A synthetic microbial community used as a bioinoculant can overcome rice production constraints in acid soils.

A customised synthetic microbial community (SynCom) composed of carefully selected rhizosphere-competent bacterial strains improved rice growth, yield and resistance to soil acidity and Al toxicity.

Stable-Isotope-Informed, Genome-Resolved Metagenomics Uncovers Potential Cross-Kingdom Interactions in Rhizosphere Soil.

The functioning, health, and productivity of soil are intimately tied to a complex network of interactions, particularly in plant root-associated rhizosphere soil. We conducted a stable-isotope-informed, genome-resolved metagenomic study to trace carbon from Avena fatua grown in a 13CO2 atmosphere into soil. We collected paired rhizosphere and nonrhizosphere soil at 6 and 9 weeks of plant growth and extracted DNA that was then separated by density using ultracentrifugation. Thirty-two fractions from each of five samples were grouped by density, sequenced, assembled, and binned to generate 55 unique bacterial genomes that were ≥70% complete. We also identified complete 18S rRNA sequences of several 13C-enriched microeukaryotic bacterivores and fungi. We generated 10 circularized bacteriophage (phage) genomes, some of which were the most labeled entities in the rhizosphere, suggesting that phage may be important agents of turnover of plant-derived C in soil. CRISPR locus targeting connected one of these phage to a Burkholderiales host predicted to be a plant pathogen. Another highly labeled phage is predicted to replicate in a Catenulispora sp., a possible plant growth-promoting bacterium. We searched the genome bins for traits known to be used in interactions involving bacteria, microeukaryotes, and plant roots and found DNA from heavily 13C-labeled bacterial genes thought to be involved in modulating plant signaling hormones, plant pathogenicity, and defense against microeukaryote grazing. Stable-isotope-informed, genome-resolved metagenomics indicated that phage can be important agents of turnover of plant-derived carbon in soil. IMPORTANCE Plants grow in intimate association with soil microbial communities; these microbes can facilitate the availability of essential resources to plants. Thus, plant productivity commonly depends on interactions with rhizosphere bacteria, viruses, and eukaryotes. Our work is significant because we identified the organisms that took up plant-derived organic C in rhizosphere soil and determined that many of the active bacteria are plant pathogens or can impact plant growth via hormone modulation. Further, by showing that bacteriophage accumulate CO2-derived carbon, we demonstrated their vital roles in redistribution of plant-derived C into the soil environment through bacterial cell lysis. The use of stable-isotope probing (SIP) to identify consumption (or lack thereof) of root-derived C by key microbial community members within highly complex microbial communities opens the way for assessing manipulations of bacteria and phage with potentially beneficial and detrimental traits, ultimately providing a path to improved plant health and soil carbon storage.

Fungal-Bacterial Cooccurrence Patterns Differ between Arbuscular Mycorrhizal Fungi and Nonmycorrhizal Fungi across Soil Niches.

Soil bacteria and fungi are known to form niche-specific communities that differ between actively growing and decaying roots. Yet almost nothing is known about the cross-kingdom interactions that frame these communities and the environmental filtering that defines these potentially friendly or competing neighbors. We explored the temporal and spatial patterns of soil fungal (mycorrhizal and nonmycorrhizal) and bacterial cooccurrence near roots of wild oat grass, Avena fatua, growing in its naturalized soil in a greenhouse experiment. Amplicon sequences of the fungal internal transcribed spacer (ITS) and bacterial 16S rRNA genes from rhizosphere and bulk soils collected at multiple plant growth stages were used to construct covariation-based networks as a step toward identifying fungal-bacterial associations. Corresponding stable-isotope-enabled metagenome-assembled genomes (MAGs) of bacteria identified in cooccurrence networks were used to inform potential mechanisms underlying the observed links. Bacterial-fungal networks were significantly different in rhizosphere versus bulk soils and between arbuscular mycorrhizal fungi (AMF) and nonmycorrhizal fungi. Over 12 weeks of plant growth, nonmycorrhizal fungi formed increasingly complex networks with bacteria in rhizosphere soils, while AMF more frequently formed networks with bacteria in bulk soils. Analysis of network-associated bacterial MAGs suggests that some of the fungal-bacterial links that we identified are potential indicators of bacterial breakdown and consumption of fungal biomass, while others intimate shared ecological niches.IMPORTANCE Soils near living and decomposing roots form distinct niches that promote microorganisms with distinctive environmental preferences and interactions. Yet few studies have assessed the community-level cooccurrence of bacteria and fungi in these soil niches as plant roots grow and senesce. With plant growth, we observed increasingly complex cooccurrence networks between nonmycorrhizal fungi and bacteria in the rhizosphere, while mycorrhizal fungal (AMF) and bacterial cooccurrence was more pronounced in soil further from roots, in the presence of decaying root litter. This rarely documented phenomenon suggests niche sharing of nonmycorrhizal fungi and bacteria, versus niche partitioning between AMF and bacteria; both patterns are likely driven by C substrate availability and quality. Although the implications of species cooccurrence are fiercely debated, MAGs matching the bacterial nodes in our networks possess the functional potential to interact with the fungi that they are linked to, suggesting an ecological significance of fungal-bacterial cooccurrence patterns.

Microbial functional genes commonly respond to elevated carbon dioxide.

Atmospheric CO2 concentration is increasing, largely due to anthropogenic activities. Previous studies of individual free-air CO2 enrichment (FACE) experimental sites have shown significant impacts of elevated CO2 (eCO2) on soil microbial communities; however, no common microbial response patterns have yet emerged, challenging our ability to predict ecosystem functioning and sustainability in the future eCO2 environment. Here we analyzed 66 soil microbial communities from five FACE sites, and showed common microbial response patterns to eCO2, especially for key functional genes involved in carbon and nitrogen fixation (e.g., pcc/acc for carbon fixation, nifH for nitrogen fixation), carbon decomposition (e.g., amyA and pulA for labile carbon decomposition, mnp and lcc for recalcitrant carbon decomposition), and greenhouse gas emissions (e.g., mcrA for methane production, norB for nitrous oxide production) across five FACE sites. Also, the relative abundance of those key genes was generally increased and directionally associated with increased biomass, soil carbon decomposition, and soil moisture. In addition, a further literature survey of more disparate FACE experimental sites indicated increased biomass, soil carbon decay, nitrogen fixation, methane and nitrous oxide emissions, plant and soil carbon and nitrogen under eCO2. A conceptual framework was developed to link commonly responsive functional genes with ecosystem processes, such as pcc/acc vs. soil carbon storage, amyA/pulA/mnp/lcc vs. soil carbon decomposition, and nifH vs. nitrogen availability, suggesting that such common responses of microbial functional genes may have the potential to predict ecosystem functioning and sustainability in the future eCO2 environment.

Stable isotope informed genome-resolved metagenomics reveals that Saccharibacteria utilize microbially-processed plant-derived carbon.

BACKGROUND: The transformation of plant photosynthate into soil organic carbon and its recycling to CO2 by soil microorganisms is one of the central components of the terrestrial carbon cycle. There are currently large knowledge gaps related to which soil-associated microorganisms take up plant carbon in the rhizosphere and the fate of that carbon. RESULTS: We conducted an experiment in which common wild oats (Avena fatua) were grown in a 13CO2 atmosphere and the rhizosphere and non-rhizosphere soil was sampled for genomic analyses. Density gradient centrifugation of DNA extracted from soil samples enabled distinction of microbes that did and did not incorporate the 13C into their DNA. A 1.45-Mbp genome of a Saccharibacteria (TM7) was identified and, despite the microbial complexity of rhizosphere soil, curated to completion. The genome lacks many biosynthetic pathways, including genes required to synthesize DNA de novo. Rather, it requires externally derived nucleotides for DNA and RNA synthesis. Given this, we conclude that rhizosphere-associated Saccharibacteria recycle DNA from bacteria that live off plant exudates and/or phage that acquired 13C because they preyed upon these bacteria and/or directly from the labeled plant DNA. Isotopic labeling indicates that the population was replicating during the 6-week period of plant growth. Interestingly, the genome is ~ 30% larger than other complete Saccharibacteria genomes from non-soil environments, largely due to more genes for complex carbon utilization and amino acid metabolism. Given the ability to degrade cellulose, hemicellulose, pectin, starch, and 1,3-ß-glucan, we predict that this Saccharibacteria generates energy by fermentation of soil necromass and plant root exudates to acetate and lactate. The genome also encodes a linear electron transport chain featuring a terminal oxidase, suggesting that this Saccharibacteria may respire aerobically. The genome encodes a hydrolase that could breakdown salicylic acid, a plant defense signaling molecule, and genes to interconvert a variety of isoprenoids, including the plant hormone zeatin. CONCLUSIONS: Rhizosphere Saccharibacteria likely depend on other bacteria for basic cellular building blocks. We propose that isotopically labeled CO2 is incorporated into plant-derived carbon and then into the DNA of rhizosphere organisms capable of nucleotide synthesis, and the nucleotides are recycled into Saccharibacterial genomes.

Dynamic root exudate chemistry and microbial substrate preferences drive patterns in rhizosphere microbial community assembly.

Like all higher organisms, plants have evolved in the context of a microbial world, shaping both their evolution and their contemporary ecology. Interactions between plant roots and soil microorganisms are critical for plant fitness in natural environments. Given this co-evolution and the pivotal importance of plant-microbial interactions, it has been hypothesized, and a growing body of literature suggests, that plants may regulate the composition of their rhizosphere to promote the growth of microorganisms that improve plant fitness in a given ecosystem. Here, using a combination of comparative genomics and exometabolomics, we show that pre-programmed developmental processes in plants (Avena barbata) result in consistent patterns in the chemical composition of root exudates. This chemical succession in the rhizosphere interacts with microbial metabolite substrate preferences that are predictable from genome sequences. Specifically, we observed a preference by rhizosphere bacteria for consumption of aromatic organic acids exuded by plants (nicotinic, shikimic, salicylic, cinnamic and indole-3-acetic). The combination of these plant exudation traits and microbial substrate uptake traits interact to yield the patterns of microbial community assembly observed in the rhizosphere of an annual grass. This discovery provides a mechanistic underpinning for the process of rhizosphere microbial community assembly and provides an attractive direction for the manipulation of the rhizosphere microbiome for beneficial outcomes.

The interconnected rhizosphere: High network complexity dominates rhizosphere assemblages.

While interactions between roots and microorganisms have been intensively studied, we know little about interactions among root-associated microbes. We used random matrix theory-based network analysis of 16S rRNA genes to identify bacterial networks associated with wild oat (Avena fatua) over two seasons in greenhouse microcosms. Rhizosphere networks were substantially more complex than those in surrounding soils, indicating the rhizosphere has a greater potential for interactions and niche-sharing. Network complexity increased as plants grew, even as diversity decreased, highlighting that community organisation is not captured by univariate diversity. Covariations were predominantly positive (> 80%), suggesting that extensive mutualistic interactions may occur among rhizosphere bacteria; we identified quorum-based signalling as one potential strategy. Putative keystone taxa often had low relative abundances, suggesting low-abundance taxa may significantly contribute to rhizosphere function. Network complexity, a previously undescribed property of the rhizosphere microbiome, appears to be a defining characteristic of this habitat.

Successional Trajectories of Rhizosphere Bacterial Communities over Consecutive Seasons.

UNLABELLED: It is well known that rhizosphere microbiomes differ from those of surrounding soil, and yet we know little about how these root-associated microbial communities change through the growing season and between seasons. We analyzed the response of soil bacteria to roots of the common annual grass Avena fatua over two growing seasons using high-throughput sequencing of 16S rRNA genes. Over the two periods of growth, the rhizosphere bacterial communities followed consistent successional patterns as plants grew, although the starting communities were distinct. Succession in the rhizosphere was characterized by a significant decrease in both taxonomic and phylogenetic diversity relative to background soil communities, driven by reductions in both richness and evenness of the bacterial communities. Plant roots selectively stimulated the relative abundance of Alphaproteobacteria, Betaproteobacteria, and Bacteroidetes but reduced the abundance of Acidobacteria, Actinobacteria, and Firmicutes. Taxa that increased in relative abundance in the rhizosphere soil displayed phylogenetic clustering, suggesting some conservation and an evolutionary basis for the response of complex soil bacterial communities to the presence of plant roots. The reproducibility of rhizosphere succession and the apparent phylogenetic conservation of rhizosphere competence traits suggest adaptation of the indigenous bacterial community to this common grass over the many decades of its presence. IMPORTANCE: We document the successional patterns of rhizosphere bacterial communities associated with a "wild" annual grass, Avena fatua, which is commonly a dominant plant in Mediterranean-type annual grasslands around the world; the plant was grown in its grassland soil. Most studies documenting rhizosphere microbiomes address "domesticated" plants growing in soils to which they are introduced. Rhizosphere bacterial communities exhibited a pattern of temporal succession that was consistent and repeatable over two growing seasons. There are few studies assessing the reproducibility over multiple seasons. Through the growing season, the rhizosphere community became progressively less diverse, likely reflecting root homogenization of soil microniches. Phylogenetic clustering of the rhizosphere dynamic taxa suggests evolutionary adaptation to Avena roots. The reproducibility of rhizosphere succession and the apparent phylogenetic conservation of rhizosphere competence traits suggest adaptation of the indigenous bacterial community to this common grass over the many decades of its presence.

Elevated CO2 shifts the functional structure and metabolic potentials of soil microbial communities in a C4 agroecosystem.

Atmospheric CO2 concentration is continuously increasing, and previous studies have shown that elevated CO2 (eCO2) significantly impacts C3 plants and their soil microbial communities. However, little is known about effects of eCO2 on the compositional and functional structure, and metabolic potential of soil microbial communities under C4 plants. Here we showed that a C4 maize agroecosystem exposed to eCO2 for eight years shifted the functional and phylogenetic structure of soil microbial communities at both soil depths (0-5 cm and 5-15 cm) using EcoPlate and functional gene array (GeoChip 3.0) analyses. The abundances of key genes involved in carbon (C), nitrogen (N) and phosphorus (P) cycling were significantly stimulated under eCO2 at both soil depths, although some differences in carbon utilization patterns were observed between the two soil depths. Consistently, CO2 was found to be the dominant factor explaining 11.9% of the structural variation of functional genes, while depth and the interaction of depth and CO2 explained 5.2% and 3.8%, respectively. This study implies that eCO2 has profound effects on the functional structure and metabolic potential/activity of soil microbial communities associated with C4 plants, possibly leading to changes in ecosystem functioning and feedbacks to global change in C4 agroecosystems.

Effects of selected root exudate components on soil bacterial communities.

Low-molecular-weight organic compounds in root exudates play a key role in plant-microorganism interactions by influencing the structure and function of soil microbial communities. Model exudate solutions, based on organic acids (OAs) (quinic, lactic, maleic acids) and sugars (glucose, sucrose, fructose), previously identified in the rhizosphere of Pinus radiata, were applied to soil microcosms. Root exudate compound solutions stimulated soil dehydrogenase activity and the addition of OAs increased soil pH. The structure of active bacterial communities, based on reverse-transcribed 16S rRNA gene PCR, was assessed by denaturing gradient gel electrophoresis and PhyloChip microarrays. Bacterial taxon richness was greater in all treatments than that in control soil, with a wide range of taxa (88-1043) responding positively to exudate solutions and fewer (<24) responding negatively. OAs caused significantly greater increases than sugars in the detectable richness of the soil bacterial community and larger shifts of dominant taxa. The greater response of bacteria to OAs may be due to the higher amounts of added carbon, solubilization of soil organic matter or shifts in soil pH. Our results indicate that OAs play a significant role in shaping soil bacterial communities and this may therefore have a significant impact on plant growth.

Changes to the structure of Sphingomonas spp. communities associated with biodegradation of the herbicide isoproturon in soil.

The phenyl-urea herbicide isoproturon is a major contaminant of surface and ground-water in agricultural catchments. Earlier work suggested that within-field spatial variation of isoproturon degradation rate resulted from interactions between catabolizing Sphingomonas spp. and pH. In the current study, changes to the structure of Sphingomonas communities during isoproturon catabolism were investigated using Sphingomonas-specific 16S rRNA gene primers. Growth-linked catabolism at high-pH (>7.5) sites was associated with the appearance of multiple new denaturing gradient gel electrophoresis (DGGE) bands. At low-pH sites, there was no change in DGGE banding at sites in which there was cometabolism, but at sites in which there was growth-linked catabolism, degradation was associated with the appearance of a new band not present at high pH sites. Sequencing of DGGE bands indicated that a strain related to Sphingomonas mali proliferated at low pH sites, while strain Sphingomonas sp. SRS2, a catabolic strain identified in earlier work, together with several further Sphingomonas spp., proliferated at high-pH sites. The data indicate that degradation was associated with complex changes to the structure of Sphingomonas spp. communities, the precise nature of which was spatially variable.