Metagenomics reveals effects of fluctuating water conditions on functional pathways in plant litter microbial community.

Climate change modifies environmental conditions, resulting in altered precipitation patterns, moisture availability and nutrient distribution for microbial communities. Changes in water availability are projected to affect a range of ecological processes, including the decomposition of plant litter and carbon cycling. However, a detailed understanding of microbial stress response to drought/flooding is missing. In this study, an intermittent lake is taken up as a model for changes in water availability and how they affect the functional pathways in microbial communities of the decomposing Phragmites australis litter. The results show that most enriched functions in both habitats belonged to the classes of Carbohydrates and Clustering-based subsystems (terms with unknown function) from SEED subsystems classification. We confirmed that changes in water availability resulted in altered functional makeup of microbial communities. Our results indicate that microbial communities under more frequent water stress (due to fluctuating conditions) could sustain an additional metabolic cost due to the production or uptake of compatible solutes to maintain cellular osmotic balance. Nevertheless, although prolonged submergence seemed to have a negative impact on several functional traits in the fungal community, the decomposition rate was not affected.

De novo transcriptome assembly of hyperaccumulating Noccaea praecox for gene discovery.

Hyperaccumulators are a group of plant species that accumulate high concentrations of one or more metal(loid)s in their above-ground tissues without showing any signs of toxicity. Several hyperaccumulating species belong to the Brassicaceae family, among them the Cd and Zn hyperaccumulator Noccaea praecox. In this paper, we present de novo transcriptome assembled from two naturally occurring N. praecox populations growing in (i) metal-enriched soil and (ii) soil non-contaminated with metals (control site). Total RNA was extracted from the leaves of both populations. We obtained 801,935,101 reads, which were successfully assembled and annotated. The resulting assembly contains 135,323 transcripts, with 103,396 transcripts (76.4%) annotated with at least one function and encoding 53,142 putative proteins. Due to its close relationship with the hyperaccumulating model species N. cearulescens, it will be possible to derive protein functions from sequence comparisons with this species. Comparisons will highlight common and differing pathways of metal acquisition, storage, and detoxification which will allow us to expand our knowledge of these processes.

Original Leaf Colonisers Shape Fungal Decomposer Communities of Phragmites australis in Intermittent Habitats.

Common reed (Phragmites australis) has high biomass production and is primarily subjected to decomposition processes affected by multiple factors. To predict litter decomposition dynamics in intermittent lakes, it is critical to understand how communities of fungi, as the primary decomposers, form under different habitat conditions. This study reports the shotgun metagenomic sequencing of the initial fungal communities on common reed leaves decomposing under different environmental conditions. We demonstrate that a complex network of fungi forms already on the plant persists into the decomposition phase. Phragmites australis leaves contained at least five fungal phyla, with abundant Ascomycota (95.7%) and Basidiomycota (4.1%), identified as saprotrophs (48.6%), pathotrophs (22.5%), and symbiotrophs (12.6%). Most of the correlations between fungi in fresh and decomposing leaves were identified as co-occurrences (positive correlations). The geographic source of litter and leaf age did not affect the structure and diversity of fungal communities. Keystone taxa were mostly moisture-sensitive. Our results suggest that habitat has a strong effect on the formation of the fungal communities through keystone taxa. Nevertheless, it can also alter the proportions of individual fungal groups in the community through indirect effects on competition between the fungal taxa and their exploitation of favourable conditions.

Grapevine Leafroll-Associated Virus 3 in Single and Mixed Infections Triggers Changes in the Oxidative Balance of Four Grapevine Varieties.

With the aim to characterize changes caused by grapevine leafroll-associated virus 3 (GLRaV-3) singly or in coinfection with other viruses and to potentially determine genotype-specific or common markers of viral infection, thirty-six parameters, including nutrient status, oxidative stress parameters, and primary metabolism as well as symptoms incidence were investigated in 'Cabernet Franc,' 'Merlot,' 'Pinot Noir,' and 'Tribidrag' grapevine varieties. Host responses were characterized by changes in cellular redox state rather than disturbances in nutrient status and primary metabolic processes. Superoxide dismutase, hydrogen peroxide, and proteins were drastically affected regardless of the type of isolate, the host, and the duration of the infection, so they present cellular markers of viral infection. No clear biological pattern could be ascertained for each of the GLRaV-3 genotypes. There is a need to provide a greater understanding of virus epidemiology in viticulture due to the increasing natural disasters and climate change to provide for global food production security. Finding grape varieties that will be able to cope with those changes can aid in this task. Among the studied grapevine varieties, autochthonous 'Tribidrag' seems to be more tolerant to symptoms development despite numerous physiological changes caused by viruses.

1,8-dihydroxy naphthalene (DHN) - melanin confers tolerance to cadmium in isolates of melanised dark septate endophytes.

The contribution of 1,8-dihydroxy naphthalene (DHN) melanin to cadmium (Cd) tolerance in two dark septate endophytes (DSE) of the genus Cadophora with different melanin content was investigated in vitro. The DSE isolate Cad#148 with higher melanin content showed higher tolerance to Cd than the less melanised Cad#149. Melanin synthesis was significantly reduced by Cd in both isolates with uninhibited melanin synthesis, in a dose-dependent manner. Inhibition of melanin synthesis by tricyclazole reduced the relative growth of Cad#148 exposed to Cd and did not affect Cad#149. Cd accumulation was not altered by tricyclazole in the two isolates, but it increased catalase and reduced glutathione reductase activity in more melanised Cad#148, indicating higher stress levels. In contrast, in Cad#149 the enzyme activity was less affected by tricyclazole, indicating a more pronounced role of melanin-independent Cd tolerance mechanisms. Cd ligand environment in fungal mycelia was analysed by extended EXAFS (X-ray absorption fine structure). It revealed that Cd was mainly bound to O- and S-ligands, including hydroxyl, carboxyl, phosphate and thiol groups. A similar proportion of S- and O- ligands (~35% and ~65%) were found in both isolates with uninhibited melanin synthesis. Among O-ligands two types with Cd-O-C- and Cd-O-P- coordination were identified. Tricyclazole altered Cd-O- ligand environment in both fungal isolates by reducing the proportion of Cd-O-C- and increasing the proportion of Cd-O-P coordination. DHN-melanin, among other tolerance mechanisms, significantly contributes to Cd tolerance in more melanised DSE fungi by immobilising Cd to hydroxyl groups and maintaining the integrity of the fungal cell wall.

Dataset on endophytic and rhizoplane fungi on the roots of wild grapevine in Croatia and Bosnia and Herzegovina.

The data presented in this article are related to the research article entitled "Root-associated community composition and co-occurrence patterns of fungi in wild grapevine". This dataset documents the diversity of endophytic and rhizoplane fungi found on the roots of 38 wild grapevine plants growing at four locations (Krka, Neretva, Psunj and Paklenica) in Croatia and Bosnia and Herzegovina. We record here 48 identified fungal operational taxonomic units (OTUs) from 3 phyla, 23 families and 30 genera. The material in this Data in Brief paper comprised the data on the identification of OTUs with their corresponding ecological niche, abundance distribution of each fungal OTU recorded in each of the host and location, restrictedness and with fit to the Sloan neutral community model.

Breeding Buckwheat for Increased Levels of Rutin, Quercetin and Other Bioactive Compounds with Potential Antiviral Effects.

Common buckwheat (Fagopyrum esculentum Moench) and Tartary buckwheat (Fagopyrum tataricum (L.) Gaertn.) are sources of many bioactive compounds, such as rutin, quercetin, emodin, fagopyrin and other (poly)phenolics. In damaged or milled grain under wet conditions, most of the rutin in common and Tartary buckwheat is degraded to quercetin by rutin-degrading enzymes (e.g., rutinosidase). From Tartary buckwheat varieties with low rutinosidase activity it is possible to prepare foods with high levels of rutin, with the preserved initial levels in the grain. The quercetin from rutin degradation in Tartary buckwheat grain is responsible in part for inhibition of α-glucosidase in the intestine, which helps to maintain normal glucose levels in the blood. Rutin and emodin have the potential for antiviral effects. Grain embryos are rich in rutin, so breeding buckwheat with the aim of producing larger embryos may be a promising strategy to increase the levels of rutin in common and Tartary buckwheat grain, and hence to improve its nutritional value.

Genetic Structure and Relationships among Wild and Cultivated Grapevines from Central Europe and Part of the Western Balkan Peninsula.

The genetic diversity and relationship between wild (Vitis vinifera L. subsp. sylvestris (Gmel.) Hegi and cultivated (V. vinifera L. subsp. vinifera) grapevine in the western Balkan region and Central Europe have not been studied together previously, although this area has a rich viticultural past. Here, we studied wild grapevine populations sampled from their natural habitats in several countries of the western Balkan region and Central Europe. Their genetic diversity and structure were compared to cultivars that are traditionally in use in this region. A sample set of 243 accessions was genotyped at 20 nuclear microsatellite loci, including 167 sylvestris and 76 diverse vinifera cultivars. The genetic diversity of the wild grapevines was lower than that of cultivars by all genetic parameters. Both hierarchical and nonhierarchical clustering methods differentiated two main groups, indicating clear separation between wild and cultivated vines but also revealed clear gene flow between the cultivated and wild gene pools through overlaps and admixed ancestry values in the graphs. There was greater affinity to the wild grapes in Central European cultivars than in Balkan cultivars. Fine arrangement of the structure among cultivated grapevines showed differentiation among Central European and Balkan cultivars. These results confirm the divergence of wild grapes from vinifera and highlight the "crossroad" role of the western Balkan peninsula in the broader context of European viticulture.

Effect of phosphorus supply on root traits of two Brassica oleracea L. genotypes.

BACKGROUND: Phosphorus (P) deficiency limits crop production worldwide. Crops differ in their ability to acquire and utilise the P available. The aim of this study was to determine root traits (root exudates, root system architecture (RSA), tissue-specific allocation of P, and gene expression in roots) that (a) play a role in P-use efficiency and (b) contribute to large shoot zinc (Zn) concentration in Brassica oleracea. RESULTS: Two B. oleracea accessions (var. sabellica C6, a kale, and var. italica F103, a broccoli) were grown in a hydroponic system or in a high-throughput-root phenotyping (HTRP) system where they received Low P (0.025 mM) or High P (0.25 mM) supply for 2 weeks. In hydroponics, root and shoot P and Zn concentrations were measured, root exudates were profiled using both Fourier-Transform-Infrared spectroscopy and gas-chromatography-mass spectrometry and previously published RNAseq data from roots was re-examined. In HTRP experiments, RSA (main and lateral root number and lateral root length) was assessed and the tissue-specific distribution of P was determined using micro-particle-induced-X-ray emission. The C6 accession had greater root and shoot biomass than the F103 accession, but the latter had a larger shoot P concentration than the C6 accession, regardless of the P supply in the hydroponic system. The F103 accession had a larger shoot Zn concentration than the C6 accession in the High P treatment. Although the F103 accession had a larger number of lateral roots, which were also longer than in the C6 accession, the C6 accession released a larger quantity and number of polar compounds than the F103 accession. A larger number of P-responsive genes were found in the Low P treatment in roots of the F103 accession than in roots of the C6 accession. Expression of genes linked with "phosphate starvation" was up-regulated, while those linked with iron homeostasis were down-regulated in the Low P treatment. CONCLUSIONS: The results illustrate large within-species variability in root acclimatory responses to P supply in the composition of root exudates, RSA and gene expression, but not in P distribution in root cross sections, enabling P sufficiency in the two B. oleracea accessions studied.

Breeding buckwheat for nutritional quality.

Common buckwheat (Fagopyrum esculentum Moench, CB) and Tartary buckwheat (Fagopyrum tataricum (L.) Gaertn., TB) are used in human nutrition. The idea to screen in the haploid phase for genes affecting low amylose concentration opens the possibility for the effective search of low amylose (waxy) genotypes in CB populations. Self-pollinated homozygous plants of TB might allow us to use a part of endosperm for screening of amylose content. Phenolic substances have a significant inhibitory effect on the digestion of CB and TB proteins, thus metabolites may have impact on protein digestibility. Digestion-resistant peptides are largely responsible for the bile acid elimination. Breeding to diminish polyphenols and anti-nutritional substances might have negative effects on the resistance of plants against pests, diseases and UV-radiation. Bread and pasta are popular CB and TB dishes. During dough making most of CB or TB rutin is degraded to quercetin by rutin-degrading enzymes. The new trace-rutinosidase TB variety makes possible making TB bread with considerable amount of rutin, preserving the initial rutin from flour. Breeding CB and TB for larger embryos would make it possible to increase protein, rutin, and essential minerals concentration in CB and TB grain.

Comparison of lovastatin, citrinin and pigment production of different Monascus purpureus strains grown on rice and millet.

Six Monascus purpureus (red mould) strains were cultivated on brown rice and millet as substrates. They underwent strain selection for high lovastatin and pigment production, and low citrinin mycotoxin production, with particular reference to potential for millet as substrate. For most of these strains, substrate dry matter loss was 54-60% on rice and 46-48% on millet, although the 'MOPU GS1' strain showed 18% and 17% dry matter loss, respectively. 'MOPU GS1' was also the only strain with detectable levels of lovastatin (1.3 and 1.6 mg lovastatin/g substrate dry weight [dw], respectively) and citrinin under the limit of detection. In the other strains, citrinin varied from 0.3 to 18.2 mg/g substrate dw. Among the six strains, 'EBY3' provided high pigment production when grown on rice, although it produced 1.1 mg citrinin/g substrate dw. Millet showed good potential as an alternative substrate to rice, due to higher lovastatin and lower citrinin production; however, rice was the better substrate for production of M. purpureus pigments.

Links Between Genetic Groups, Host Specificity, and Ergot-Alkaloid Profiles within Claviceps purpurea (Fr.) Tul. on Slovenian Grasses.

In the present study, the genetic relationships and ergot-alkaloid production of the fungus Claviceps purpurea on grasses were investigated, to determine any associations between grass host specificity, ergot-alkaloid production, and geographic origin. C. purpurea sclerotia were obtained from wild and cultivated grasses along a 300-km climatic gradient, from sub-Mediterranean to continental climates. Twenty-one infected grass samples provided 39 sclerotia for analysis of the ergot alkaloids ergometrine, ergosine, ergotamine, ergocornine, ergocryptine, and ergocristine, and their "-inine" epimers, using liquid chromatography-tandem mass spectrometry. C. purpurea ribosomal DNA underwent molecular classification to determine any grass host or geographic specificity of ergot-alkaloid composition for the different operational taxonomic units. Molecular analysis of sclerotia ribosomal DNA showed three genetic groups, with some associations with specific grass host taxonomic groups. The ergot-alkaloid composition data were in agreement with the data obtained by molecular methods. The most frequent ergot-alkaloid epimers were ergocristine, and ergosine. The total ergot-alkaloid concentrations in sclerotia varied from 59 to 4,200 mg kg-1, which corresponds to 0.059 to 4.2 mg kg-1 in animal feed (assuming ergot alkaloids at 1,000 mg kg-1 sclerotia). Therefore, grasses can be associated with significant levels of ergot alkaloids. In addition, the ergot-alkaloid compositions of C. purpurea sclerotia can be different for infections with different C. purpurea genetic groups, because these show different ergot-alkaloid compositions.

Hypoxia and inactivity related physiological changes precede or take place in absence of significant rearrangements in bacterial community structure: The PlanHab randomized trial pilot study.

We explored the assembly of intestinal microbiota in healthy male participants during the randomized crossover design of run-in (5 day) and experimental phases (21-day normoxic bed rest (NBR), hypoxic bed rest (HBR) and hypoxic ambulation (HAmb) in a strictly controlled laboratory environment, with balanced fluid and dietary intakes, controlled circadian rhythm, microbial ambiental burden and 24/7 medical surveillance. The fraction of inspired O2 (FiO2) and partial pressure of inspired O2 (PiO2) were 0.209 and 133.1 ± 0.3 mmHg for NBR and 0.141 ± 0.004 and 90.0 ± 0.4 mmHg for both hypoxic variants (HBR and HAmb; ~4000 m simulated altitude), respectively. A number of parameters linked to intestinal environment such as defecation frequency, intestinal electrical conductivity (IEC), sterol and polyphenol content and diversity, indole, aromaticity and spectral characteristics of dissolved organic matter (DOM) were measured (64 variables). The structure and diversity of bacterial microbial community was assessed using 16S rRNA amplicon sequencing. Inactivity negatively affected frequency of defecation and in combination with hypoxia increased IEC (p < 0.05). In contrast, sterol and polyphenol diversity and content, various characteristics of DOM and aromatic compounds, the structure and diversity of bacterial microbial community were not significantly affected over time. A new in-house PlanHab database was established to integrate all measured variables on host physiology, diet, experiment, immune and metabolic markers (n = 231). The observed progressive decrease in defecation frequency and concomitant increase in IEC suggested that the transition from healthy physiological state towards the developed symptoms of low magnitude obesity-related syndromes was dose dependent on the extent of time spent in inactivity and preceded or took place in absence of significant rearrangements in bacterial microbial community. Species B. thetaiotamicron, B. fragilis, B. dorei and other Bacteroides with reported relevance for dysbiotic medical conditions were significantly enriched in HBR, characterized with most severe inflammation symptoms, indicating a shift towards host mucin degradation and proinflammatory immune crosstalk.

Phenolic Responses to Esca-Associated Fungi in Differently Decayed Grapevine Woods from Different Trunk Parts of 'Cabernet Sauvignon'.

Alterations in phenolic contents were studied in Esca symptomatic (Sym) and asymptomatic (Asym) vines of 'Cabernet Sauvignon' based on wood conditions (healthy, HLT; necrotic, Nec; and rotten, Rot) and vine parts (head, trunk, and rootstock). In Asym vines, only Alternaria alternate was identified in Nec wood, while the HLT wood of Sym vines was colonized by Botryospaeriaceae sp. and Aureobasidium pullulans, Nec wood by Fomitiporia mediterranea, and Rot wood by Fomitiporia mediterranea and Phaeomoniella chlamydospora. Esca infection caused a significant accumulation of gallic acid, total flavanols, stilbenes (STB), and total analyzed phenolics (TAP) in all studied woods, especially in Nec wood. In Asym vines, TAP in the head increased with necrosis, but in Sym it decreased, while TAP in the trunk and rootstock of Sym showed an opposite response. The significantly highest contents of procyanidins (Pcys), catechin, epicatechin, epicatechin gallates, and Pcys dimers and tetramers were measured in HLT wood in the head and in Nec wood in the trunk of Sym vines. The significant increase of STB content was not caused only by Esca infection in HLT wood but also by necrosis in Asym vines, especially of ε-viniferin glucoside, resveratrol glycosides, and astringin. The obtained results suggest that the alteration in phenolics differed not only due to Esca infection but also due to the wood conditions and vine part, which might reflect the impact of the duration of the presence of the pathogen in different parts of the vine.

Temporal changes in fungal communities from buckwheat seeds and their effects on seed germination and seedling secondary metabolism.

Seed-associated fungal communities affect multiple parameters of seed quality at all stages of production, from seed development to post-harvest storage and germination. We therefore investigated the diversity and dynamics of fungal communities in the seeds of common buckwheat (Fagopyrum esculentum) and Tartary buckwheat (F. tataricum) from harvest to 1 y of storage. Fungal populations in seeds were relatively stable, comprised mainly of field fungi. Incidence of fungi was most likely determined by fungal interspecies direct interactions, as well as by their synthesis of volatile organic compounds. Most prominent antagonistic interactions were seen for two plant pathogens, Alternaria alternata on Botrytis cinerea. Detrimental effects of the fungi on seed germination and seedling development were related to fungal extracellular enzyme activity, and in particular to amylase, cellulase and, polyphenol oxidase. Polyphenol and tannin concentrations in buckwheat seedlings were related to fungal growth rate and intensity of fungal cellulase activity, respectively, which suggests that physical penetration of the fungi through the host tissues is probably the stimulus for the activation of plant defence reactions in these seedlings.

Distribution and diversity of arbuscular mycorrhizal fungi in grapevines from production vineyards along the eastern Adriatic coast.

The colonisation and diversity of arbuscular mycorrhizal fungi (AMF) on roots of grapevines were investigated in production vineyards located along a 500-km-long stretch of karst along the coast of the Adriatic Sea. AMF communities on roots of grapevines were analysed using temporal temperature gel electrophoresis and sequencing of the 18S and internal transcribed spacer segments of the rDNA operon. The AMF colonisation of these grapevines roots was consistent along the whole of this east Adriatic karst region, at 64 to 82% of fine roots. The comparison of the AMF communities on the roots of these grapevines showed that the fungal community associated with grapevine roots seems to be relatively stable, with inter-vineyard variability comparable to intra-vineyard variability. Some of the changes in the fungal communities were attributed to environmental factors (plant-available P) and location of the vineyard, although the latter could also have been influenced by an unmeasured environmental factor. A total of 27 taxa of fungi were identified, including taxa from Glomus group B, based on the sequencing of 18S rDNA. Sequencing of the internal transcribed spacer rDNA yielded 30 different fungal taxa, which comprised eight different Glomeromycota taxa, including Glomus sinuosum and Glomus indicum. To our knowledge, this is the first report of grapevine colonisation by G. indicum.

Mycorrhizal status and diversity of fungal endophytes in roots of common buckwheat (Fagopyrum esculentum) and tartary buckwheat (F. tataricum).

To determine the mycorrhizal status and to identify the fungi colonising the roots of the plants, common buckwheat (Fagopyrum esculentum) and tartary buckwheat (F. tataricum) were inoculated with an indigenous fungal mixture from a buckwheat field. Root colonisation was characterised by the hyphae and distinct microsclerotia of dark septate endophytes, with occasional arbuscules and vesicles of arbuscular mycorrhizal fungi. Sequences of arbuscular mycorrhizal fungi colonising tartary buckwheat clustered close to the Glomus species group A. Sequences with similarity to the Ceratobasidium/Rhizoctonia complex, a putative dark septate endophyte fungus, were amplified from the roots of both common and tartary buckwheat. To the best of our knowledge, this is the first report of arbuscular mycorrhizal colonisation in tartary buckwheat and the first molecular characterisation of these fungi that can colonise both of these economically important plant species.