rhizosphere.

Induced resistance (IR), a phenotypic state induced by an exogenous stimulus and characterized by enhanced resistance to future (a)biotic challenge, is an important component of plant immunity. Numerous IR-inducing stimuli have been described in various plant species, but relatively little is known about 'core' systemic responses shared by these distinct IR stimuli and the effects of IR on plant-associated microbiota. In this study, rice (Oryza sativa) leaves were treated with four distinct IR stimuli (ß-aminobutyric acid, acibenzolar-S-methyl, dehydroascorbic acid, and piperonylic acid) capable of inducing systemic IR against the root-knot nematode Meloidogyne graminicola and evaluated their effect on the root transcriptome and exudome, and root-associated nematode communities. Our results reveal shared transcriptional responses-notably induction of jasmonic acid and phenylpropanoid metabolism-and shared alterations to the exudome that include increased amino acid, benzoate, and fatty acid exudation. In rice plants grown in soil from a rice field, IR stimuli significantly affected the composition of rhizosphere nematode communities 3 d after treatment, but by 14 d after treatment these changes had largely reverted. Notably, IR stimuli did not reduce nematode diversity, which suggests that IR might offer a sustainable option for managing plant-parasitic nematodes.

Fusarium oxysporum Disrupts Microbiome-Metabolome Networks in Arabidopsis thaliana Roots.

While the plant host metabolome drives distinct enrichment of detrimental and beneficial members of the microbiome, the mechanistic interomics relationships remain poorly understood. Here, we studied microbiome and metabolome profiles of two Arabidopsis thaliana accessions after Fusarium oxysporum f.sp. mathioli (FOM) inoculation, Landsberg erecta (Ler-0) being susceptible and Col-0 being resistant against FOM. By using bacterial and fungal amplicon sequencing and targeted metabolite analysis, we observed highly dynamic microbiome and metabolome profiles across FOM host progression, while being markedly different between FOM-inoculated and noninoculated Col-0 and Ler-0. Co-occurrence network analysis revealed more robust microbial networks in the resistant Col-0 compared to Ler-0 during FOM infection. Correlation analysis revealed distinct metabolite-OTU correlations in Ler-0 compared with Col-0 which could possibly be explained by missense variants of the Rfo3 and Rlp2 genes in Ler-0. Remarkably, we observed positive correlations in Ler-0 between most of the analyzed metabolites and the bacterial phyla Proteobacteria, Bacteroidetes, Planctomycetes, Acidobacteria, and Verrucomicrobia, and negative correlations with Actinobacteria, Firmicutes, and Chloroflexi. The glucosinolates 4-methyoxyglucobrassicin, glucoerucin and indole-3 carbinol, but also phenolic compounds were strongly correlating with the relative abundances of indicator and hub OTUs and thus could be active in structuring the A. thaliana root-associated microbiome. Our results highlight interactive effects of host plant defense and root-associated microbiota on Fusarium infection and progression. Our findings provide significant insights into plant interomic dynamics during pathogen invasion and could possibly facilitate future exploitation of microbiomes for plant disease control. IMPORTANCE Plant health and fitness are determined by plant-microbe interactions which are guided by host-synthesized metabolites. To understand the orchestration of this interaction, we analyzed the distinct interomic dynamics in resistant and susceptible Arabidopsis ecotypes across different time points after infection with Fusarium oxysporum (FOM). Our results revealed distinct microbial profiles and network resilience during FOM infection in the resistant Col-0 compared with the susceptible Ler-0 and further pinpointed specific microbe-metabolite associations in the Arabidopsis microbiome. These findings provide significant insights into plant interomics dynamics that are likely affecting fungal pathogen invasion and could possibly facilitate future exploitation of microbiomes for plant disease control.

Genetic disruption of Arabidopsis secondary metabolite synthesis leads to microbiome-mediated modulation of nematode invasion.

In-depth understanding of metabolite-mediated plant-nematode interactions can guide us towards novel nematode management strategies. To improve our understanding of the effects of secondary metabolites on soil nematode communities, we grew Arabidopsis thaliana genetically altered in glucosinolate, camalexin, or flavonoid synthesis pathways, and analyzed their root-associated nematode communities using metabarcoding. To test for any modulating effects of the associated microbiota on the nematode responses, we characterized the bacterial and fungal communities. Finally, as a proxy of microbiome-modulating effects on nematode invasion, we isolated the root-associated microbiomes from the mutants and tested their effect on the ability of the plant parasitic nematode Meloidogyne incognita to penetrate tomato roots. Most mutants had altered relative abundances of several nematode taxa with stronger effects on the plant parasitic Meloidogyne hapla than on other root feeding taxa. This probably reflects that M. hapla invades and remains embedded within root tissues and is thus intimately associated with the host. When transferred to tomato, microbiomes from the flavonoid over-producing pap1-D enhanced M. incognita root-invasion, whereas microbiomes from flavonoid-deficient mutants reduced invasion. This suggests microbiome-mediated effect of flavonoids on Meloidogyne infectivity plausibly mediated by the alteration of the abundances of specific microbial taxa in the transferred microbiomes, although we could not conclusively pinpoint such causative microbial taxa.

Rice diterpenoid phytoalexins are involved in defence against parasitic nematodes and shape rhizosphere nematode communities.

Rice diterpenoid phytoalexins (DPs) are secondary metabolites with a well known role in resistance to foliar pathogens. As DPs are also known to be produced and exuded by rice roots, we hypothesised that they might play an important role in plant-nematode interactions, and particularly in defence against phytoparasitic nematodes. We used transcriptome analysis on rice roots to analyse the effect of infection by the root-knot nematode Meloidogyne graminicola or treatment with resistance-inducing chemical stimuli on DP biosynthesis genes, and assessed the susceptibility of mutant rice lines impaired in DP biosynthesis to M. graminicola. Moreover, we grew these mutants and their wild-type in field soil and used metabarcoding to assess the effect of impairment in DP biosynthesis on rhizosphere and root nematode communities. We show that M. graminicola suppresses DP biosynthesis genes early in its invasion process and, conversely, that resistance-inducing stimuli transiently induce the biosynthesis of DPs. Moreover, we show that loss of DPs increases susceptibility to M. graminicola. Metabarcoding on wild-type and DP-deficient plants grown in field soil reveals that DPs significantly alter the composition of rhizosphere and root nematode communities. Diterpenoid phytoalexins are important players in basal and inducible defence against nematode pathogens of rice and help shape rice-associated nematode communities.

Arabidopsis assemble distinct root-associated microbiomes through the synthesis of an array of defense metabolites.

Plant associated microbiomes are known to confer fitness advantages to the host. Understanding how plant factors including biochemical traits influence host associated microbiome assembly could facilitate the development of microbiome-mediated solutions for sustainable plant production. Here, we examined microbial community structures of a set of well-characterized Arabidopsis thaliana mutants disrupted in metabolic pathways for the production of glucosinolates, flavonoids, or a number of defense signalling molecules. A. thaliana lines were grown in a natural soil and maintained under greenhouse conditions for 4 weeks before collection of roots for bacterial and fungal community profiling. We found distinct relative abundances and diversities of bacterial and fungal communities assembled in the individual A. thaliana mutants compared to their parental lines. Bacterial and fungal genera were mostly enriched than depleted in secondary metabolite and defense signaling mutants, except for flavonoid mutations on fungi communities. Bacterial genera Azospirillum and Flavobacterium were significantly enriched in most of the glucosinolate, flavonoid and signalling mutants while the fungal taxa Sporobolomyces and Emericellopsis were enriched in several glucosinolates and signalling mutants. Whilst the present study revealed marked differences in microbiomes of Arabidopsis mutants and their parental lines, it is suggestive that unknown enzymatic and pleiotropic activities of the mutated genes could contribute to the identified host-associated microbiomes. Notwithstanding, this study revealed interesting gene-microbiota links, and thus represents valuable resource data for selecting candidate A. thaliana mutants for analyzing the links between host genetics and the associated microbiome.

Phytohormones selectively affect plant parasitic nematodes associated with Arabidopsis roots.

Phytohormones may affect plant-nematode interactions directly as chemo-attractants or -repellents, or indirectly through the root-associated microbiome or through host defense mechanisms. However, the exact roles of phytohormones in these complex plant-soil-nematode interactions are not well understood. We used Arabidopsis thaliana mutants impaired in phytohormone synthesis or sensitivity to elucidate their role in root-nematode interactions. As root-associated microorganisms may modulate these interactions, we explored correlations between the relative abundances of root-associated nematodes, and bacteria and fungi using amplicon sequencing. We found distinct shifts in relative abundances of a range of nematode taxa in the A. thaliana phytohormone mutants. The root knot nematode Meloidogyne hapla, a sedentary endoparasitic species that is in intimate contact with the host, was highly enriched in JA-, SA- and SL-impaired lines, and in an ET-insensitive line. Positive or negative correlations between specific microbial and nematode taxa were observed, but, as the inference of causal relationships between microbiome responses and effects on nematode communities is premature, this should be studied in detail in future studies. In conclusion, genetic derailment of hormonal balances generally rendered plants vulnerable to endoparasitic nematode attack. Furthermore, preliminary data suggest that this effect may be partially modulated by the associated microbiome.

Benzoxazinoids selectively affect maize root-associated nematode taxa.

Although the effects of plant secondary metabolites on plant defence have been studied for decades, the exact roles of secondary metabolites in shaping plant-associated microbial and nematode communities remain elusive. We evaluated the effects of benzoxazinoids, a group of secondary metabolites present in several cereals, on root-associated nematodes. We employed 18S rRNA metabarcoding to compare maize root-associated nematode communities in a bx1 knockout maize line impaired in benzoxazinoid synthesis and in its parental wild type. Both genotype and plant age affected the composition of the nematode community in the roots, and the effects of benzoxazinoids on nematode communities were stronger in the roots than in the rhizosphere. Differential abundance analysis and quantitative PCR showed that the root lesion nematode Pratylenchus neglectus was enriched in the bx1 mutant line, while another root lesion nematode, Pratylenchus crenatus, was reduced. Correlation analysis showed that benzoxazinoid concentrations in maize roots mostly correlated negatively with the relative abundance of nematode sequence reads. However, positive correlations between benzoxazinoids and nematode taxa, including several plant-parasitic nematodes, were also identified. Our detailed nematode community analysis suggests differential and selective effects of benzoxazinoids on soil nematodes depending on both the nematode species and the benzoxazinoid compound.

Comparative Genomics, Pangenome, and Phylogenomic Analyses of Brenneria spp., and Delineation of Brenneria izadpanahii sp. nov.

Brenneria species are bacterial plant pathogens mainly affecting woody plants. Association of all members with devastating disorders (e.g., acute oak decline in Iran and United Kingdom) are due to adaptation and pathogenic behavior in response to host and environmental factors. Some species, including B. goodwinii, B. salicis, and B. nigrifluens, also show endophytic residence. Here we show that all species including novel Brenneria sp. are closely related. Gene-based and genome/pangenome-based phylogeny divide the genus into two distinct lineages, Brenneria clades A and B. The two clades were functionally distinct and were consistent with their common and special potential activities as determined via annotation of functional domains. Pangenome analysis demonstrated that the core pathogenicity factors were highly conserved, an hrp gene cluster encoding a type III secretion system was found in all species except B. corticis. An extensive repertoire of candidate virulence factors was identified. Comparative genomics indicated a repertoire of plant cell wall degrading enzymes, metabolites/antibiotics, and numerous prophages providing new insights into Brenneria-host interactions and appropriate targets for further characterization. This work not only documented the genetic differentiation of Brenneria species but also delineates a more functionally driven understanding of Brenneria by comparison with relevant Pectobacteriaceae thereby substantially enriching the extent of information available for functional genomic investigations.

Mass Spectrometry-Based Metabolomics Reveals a Concurrent Action of Several Chemical Mechanisms in Arabidopsis-Fusarium oxysporum Compatible and Incompatible Interactions.

Fusarium oxysporum is a destructive root-infecting plant pathogen that causes significant yield losses in many economically important crop species. Hence, a deeper understanding of pathogen infection strategies is needed. With liquid chromatography-tandem mass spectrometry and gas chromatography-time of flight mass spectrometry platforms, we analyzed the metabolic changes in a time-course experiment with Arabidopsis accessions either resistant (Col-0) or susceptible (Ler-0) to isolates of Fusarium oxysporum forma specialis matthioli infection. We showed a concurrent effect of Fusarium-derived polyols and the mycotoxin beauvericin in the suppression of the immune response of susceptible hosts. A significant increase in oxidized glutathione in the resistant host was probably associated with effective reactive oxygen species-mediated resistance responses. Through a combination of targeted and untargeted metabolomics, we demonstrated the concurrent action of several Arabidopsis defense systems as well as the concurrent action of several virulence systems in the fungal attack of susceptible Arabidopsis.

Breeding selection imposed a differential selective pressure on the wheat root-associated microbiome.

Plants-microbiome associations are the result of millions of years of co-evolution. Due to breeding-accelerated plant evolution in non-native and highly managed soil, plant-microbe links could have been lost. We hypothesized that post-domestication breeding of wheat changed the root-associated microbiome. To test this, we analyzed root-associated fungal and bacterial communities shortly after emergence of seedlings representing a transect of wheat evolution including modern wheat, landraces and ancestors. Numbers of observed microbial taxa were highest in landraces bred in low-input agricultural systems, and lowest in ancestors that had evolved in native soils. The microbial communities of modern cultivars were different from those of landraces and ancestors. Old wheat accessions enriched Acidobacteria and Actinobacteria, while modern cultivars enriched OTUs from Candidatus Saccharibacteria, Verrucomicrobia and Firmicutes. The fungal pathogens Fusarium, Neoascochyta and Microdochium enriched in modern cultivars. Both bacterial and fungal communities followed a neutral assembly model when bulk soil was considered as the source community, but accessions of the ancient Triticum turgidum and T. monococcum created a more isolated environment in their roots. In conclusion, wheat root-associated microbiomes have dramatically changed through a transect of breeding history.

Pathogen Infection and Host-Resistance Interactively Affect Root-Associated Fungal Communities in Watermelon.

Interactions of pathogen infection, host plant resistance, and fungal communities are poorly understood. Although the use of resistant watermelon cultivars is an effective control measure of watermelon wilt disease, fungal communities may also have significant effects on the development of the soil-borne pathogen complexes. We characterized the root and rhizosphere fungal communities associated with healthy and diseased watermelons of three different cultivars with different susceptibilities toward wilt disease by paired-end Illumina MiSeq sequencing. Thirty watermelon plants including highly wilt-resistant, moderately resistant, and susceptible cultivars were collected from a greenhouse, half of which showing clear wilt symptoms and the other half with no symptoms. Patterns of watermelon wilt disease and the response of the fungal communities varied among the three cultivars. The amount of the pathogen Fusarium oxysporum f. sp. niveum was higher in diseased root and rhizosphere samples, particularly in the susceptible cultivar, and was significantly positively correlated with the disease index of Fusarium wilt. Plant health had significant effects on root-associated fungal communities, whereas only the highly resistant cultivar had significant effects only on the rhizosphere fungal communities. Co-occurrence networks revealed a higher complexity of fungal communities in the symptom-free roots compared to diseased roots. In addition, networks from roots of the highly resistant plants showing symptoms had a higher complexity compared to the susceptible cultivars. Keystone species were identified for the plants with different symptom severity and the different cultivars in the root and rhizosphere, such as Fusarium oxysporum, Monosporascus cannonballus, and Mortierella alpina. Overall, the most important factor determining fungal communities in the roots was plant symptom severity, whereas in the rhizosphere, plant genotype was the most important factor determining fungal communities.

Fusarium Head Blight Modifies Fungal Endophytic Communities During Infection of Wheat Spikes.

Fusarium head blight (FHB) is a devastating disease of wheat heads. It is caused by several species from the genus Fusarium. Several endophytic fungi also colonize wheat spikes asymptomatically. Pathogenic and commensal fungi share and compete for the same niche and thereby influence plant performance. Understanding the natural dynamics of the fungal community and how the pre-established species react to pathogen attack can provide useful information on the disease biology and the potential use of some of these endophytic organisms in disease control strategies. Fungal community composition was assessed during anthesis as well as during FHB attack in wheat spikes during 2016 and 2017 in two locations. Community metabarcoding revealed that endophyte communities are dominated by basidiomycete yeasts before anthesis and shift towards a more opportunistic ascomycete-rich community during kernel development. These dynamics are interrupted when Fusarium spp. colonize wheat spikes. The Fusarium pathogens appear to exclude other fungi from floral tissues as they are associated with a reduction in community diversity, especially in the kernel which they colonize rapidly. Similarly, the presence of several endophytes was negatively correlated with Fusarium spp. and linked with spikes that stayed healthy despite exposure to the pathogen. These endophytes belonged to the genera Cladosporium, Itersonillia and Holtermanniella. These findings support the hypothesis that some naturally occurring endophytes could outcompete or prevent FHB and represent a source of potential biological control agents in wheat.

Maize synthesized benzoxazinoids affect the host associated microbiome.

BACKGROUND: Plants actively shape their associated microbial communities by synthesizing bio-active substances. Plant secondary metabolites are known for their signaling and plant defense functions, yet little is known about their overall effect on the plant microbiome. In this work, we studied the effects of benzoxazinoids (BXs), a group of secondary metabolites present in maize, on the host-associated microbial structure. Using BX knock-out mutants and their W22 parental lines, we employed 16S and ITS2 rRNA gene amplicon analysis to characterize the maize microbiome at early growth stages. RESULTS: Rhizo-box experiment showed that BXs affected microbial communities not only in roots and shoots, but also in the rhizosphere. Fungal richness in roots was more affected by BXs than root bacterial richness. Maize genotype (BX mutants and their parental lines) as well as plant age explained both fungal and bacterial community structure. Genotypic effect on microbial communities was stronger in roots than in rhizosphere. Diverse, but specific, microbial taxa were affected by BX in both roots and shoots, for instance, many plant pathogens were negatively correlated to BX content. In addition, a co-occurrence analysis of the root microbiome revealed that BXs affected specific groups of the microbiome. CONCLUSIONS: This study provides insights into the role of BXs for microbial community assembly in the rhizosphere and in roots and shoots. Coupling the quantification of BX metabolites with bacterial and fungal communities, we were able to suggest a gatekeeper role of BX by showing its correlation with specific microbial taxa and thus providing insights into effects on specific fungal and bacterial taxa in maize roots and shoots. Root microbial co-occurrence networks revealed that BXs affect specific microbial clusters.

Fungicides have complex effects on the wheat phyllosphere mycobiome.

Effects of fungicide treatments on non-target fungi in the phyllosphere are not well known. We studied community composition and dynamics of target (Puccinia striiformis) and non-target fungi in wheat that was heavily infected with yellow rust. Mycobiotas in bulk leaf samples and individual leaves were studied by metabarcoding targeting the internal transcribed spacer-1 (ITS1) region of the ribosomal DNA. The amount of yellow rust in individual samples was quantified by qPCR (quantitative PCR). In addition, septoria tritici blotch (Zymoseptoria tritici), powdery mildew (Blumeria graminis), tan spot (Pyrenophora tritici-repentis), and yellow rust (P. striiformis) were visually evaluated. We showed how fungal communities were affected by three different broad-spectrum fungicides that had been applied at different timings and doses to control Puccinia striiformis. We showed that fungal content was relatively constant even after fungicide treatments. Principal component analysis demonstrated that communities from fungicide-treated plots could be separated from the communities in non-treated plots. We observed effects of fungicide treatments on fungal communities using different dose, timing and products. Some fungi, including the target organism P. striiformis were effectively controlled by most of the fungicide applications whereas some yeasts and also P. tritici-repentis increased after treatments. We demonstrated the feasibility of using metabarcoding as a supplement to visual assessments of fungicide effects on target as well as non-target fungi.

Fungal Communities Including Plant Pathogens in Near Surface Air Are Similar across Northwestern Europe.

Information on the diversity of fungal spores in air is limited, and also the content of airborne spores of fungal plant pathogens is understudied. In the present study, a total of 152 air samples were taken from rooftops at urban settings in Slagelse, DK, Wageningen NL, and Rothamsted, UK together with 41 samples from above oilseed rape fields in Rothamsted. Samples were taken during 10-day periods in spring and autumn, each sample representing 1 day of sampling. The fungal content of samples was analyzed by metabarcoding of the fungal internal transcribed sequence 1 (ITS1) and by qPCR for specific fungi. The metabarcoding results demonstrated that season had significant effects on airborne fungal communities. In contrast, location did not have strong effects on the communities, even though locations were separated by up to 900 km. Also, a number of plant pathogens had strikingly similar patterns of abundance at the three locations. Rooftop samples were more diverse than samples taken above fields, probably reflecting greater mixing of air from a range of microenvironments for the rooftop sites. Pathogens that were known to be present in the crop were also found in air samples taken above the field. This paper is one of the first detailed studies of fungal composition in air with the focus on plant pathogens and shows that it is possible to detect a range of pathogens in rooftop air samplers using metabarcoding.

Oomycete Communities Associated with Reed Die-Back Syndrome.

Phragmites australis (Cav.) Trin. ex Steud. die-back is a widely-studied phenomenon that was first discovered in northern Europe and that, until recently, was almost unknown in the Mediterranean basin. It has been described as a complex syndrome affecting reed populations leading to their retreat and decline, with significant impacts on valuable ecosystem services. Among the factors that cause the decline, soil-living microorganisms can be crucial. The aims of this study were to analyze the diversity of oomycetes communities associated with reed stands, and to understand whether they could play a key role in the decline. Variations in the structure of oomycetes communities were studied by metabarcoding of the internal transcribed spacer (ITS) 1 region of ribosomal DNA, from the sediments of five Italian freshwater ecosystems. They were chosen to cover a large variability in terms of surface area, water depth, microclimate, and presence of documented reed retreat. From 96 samples collected from reed roots, rhizosphere, and bulk soil, we assembled 207661 ITS1 reads into 523 OTUs. We demonstrated that oomycete communities were structured by several factors, among which the most important was die-back occurrence. Our study also indicates that Pythiogeton spp. could be potentially involved in the development of die-back. The role of heavy metals in the soil was also explored, and cadmium concentration was shown to affect oomycetes distribution. This study represents a significant step forward for the characterization of microbial communities associated with reed die-back syndrome and helps to gain knowledge of the complexity of these important wet ecosystems.