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April 21, 2020  |  

Complete genome sequence of Caulobacter flavus RHGG3T, a type species of the genus Caulobacter with plant growth-promoting traits and heavy metal resistance.

Caulobacter flavus RHGG3T, a novel type species in the genus Caulobacter, originally isolated from rhizosphere soil of watermelon (Citrullus lanatus), has the ability to improve the growth of watermelon seedling and tolerate heavy metals. In vitro, C. flavus RHGG3T was able to solubilize phosphate (80.56 mg L-1), produce indole-3-acetic acid (IAA) (11.58 mg L-1) and was resistant to multiple heavy metals (copper, zinc, cadmium, cobalt and lead). Inoculating watermelon with this strain increased shoot and root length by 22.1% and 43.7%, respectively, and the total number of lateral roots by 55.9% compared to non-inoculated watermelon. In this study, we present the complete genome sequence of C. flavus RHGG3T, which was comprised of a single circular chromosome of 5,659,202 bp with a G?+?C content of 69.25%. An annotation analysis revealed that the C. flavus RHGG3T genome contained 5172 coding DNA sequences, 9 rRNA and 55 tRNA genes. Genes related to plant growth promotion (PGP), such as those associated with phosphate solubilization, nitrogen fixation, IAA, phenazine, volatile compounds, spermidine and cobalamin synthesis, were found in the C. flavus RHGG3T genome. Some genes responsible for heavy metal tolerance were also identified. The genome sequence of strain RHGG3T reported here provides new insight into the molecular mechanisms underlying the promotion of plant growth and the resistance to heavy metals in C. flavus. This study will be valuable for further exploration of the biotechnological applications of strain RHGG3T in agriculture.

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September 22, 2019  |  

Is there foul play in the leaf pocket? The metagenome of floating fern Azolla reveals endophytes that do not fix N2 but may denitrify.

Dinitrogen fixation by Nostoc azollae residing in specialized leaf pockets supports prolific growth of the floating fern Azolla filiculoides. To evaluate contributions by further microorganisms, the A. filiculoides microbiome and nitrogen metabolism in bacteria persistently associated with Azolla ferns were characterized. A metagenomic approach was taken complemented by detection of N2 O released and nitrogen isotope determinations of fern biomass. Ribosomal RNA genes in sequenced DNA of natural ferns, their enriched leaf pockets and water filtrate from the surrounding ditch established that bacteria of A. filiculoides differed entirely from surrounding water and revealed species of the order Rhizobiales. Analyses of seven cultivated Azolla species confirmed persistent association with Rhizobiales. Two distinct nearly full-length Rhizobiales genomes were identified in leaf-pocket-enriched samples from ditch grown A. filiculoides. Their annotation revealed genes for denitrification but not N2 -fixation. 15 N2 incorporation was active in ferns with N. azollae but not in ferns without. N2 O was not detectably released from surface-sterilized ferns with the Rhizobiales. N2 -fixing N. azollae, we conclude, dominated the microbiome of Azolla ferns. The persistent but less abundant heterotrophic Rhizobiales bacteria possibly contributed to lowering O2 levels in leaf pockets but did not release detectable amounts of the strong greenhouse gas N2 O.© 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

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September 22, 2019  |  

Comprehensive exploration of the rumen microbial ecosystem with advancements in metagenomics

Ruminant farming and its environmental impact has long remained an economic concern. Metagenomics unravel the vast structural and functional diversity of the rumen microbial community that plays a major role in animal nutrition. Hereby, we summarize rumen metagenomic studies that have enhanced the knowledge of rumen microbe dynamics subsequently leading to development of better feed strategies to improve livestock production and reduce methane emissions.

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September 22, 2019  |  

Lack of thinning effects over inter-annual changes in soil fungal community and diversity in a Mediterranean pine forest

Predicted changes in global climate might negatively affect the soil microbiome and associated ecosystem processes in Mediterranean forests. Forest treatments, such as forest thinning, have been suggested to mitigate climate change impacts on vegetation by reducing competition between trees, thus increasing water availability. Studies addressing the combined effects of climate and forest thinning on belowground fungal communities are still scarce, being fundamental to elaborate adaptive strategies to global warming. The aim of this study was to evaluate the tree density reduction effects on soil fungal communities and their response to inter-annual changes in weather conditions. The temporal dynamics of soil fungal communities in relation to these two drivers (i.e., forest management and weather conditions) were studied from 2009 until 2014 in a set of 12 pairs of thinned and un-thinned plots dominated by Pinus pinaster Ait. Thinning (from 30% up to 70% reduction in stand basal area) was conducted in 2009 and soil fungal community composition was studied during 4?years. Here, we used autumn precipitation and temperature to describe the impact of inter-annual weather changes. We used Pacific Biosciences sequencing of fungal ITS2 amplicons to study fungal communities in soil samples. Forest thinning did not significantly affect fungal community composition nor fungal species richness and diversity, indicating that the soil fungal community is resistant to forest thinning regardless of its intensity. However, fungal species composition changed progressively across years, both at the species level and with regards to functional guilds. These changes in community composition were partly driven by inter-annual variation in precipitation and temperature, with free-living fungi increasing in abundance under wetter conditions, and symbiotic fungi being more prominent under drier and colder conditions. The results indicate that mycorrhizal communities in Mediterranean forest ecosystems can resist forest thinning, if enough trees and functional roots from thinned trees are retained.

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September 22, 2019  |  

A comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms.

Several fungi-specific primers target the 18S rRNA gene sequence, one of the prominent markers for fungal classification. The design of most primers goes back to the last decades. Since then, the number of sequences in public databases increased leading to the discovery of new fungal groups and changes in fungal taxonomy. However, no reevaluation of primers was carried out and relevant information on most primers is missing. With this study, we aimed to develop an 18S rRNA gene sequence primer toolkit allowing an easy selection of the best primer pair appropriate for different sequencing platforms, research aims (biodiversity assessment versus isolate classification) and target groups.We performed an intensive literature research, reshuffled existing primers into new pairs, designed new Illumina-primers, and annealing blocking oligonucleotides. A final number of 439 primer pairs were subjected to in silico PCRs. Best primer pairs were selected and experimentally tested. The most promising primer pair with a small amplicon size, nu-SSU-1333-5’/nu-SSU-1647-3′ (FF390/FR-1), was successful in describing fungal communities by Illumina sequencing. Results were confirmed by a simultaneous metagenomics and eukaryote-specific primer approach. Co-amplification occurred in all sample types but was effectively reduced by blocking oligonucleotides.The compiled data revealed the presence of an enormous diversity of fungal 18S rRNA gene primer pairs in terms of fungal coverage, phylum spectrum and co-amplification. Therefore, the primer pair has to be carefully selected to fulfill the requirements of the individual research projects. The presented primer toolkit offers comprehensive lists of 164 primers, 439 primer combinations, 4 blocking oligonucleotides, and top primer pairs holding all relevant information including primer’s characteristics and performance to facilitate primer pair selection.

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September 22, 2019  |  

Complete genome sequence of Paenibacillus polymyxa YC0136, a plant growth–promoting rhizobacterium isolated from tobacco rhizosphere.

Paenibacillus polymyxa strain YC0136 is a plant growth-promoting rhizobacterium with antimicrobial activity, which was isolated from tobacco rhizosphere. Here, we report the complete genome sequence of P. polymyxa YC0136. Several genes with antifungal and antibacterial activity were discovered. Copyright © 2017 Liu et al.

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September 22, 2019  |  

Community profiling of Fusarium in combination with other plant associated fungi in different crop species using SMRT Sequencing.

Fusarium head blight, caused by fungi from the genus Fusarium, is one of the most harmful cereal diseases, resulting not only in severe yield losses but also in mycotoxin contaminated and health-threatening grains. Fusarium head blight is caused by a diverse set of species that have different host ranges, mycotoxin profiles and responses to agricultural practices. Thus, understanding the composition of Fusarium communities in the field is crucial for estimating their impact and also for the development of effective control measures. Up to now, most molecular tools that monitor Fusarium communities on plants are limited to certain species and do not distinguish other plant associated fungi. To close these gaps, we developed a sequencing-based community profiling methodology for crop-associated fungi with a focus on the genus Fusarium. By analyzing a 1600 bp long amplicon spanning the highly variable segments ITS and D1-D3 of the ribosomal operon by PacBio SMRT sequencing, we were able to robustly quantify Fusarium down to species level through clustering against reference sequences. The newly developed methodology was successfully validated in mock communities and provided similar results as the culture-based assessment of Fusarium communities by seed health tests in grain samples from different crop species. Finally, we exemplified the newly developed methodology in a field experiment with a wheat-maize crop sequence under different cover crop and tillage regimes. We analyzed wheat straw residues, cover crop shoots and maize grains and we could reveal that the cover crop hairy vetch (Vicia villosa) acts as a potent alternative host for Fusarium (OTU F.ave/tri) showing an eightfold higher relative abundance compared with other cover crop treatments. Moreover, as the newly developed methodology also allows to trace other crop-associated fungi, we found that vetch and green fallow hosted further fungal plant pathogens including Zymoseptoria tritici. Thus, besides their beneficial traits, cover crops can also entail phytopathological risks by acting as alternative hosts for Fusarium and other noxious plant pathogens. The newly developed sequencing based methodology is a powerful diagnostic tool to trace Fusarium in combination with other fungi associated to different crop species.

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September 22, 2019  |  

Contrasting distribution patterns between aquatic and terrestrial Phytophthora species along a climatic gradient are linked to functional traits.

Diversity of microbial organisms is linked to global climatic gradients. The genus Phytophthora includes both aquatic and terrestrial plant pathogenic species that display a large variation of functional traits. The extent to which the physical environment (water or soil) modulates the interaction of microorganisms with climate is unknown. Here, we explored the main environmental drivers of diversity and functional trait composition of Phytophthora communities. Communities were obtained by a novel metabarcoding setup based on PacBio sequencing of river filtrates in 96 river sites along a geographical gradient. Species were classified as terrestrial or aquatic based on their phylogenetic clade. Overall, terrestrial and aquatic species showed contrasting patterns of diversity. For terrestrial species, precipitation was a stronger driver than temperature, and diversity and functional diversity decreased with decreasing temperature and precipitation. In cold and dry areas, the dominant species formed resistant structures and had a low optimum temperature. By contrast, for aquatic species, temperature and water chemistry were the strongest drivers, and diversity increased with decreasing temperature and precipitation. Within the same area, environmental filtering affected terrestrial species more strongly than aquatic species (20% versus 3% of the studied communities, respectively). Our results highlight the importance of functional traits and the physical environment in which microorganisms develop their life cycle when predicting their distribution under changing climatic conditions. Temperature and rainfall may be buffered differently by water and soil, and thus pose contrasting constrains to microbial assemblies.

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September 22, 2019  |  

Impacts of experimentally accelerated forest succession on belowground plant and fungal communities

Understanding how soil processes, belowground plant and fungal species composition, and nutrient cycles are altered by disturbances is essential for understanding the role forests play in mitigating global climate change. Here we ask: How are root and fungal communities altered in a mid-successional forest during shifts in dominant tree species composition? This study utilizes the Forest Accelerated Succession ExperimenT (FASET) at the University of Michigan Biological Station (UMBS) as a platform for addressing this question. FASET consists of a 39-ha treatment in which all mature early successional aspen (Populus spp.) and paper birch (Betula papyrifera) were killed by stem-girdling in 2008. Four years after girdling, neither overall fungal diversity indices, plant diversity indices, nor root biomass differed between girdled (treated) and non-girdled (reference) stands. However, experimental advancement of succession by removal of aspen and birch resulted in 1) a shift in fungal functional groups, with significantly less ectomycorrhizal fungi, 2) a trend toward less arbuscular mycorrhizal fungi, and 3) a significant increase in the proportion of saprotrophs in girdled stands. In addition to shifts in functional groups between treated and untreated stands, ectomycorrhizal fungi proportions were negatively correlated with NH4+ and total dissolved inorganic nitrogen (DIN) in soil. This research illustrates the propensity for disturbances in forest ecosystems to shift fungal community composition, which has implications for carbon storage and nutrient cycling in soils under future climate scenarios.

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September 22, 2019  |  

Soil microbial communities are shaped by plant-driven changes in resource availability during secondary succession.

Although we understand the ecological processes eliciting changes in plant community composition during secondary succession, we do not understand whether co-occurring changes in plant detritus shape saprotrophic microbial communities in soil. In this study, we investigated soil microbial composition and function across an old-field chronosequence ranging from 16 to 86 years following agricultural abandonment, as well as three forests representing potential late-successional ecosystems. Fungal and bacterial community composition was quantified from ribosomal DNA, and insight into the functional potential of the microbial community to decay plant litter was gained from shotgun metagenomics and extracellular enzyme assays. Accumulation of soil organic matter across the chronosequence exerted a positive and significant effect on fungal phylogenetic ß-diversity and the activity of extracellular enzymes with lignocellulolytic activity. In addition, the increasing abundance of lignin-rich C4 grasses was positively related to the composition of fungal genes with lignocellulolytic function, thereby linking plant community composition, litter biochemistry, and microbial community function. However, edaphic properties were the primary agent shaping bacterial communities, as bacterial ß-diversity and variation in functional gene composition displayed a significant and positive relationship to soil pH across the chronosequence. The late-successional forests were compositionally distinct from the oldest old fields, indicating that substantial changes occur in soil microbial communities as old fields give way to forests. Taken together, our observations demonstrate that plants govern the turnover of soil fungal communities and functional characteristics during secondary succession, due to the continual input of detritus and differences in litter biochemistry among plant species.

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September 22, 2019  |  

A community-based culture collection for targeting novel plant growth-promoting bacteria from the sugarcane microbiome.

The soil-plant ecosystem harbors an immense microbial diversity that challenges investigative approaches to study traits underlying plant-microbe association. Studies solely based on culture-dependent techniques have overlooked most microbial diversity. Here we describe the concomitant use of culture-dependent and -independent techniques to target plant-beneficial microbial groups from the sugarcane microbiome. The community-based culture collection (CBC) approach was used to access microbes from roots and stalks. The CBC recovered 399 unique bacteria representing 15.9% of the rhizosphere core microbiome and 61.6-65.3% of the endophytic core microbiomes of stalks. By cross-referencing the CBC (culture-dependent) with the sugarcane microbiome profile (culture-independent), we designed a synthetic community comprised of naturally occurring highly abundant bacterial groups from roots and stalks, most of which has been poorly explored so far. We then used maize as a model to probe the abundance-based synthetic inoculant. We show that when inoculated in maize plants, members of the synthetic community efficiently colonize plant organs, displace the natural microbiota and dominate at 53.9% of the rhizosphere microbial abundance. As a result, inoculated plants increased biomass by 3.4-fold as compared to uninoculated plants. The results demonstrate that abundance-based synthetic inoculants can be successfully applied to recover beneficial plant microbes from plant microbiota.

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September 22, 2019  |  

Molecular mechanisms of acclimatization to phosphorus starvation and recovery underlying full-length transcriptome profiling in barley (Hordeum vulgare L.).

A lack of phosphorus (P) in plants can severely constrain growth and development. Barley, one of the earliest domesticated crops, is extensively planted in poor soil around the world. To date, the molecular mechanisms of enduring low phosphorus, at the transcriptional level, in barley are still unclear. In the present study, two different barley genotypes (GN121 and GN42)-with contrasting phosphorus efficiency-were used to reveal adaptations to low phosphorus stress, at three time points, at the morphological, physiological, biochemical, and transcriptome level. GN121 growth was less affected by phosphorus starvation and recovery than that of GN42. The biomass and inorganic phosphorus concentration of GN121 and GN42 declined under the low phosphorus-induced stress and increased after recovery with normal phosphorus. However, the range of these parameters was higher in GN42 than in GN121. Subsequently, a more complete genome annotation was obtained by correcting with the data sequenced on Illumina HiSeq X 10 and PacBio RSII SMRT platform. A total of 6,182 and 5,270 differentially expressed genes (DEGs) were identified in GN121 and GN42, respectively. The majority of these DEGs were involved in phosphorus metabolism such as phospholipid degradation, hydrolysis of phosphoric enzymes, sucrose synthesis, phosphorylation/dephosphorylation and post-transcriptional regulation; expression of these genes was significantly different between GN121 and GN42. Specifically, six and seven DEGs were annotated as phosphorus transporters in roots and leaves, respectively. Furthermore, a putative model was constructed relying on key metabolic pathways related to phosphorus to illustrate the higher phosphorus efficiency of GN121 compared to GN42 under low phosphorus conditions. Results from this study provide a multi-transcriptome database and candidate genes for further study on phosphorus use efficiency (PUE).

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September 22, 2019  |  

The microbiome of the leaf surface of Arabidopsis protects against a fungal pathogen.

We have explored the importance of the phyllosphere microbiome in plant resistance in the cuticle mutants bdg (BODYGUARD) or lacs2.3 (LONG CHAIN FATTY ACID SYNTHASE 2) that are strongly resistant to the fungal pathogen Botrytis cinerea. The study includes infection of plants under sterile conditions, 16S ribosomal DNA sequencing of the phyllosphere microbiome, and isolation and high coverage sequencing of bacteria from the phyllosphere. When inoculated under sterile conditions bdg became as susceptible as wild-type (WT) plants whereas lacs2.3 mutants retained the resistance. Adding washes of its phyllosphere microbiome could restore the resistance of bdg mutants, whereas the resistance of lacs2.3 results from endogenous mechanisms. The phyllosphere microbiome showed distinct populations in WT plants compared to cuticle mutants. One species identified as Pseudomonas sp isolated from the microbiome of bdg provided resistance to B. cinerea on Arabidopsis thaliana as well as on apple fruits. No direct activity was observed against B. cinerea and the action of the bacterium required the plant. Thus, microbes present on the plant surface contribute to the resistance to B. cinerea. These results open new perspectives on the function of the leaf microbiome in the protection of plants.© 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

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September 22, 2019  |  

Ecological genomics of tropical trees: how local population size and allelic diversity of resistance genes relate to immune responses, cosusceptibility to pathogens, and negative density dependence

In tropical forests, rarer species show increased sensitivity to species-specific soil pathogens and more negative effects of conspecific density on seedling survival (NDD). These patterns suggest a connection between ecology and immunity, perhaps because small population size disproportionately reduces genetic diversity of hyperdiverse loci such as immunity genes. In an experiment examining seedling roots from six species in one tropical tree community, we found that smaller populations have reduced amino acid diversity in pathogen resistance (R) genes but not the transcriptome in general. Normalized R gene amino acid diversity varied with local abundance and prior measures of differences in sensitivity to conspecific soil and NDD. After exposure to live soil, species with lower R gene diversity had reduced defence gene induction, more cosusceptibility of maternal cohorts to colonization by potentially pathogenic fungi, reduced root growth arrest (an R gene-mediated response) and their root-associated fungi showed lower induction of self-defence (antioxidants). Local abundance was not related to the ability to induce immune responses when pathogen recognition was bypassed by application of salicylic acid, a phytohormone that activates defence responses downstream of R gene signalling. These initial results support the hypothesis that smaller local tree populations have reduced R gene diversity and recognition-dependent immune responses, along with greater cosusceptibility to species-specific pathogens that may facilitate disease transmission and NDD. Locally rare species may be less able to increase their equilibrium abundance without genetic boosts to defence via immigration of novel R gene alleles from a larger and more diverse regional population.

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September 22, 2019  |  

Soil microclimate changes affect soil fungal communities in a Mediterranean pine forest.

Soil microclimate is a potentially important regulator of the composition of plant-associated fungal communities in climates with significant drought periods. Here, we investigated the spatio-temporal dynamics of soil fungal communities in a Mediterranean Pinus pinaster forest in relation to soil moisture and temperature. Fungal communities in 336 soil samples collected monthly over 1 year from 28 long-term experimental plots were assessed by PacBio sequencing of ITS2 amplicons. Total fungal biomass was estimated by analysing ergosterol. Community changes were analysed in the context of functional traits. Soil fungal biomass was lowest during summer and late winter and highest during autumn, concurrent with a greater relative abundance of mycorrhizal species. Intra-annual spatio-temporal changes in community composition correlated significantly with soil moisture and temperature. Mycorrhizal fungi were less affected by summer drought than free-living fungi. In particular, mycorrhizal species of the short-distance exploration type increased in relative abundance under dry conditions, whereas species of the long-distance exploration type were more abundant under wetter conditions. Our observations demonstrate a potential for compositional and functional shifts in fungal communities in response to changing climatic conditions. Free-living fungi and mycorrhizal species with extensive mycelia may be negatively affected by increasing drought periods in Mediterranean forest ecosystems.© 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.

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