Metagenomic insights into genetic factors driving bacterial niche differentiation between bulk and rhizosphere soils.

Cellular motility is crucial for effective colonization of the rhizosphere, but it is not yet clear whether bacterial motility is particularly linked to other genetic traits. Here, we applied genome-resolved metagenomics and phylogenomics to investigate the ecological significance of cellular motility for niche differentiation and the links between the genetic makeup of motile bacteria and rhizosphere colonization within a four-decade maize field experiment. Indeed, highly diverse sets of genes encoding cellular motility, including chemotaxis, flagellar assembly and motility proteins, and utilization of polymeric carbon were the important predictors of bacterial niche differentiation between bulk and rhizosphere soils. This is well exemplified by metagenome-assembled genomes encoding high motility capacity (hmc_MAGs). Their collective abundance was, on average, sixfold higher in rhizosphere soil than in bulk soil. All bulk-soil-derived MAGs showed low motility capacities (lmc). The hmc_MAGs were highly enriched in beneficial traits involved in carbohydrate utilization, assimilatory (nasA) and dissimilatory (nirBD) nitrate reduction, inorganic phosphate solubilization (gcd), and organic phosphate mineralization (phoD). Belonging to the families Sphingomonadaceae, Burkholderiaceae and Steroidobacteraceae, the hmc_MAGs showed a ninefold greater enrichment in these traits than proteobacterial lmc_MAGs and a twofold greater enrichment than 264 genomes publicly available for the above three families, thereby substantiating that a specific rhizosphere effect acted on the microbes represented by the hmc_MAGs. The particular link between the genetic capacities for high cellular motility and increased carbohydrate depolymerization as the key determinant for plant-selected rhizosphere colonization was further substantiated by the analysis of public bulk-rhizosphere soil metagenomes retrieved from wheat and cucumber field sites.

Mycorrhiza-mediated recruitment of complete denitrifying Pseudomonas reduces N2O emissions from soil.

BACKGROUND: Arbuscular mycorrhizal fungi (AMF) are key soil organisms and their extensive hyphae create a unique hyphosphere associated with microbes actively involved in N cycling. However, the underlying mechanisms how AMF and hyphae-associated microbes may cooperate to influence N2O emissions from "hot spot" residue patches remain unclear. Here we explored the key microbes in the hyphosphere involved in N2O production and consumption using amplicon and shotgun metagenomic sequencing. Chemotaxis, growth and N2O emissions of isolated N2O-reducing bacteria in response to hyphal exudates were tested using in vitro cultures and inoculation experiments. RESULTS: AMF hyphae reduced denitrification-derived N2O emission (max. 63%) in C- and N-rich residue patches. AMF consistently enhanced the abundance and expression of clade I nosZ gene, and inconsistently increased that of nirS and nirK genes. The reduction of N2O emissions in the hyphosphere was linked to N2O-reducing Pseudomonas specifically enriched by AMF, concurring with the increase in the relative abundance of the key genes involved in bacterial citrate cycle. Phenotypic characterization of the isolated complete denitrifying P. fluorescens strain JL1 (possessing clade I nosZ) indicated that the decline of net N2O emission was a result of upregulated nosZ expression in P. fluorescens following hyphal exudation (e.g. carboxylates). These findings were further validated by re-inoculating sterilized residue patches with P. fluorescens and by an 11-year-long field experiment showing significant positive correlation between hyphal length density with the abundance of clade I nosZ gene. CONCLUSIONS: The cooperation between AMF and the N2O-reducing Pseudomonas residing on hyphae significantly reduce N2O emissions in the microsites. Carboxylates exuded by hyphae act as attractants in recruiting P. fluorescens and also as stimulants triggering nosZ gene expression. Our discovery indicates that reinforcing synergies between AMF and hyphosphere microbiome may provide unexplored opportunities to stimulate N2O consumption in nutrient-enriched microsites, and consequently reduce N2O emissions from soils. This knowledge opens novel avenues to exploit cross-kingdom microbial interactions for sustainable agriculture and for climate change mitigation. Video Abstract.

Arbuscular mycorrhizal fungi contribute to wheat yield in an agroforestry system with different tree ages.

Intercropping achieved through agroforestry is increasingly being recognized as a sustainable form of land use. In agroforestry, the roots of trees and crops are intermingled, and their interactions and the production of exudates alter the soil environment and soil microbial community. Although tree-crop interactions vary depending on the stand age of the trees, how stand age affects beneficial microorganisms, including arbuscular mycorrhizal fungi (AMF), and whether changes in soil microorganisms feed back on crop growth in agroforestry systems are unknown. We therefore conducted a long-term field study to compare changes in the soil microbial and AMF communities in a jujube/wheat agroforestry system containing trees of different stand ages: 3-year-old jujube, 8-year-old jujube, and 13-year-old jujube. Our results showed that by changing soil moisture and available phosphorus content, the stand age of the trees had a significant effect on the soil microbial and AMF communities. Soil moisture altered the composition of soil bacteria, in particular the proportions of Gram-positive and Gram-negative species, and available phosphorus had significant effects on the AMF community. A network analysis showed that older stands of trees reduced both AMF diversity and network complexity. An ordinary least squares regression analysis indicated that AMF diversity, network complexity, and stability contributed to wheat yield. Finally, structural equation modeling showed that changes in edaphic factors induced by tree age brought about significant variation in the soil microbial and AMF communities, in turn, affecting crop growth. Our study highlights the crucial roles of soil microorganisms, in particular AMF, in supporting plant growth in agroforestry systems as well as the need to consider stand age in the establishment of these systems.

High bacterial diversity and siderophore-producing bacteria collectively suppress Fusarium oxysporum in maize/faba bean intercropping.

Beyond interacting with neighboring plants, crop performance is affected by the microbiome that includes pathogens and mutualists. While the importance of plant-plant interactions in explaining overyielding in intercropping is well known, the role of the microbiome, in particular how the presence of microbes from heterospecific crop species inhibit pathogens of the focal plants in affecting yield remains hardly explored. Here we performed both field samplings and pot experiments to investigate the microbial interactions in the maize/faba bean intercropping system, with the focus on the inhibition of Fusarium oxysporum in faba bean plants. Long-term field measurements show that maize/faba bean intercropping increased crop yield, reduced the gene copies of F. oxysporum by 30-84% and increased bacterial richness and Shannon index compared to monocropping. Bacterial networks in intercropping were more stable with more hub nodes than the respective monocultures. Furthermore, the observed changes of whole microbial communities were aligned with differences in the number of siderophore-producing rhizobacteria in maize and pathogen abundances in faba bean. Maize possessed 71% more siderophore-producing rhizobacteria and 33% more synthetases genes abundance of nonribosomal peptides, especially pyochelin, relative to faba bean. This was further evidenced by the increased numbers of siderophore-producing bacteria and decreased gene copies of F. oxysporum in the rhizosphere of intercropped faba bean. Four bacteria (Pseudomonas spp. B004 and B021, Bacillus spp. B005 and B208) from 95 isolates antagonized F. oxysporum f. sp. fabae. In particular, B005, which represented a hub node in the networks, showed particularly high siderophore-producing capabilities. Intercropping increased overall bacterial diversity and network complexity and the abundance of siderophore-producing bacteria, leading to facilitated pathogen suppression and increased resistance of faba bean to F. oxysporum. This study has great agronomic implications as microorganisms might be specifically targeted to optimize intercropping practices in the future.

Nitrogen availability mediates the priming effect of soil organic matter by preferentially altering the straw carbon-assimilating microbial community.

Straw incorporation into soil increases carbon (C) sequestration but can induce priming effects (PE), the enhanced breakdown of soil organic matter. The direction and magnitude of PE and the consequences for the C balance induced by straw addition depend on nitrogen (N) availability and soil management history. Using 13C-labeled maize straw, we conducted a 56-day incubation to determine the dynamics of PE and the underlying microbial mechanisms after straw and/or mineral N addition to three soils with contrasting cultivation and fertilization histories, i) unfertilized soil (Unfertilized), ii) 8 years farmyard manure amended soil (Manured), and iii) abandoned cropland soil (Abandoned). 13C-PLFAs (phospholipid fatty acids) were analyzed to identify microbial groups utilizing straw and to explore their contribution to the PE. Straw addition increased microbial biomass (MBC), activities of enzymes related to the C and N cycles, and changed microbial community composition. SOC decomposition was enhanced by microbes activated by straw addition, leading to a positive cumulative PE ranging from 494 to 789 µg C g-1 soil. The magnitude of positive PE and straw decomposition in the manured soil was higher than that in the unfertilized and abandoned soils due to larger MBC and higher enzyme activities, resulting in a lower net SOC gain. Compared with straw only, the combination of straw addition with N fertilizer did not influence MBC, but increased positive PE (average increase of 18.1%) and straw decomposition (17.1%), further limiting SOC gain. 13C-labeled fungi: bacteria ratios and Gram-positive (G+): negative (G-) bacteria ratios increased with the increasing PE after N fertilization, but soil-derived (un-labeled) PLFAs remained stable. Random forest analysis further showed that straw C-assimilating microbial attributes are important predictors in driving the greater PE after N addition. Our study highlights the importance of straw C-assimilating fungi and G+ bacteria in mediating N-induced PE in arable soils.

Enrichment of nosZ-type denitrifiers by arbuscular mycorrhizal fungi mitigates N2 O emissions from soybean stubbles.

Hotspots of N2 O emissions are generated from legume residues during decomposition. Arbuscular mycorrhizal fungi (AMF) from co-cultivated intercropped plants may proliferate into the microsites and interact with soil microbes to reduce N2 O emissions. Yet, the mechanisms by which or how mycorrhizal hyphae affect nitrifiers and denitrifiers in the legume residues remain ambiguous. Here, a split-microcosm experiment was conducted to assess hyphae of Rhizophagus aggregatus from neighbouring maize on overall N2 O emissions from stubbles of nodulated or non-nodulated soybean. Soil microbes from fields intercropped with maize/soybean amended with fertilizer nitrogen (SS-N1) or unamended (SS-N0) were added to the soybean chamber only. AMF hyphae consistently reduced N2 O emissions by 20.8%-61.5%. Generally, AMF hyphae promoted the abundance of N2 O-consuming (nosZ-type) denitrifiers and altered their community composition. The effects were partly associated with increasing MBC and DOC. By contrast, AMF reduced the abundance of nirK-type denitrifiers in the nodulated SS-N0 treatment only and that of AOB in the non-nodulated SS-N1 treatment. Taken together, our results show that AMF reduced N2 O emissions from soybean stubbles, mainly through the promotion of N2 O-consuming denitrifiers. This holds promise for mitigating N2 O emissions by manipulating the efficacious AMF and their associated microbes in cereal/legume intercropping systems.

Rhizoplane Bacteria and Plant Species Co-determine Phosphorus-Mediated Microbial Legacy Effect.

Much effort has been directed toward increasing the availability of soil residual phosphorus (P). However, little information is available for the P fertilization-induced biotic P legacy and its mediation of plant P uptake. We collected microbial inocula from a monoculture maize field site with a 10-year P-fertilization history. A greenhouse experiment was conducted to investigate whether bacterial communities, as a result of different P-fertilization history (nil P, 33 and/or 131 kg P kg ha-1 yr-1), affected the growth of a conspecific (maize) or heterospecific (clover) plant, at two levels of current P application (5 and 30 mg P kg-1 soil; P5 and P30). Deep amplicon sequencing (16S rRNA) was used to determine the maize and clover root-associated bacterial microbiome in different rhizocompartments (rhizoplane, rhizosphere, bulk soil). For both maize and clover, rhizocompartment and host identity were the dominant factors shaping bacterial assemblages, followed by P supply level and the inoculum effect was smallest. Bacterial operational taxonomic unit (OTU) numbers decreased from bulk soil to rhizoplane, whilst specific OTUs were enriched from bulk soil to rhizoplane. A clear hierarchical habitat filtering of bacterial communities was observed in the rhizoplane of the two plant species. The functional prediction of dominant bacterial taxa in the rhizoplane differed between clover and maize, and clover microbiota were more closely associated with P metabolism and maize with carbon cycling. More connected and complex interactions were observed in the clover rhizoplane compared to maize. The microbial legacy effect caused by long-term P fertilization is overridden by host identity and rhizocompartment. Our results highlight the importance of crop diversification in improving P efficiency. The fine-tuning of rhizosphere microbiome in host metabolism indicates that the functions of microbial communities should be integrated into P management to increase P use efficiency and sustainable food production.

Enhancement of faba bean competitive ability by arbuscular mycorrhizal fungi is highly correlated with dynamic nutrient acquisition by competing wheat.

The mechanistic understanding of the dynamic processes linking nutrient acquisition and biomass production of competing individuals can be instructive in optimizing intercropping systems. Here, we examine the effect of inoculation with Funneliformis mosseae on competitive dynamics between wheat and faba bean. Wheat is less responsive to mycorrhizal inoculation. Both inoculated and uninoculated wheat attained the maximum instantaneous N and P capture approximately five days before it attained the maximum instantaneous biomass production, indicating that wheat detected the competitor and responded physiologically to resource limitation prior to the biomass response. By contrast, the instantaneous N and P capture by uninoculated faba bean remained low throughout the growth period, and plant growth was not significantly affected by competing wheat. However, inoculation substantially enhanced biomass production and N and P acquisition of faba bean. The exudation of citrate and malate acids and acid phosphatase activity were greater in mycorrhizal than in uninoculated faba bean, and rhizosphere pH tended to decrease. We conclude that under N and P limiting conditions, temporal separation of N and P acquisition by competing plant species and enhancement of complementary resource use in the presence of AMF might be attributable to the competitive co-existence of faba bean and wheat.