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Soils are a main repository of biodiversity harbouring immense diversity of microbial species that plays a central role in fundamental ecological processes and acts as the seed bank for emergence of the plant microbiome in cropland ecosystems. Crop-associated microbiomes play an important role in shaping plant performance, which includes but not limited to nutrient uptake, disease resistance, and abiotic stress tolerance. Although our understanding of structure and function of soil and plant microbiomes has been rapidly advancing, most of our knowledge comes from ecosystems in natural environment. In this review, we present an overview of the current knowledge of diversity and function of microbial communities along the soil-plant continuum in agroecosystems. To characterize the ecological mechanisms for community assembly of soil and crop microbiomes, we explore how crop host and environmental factors such as plant species and developmental stage, pathogen invasion, and land management shape microbiome structure, microbial co-occurrence patterns, and crop-microbiome interactions. Particularly, the relative importance of deterministic and stochastic processes in microbial community assembly is illustrated under different environmental conditions, and potential sources and keystone taxa of the crop microbiome are described. Finally, we highlight a few important questions and perspectives in future crop microbiome research.

Syntrophic propionate oxidation is one of the rate-limiting steps during anaerobic decomposition of organic matter in anoxic environments. Syntrophic propionate-oxidizing bacteria (SPOB) are members of the “rare biosphere” living at the edge of the thermodynamic limit in most natural habitats.

Belowground biodiversity supports multiple ecosystem functions and services that humans rely on. However, there is a dearth of studies exploring the determinants of the biodiversity-ecosystem function (BEF) relationships, particularly in intensely managed agricultural ecosystems. Here, we reported significant and positive relationships between soil biodiversity of multiple organism groups and multiple ecosystem functions in 228 agricultural fields, relating to crop yield, nutrient provisioning, element cycling, and pathogen control. The relationships were influenced by the types of organisms that soil phylotypes with larger sizes or at higher trophic levels, for example, invertebrates or protist predators, appeared to exhibit weaker or no BEF relationships when compared to those with smaller sizes or at lower trophic levels, for example, archaea, bacteria, fungi, and protist phototrophs. Particularly, we highlighted the role of soil network complexity, reflected by co-occurrence patterns among multitrophic-level organisms, in enhancing the link between soil biodiversity and ecosystem functions. Our results represent a significant advance in forecasting the impacts of belowground multitrophic organisms on ecosystem functions in agricultural systems, and suggest that soil multitrophic network complexity should be considered a key factor in enhancing ecosystem productivity and sustainability under land-use intensification.

Electromethanogenesis refers to the process whereby methanogens utilize current for the reduction of CO(2) to CH(4). Setting low cathode potentials is essential for this process. In this study, we tested if magnetite, an iron oxide mineral widespread in the environment, can facilitate the adaptation of methanogen communities to the elevation of cathode potentials in electrochemical reactors. Two-chamber electrochemical reactors were constructed with inoculants obtained from paddy field soil. We elevated cathode potentials stepwise from the initial −0.6 V versus the standard hydrogen electrode (SHE) to −0.5 V and then to −0.4 V over the 130 days of acclimation. Only weak current consumption and CH(4) production were observed in the bioreactors without magnetite. However, significant current consumption and CH(4) production were recorded in the magnetite bioreactors. The robustness of electroactivity of the magnetite bioreactors was not affected by the elevation of cathode potentials from −0.6 V to −0.4 V. However, the current consumption and CH(4) production were halted in the bioreactors without magnetite when the cathode potentials were elevated to −0.4 V. Methanogens related to Methanospirillum were enriched on the cathode surfaces of magnetite bioreactors at −0.4 V, while Methanosarcina relatively dominated in the bioreactors without magnetite. Methanobacterium also increased in the magnetite bioreactors but stayed off electrodes at −0.4 V. Apparently, the magnetite greatly facilitates the development of biocathodes, and it appears that with the aid of magnetite, Methanospirillum spp. can adapt to the high cathode potentials, performing efficient electromethanogenesis. IMPORTANCE Converting CO(2) to CH(4) through bioelectrochemistry is a promising approach to the development of green energy biotechnology. This process, however, requires low cathode potentials, which entails a cost. In this study, we tested if magnetite, a conductive iron mineral, can facilitate the adaptation of methanogens to the elevation of cathode potentials. In two-chamber reactors constructed by using inoculants obtained from paddy field soil, biocathodes developed robustly in the presence of magnetite, whereas only weak activities in CH(4) production and current consumption were observed in the bioreactors without magnetite. The elevation of cathode potentials did not affect the robustness of electroactivity of the magnetite bioreactors over the 130 days of acclimation. Methanospirillum strains were identified as the key methanogens associated with the cathode surfaces during the operation at high potentials. The findings reported in this study shed new light on the adaptation of methanogen communities to the elevated cathode potentials in the presence of magnetite.

Soil biogeochemical processes are not only gauged by the dominant taxa in the microbiome but also depend on the critical functions of its 'rare biosphere' members. Here, we evaluated the biogeographical pattern of 'rare biosphere' propionate-oxidizing syntrophs in 113 paddy soil samples collected across China. The relative abundance, activity and growth potential of propionate-oxidizing syntrophs were analysed to provide a panoramic view of syntroph biogeographical distribution at the continental scale. The relative abundances of four syntroph genera, Syntrophobacter, Pelotomaculum, Smithella and Syntrophomonas were significantly greater at the warm low latitudes than at the cool high latitudes. Correspondingly, propionate degradation was faster in the low latitude soils compared with the high latitude soils. The low rate of propionate degradation in the high latitude soils resulted in a greater increase of the total syntroph relative abundance, probably due to their initial low relative abundances and the longer incubation time for propionate consumption. The mean annual temperature (MAT) is the most important factor shaping the biogeographical pattern of propionate-oxidizing syntrophs, with the next factor being the soil's total sulfur content (TS). We suggest that the effect of MAT is related to the thermodynamic conditions, in which the endergonic constraint of propionate oxidation is leveraged with the increase of MAT. The TS effect is likely due to the ability of some propionate syntrophs to facultatively perform sulfate respiration.

Application of ferric iron is conventionally considered to inhibit methanogenesis in anoxic environments. Here we show that Methanosarcina mazei zm-15, a strain isolated from the natural wetland of Tibetan plateau, is capable of Fe(III) reduction, which significantly promotes its growth and methanogenesis. We grew Ms. mazei zm-15 in a medium containing acetate supplemented with Fe(III) in ferric citrate or ferrihydrite and to some cultures anthraquinone-2,6-disulfonate (AQDS) was applied as an electron shuttle. The reduction of Fe(III) species occurred immediately. Ferric citrate was more readily reduced than ferrihydrite. The X-ray diffraction spectra analysis showed the formation of magnetite from ferrihydrite and amorphous reduced products from ferrihydrite plus AQDS. The analysis of protein contents revealed that Fe(III) reduction contributed 36%-46% of the cell growth. The growth yield, estimated as protein increment per acetate consumed for Fe(III) reduction, increased by 20- to 30-fold compared with methanogenesis, which is in consistence with the difference in free energy available by Fe(III) reduction relative to methanogenesis. We propose that the outer-surface multiheme c-type cytochrome predicted from Ms. mazei zm-15 genome serves as the terminal reductase with the energy-converting hydrogenase and F

Syntrophy is a thermodynamically required mutualistic cooperation between fatty acid-oxidizing bacteria and methanogens that plays the important role in organic decomposition and methanogenesis in anoxic environments. In this study, three experiments were conducted to evaluate the cell-to-cell interaction in a thermophilic coculture consisting of Syntrophothermus lipocalidus and Methanocella conradii and a mesophilic coculture consisting of Syntrophomonas wolfei and Methanococcus maripaludis. First, syntrophs and methanogens were inoculated at different initial cell ratios to evaluate the growth synchronization. The quantitative PCR analysis revealed that the organism with a lower relative abundance at the beginning always grew faster, and the cell ratio converged over time to relative constant values in both the thermophilic and mesophilic cocultures. Next, intermittent ultrasound and constant shaking treatments were used to evaluate the influence of physical disturbance on microbial aggregation in the mesophilic coculture. The fluorescence in situ hybridization and scanning electron microscopy revealed that the tendency of syntrophic aggregation was not affected by the physical disturbances, although the activity was slightly depressed. Syntrophomonas dominated in the initial microbial aggregates, which, however, did not grow until Methanococcus was attached and increased to a significant extent, indicating the local growth synchronization during the formation and maturation of syntrophic aggregates. Last, microfluidic experiments revealed that whether or not Syntrophomonas or Methanococcus was loaded first, the second organism preferred moving to the place where the first organism was located, suggesting the cell-to-cell attraction between Syntrophomonas and Methanococcus. Collectively, our study demonstrated the growth synchronization and cell-to-cell attraction between the butyrate-oxidizing bacteria and methanogens for optimizing the syntrophic cooperation.

Elucidating the mechanisms underpinning the responses of abundant and rare microbial taxa to environmental disturbances is essential for understanding the biodiversity-stability relationship and maintaining microbial diversity. Here, we explored the response patterns of abundant and rare bacterial taxa to disturbances by invasive plant growth and oil contamination in agricultural soils across a large spatial scale (latitude gradient = 18.62°–46.51°). Our meta-analysis based on existing Illumina sequencing datasets showed that abundant taxa persisted under the disturbances whereas rare taxa were more easily affected, indicating the higher resilience or resistance of abundant taxa to disturbances. The responses of abundant taxa were associated with mean annual temperature at the sampling sites, while rare taxa instead showed stochastic responses. There were significantly negative linear regressions between bacterial α-diversity and community dissimilarities (disturbed vs. undisturbed soils), suggesting stronger resilience or resistance in those bacterial communities with higher α-diversity. This resilience or resistance was mainly associated with the α-diversity of abundant taxa. Our network analysis showed that the disturbances substantially decreased the strength of the connections, loosened the co-occurrence relationships, and reshaped the complex bacterial interactions. In the undisturbed soils, abundant taxa were located in central positions within the network more often than were rare taxa, while these trends were reversed in the disturbed soils. Our results suggest that abundant taxa play a dominant role in the stability and maintenance of agro-soil bacterial communities, while rare taxa could greatly influence local bacterial interactions under environmental disturbances.

The factors determining stochastic and deterministic processes that drive microbial community structure, specifically the balance of abundant and rare bacterial taxa, remain underexplored. Here we examined biogeographic patterns of abundant and rare bacterial taxa and explored environmental factors influencing their community assembly processes in agricultural fields across eastern China. More phylogenetic turnover correlating with spatial distance was observed in abundant than rare sub-communities. Homogeneous selection was the main assembly process for both the abundant and rare sub-communities; however, the abundant sub-community was more tightly clustered phylogenetically and was more sensitive to dispersal limitations than the rare sub-community. Rare sub-community of rice fields and abundant sub-community of maize fields were more governed by stochastic assembly processes, which showed higher operational taxonomic unit richness. We propose a conceptual paradigm wherein soil pH and mean annual temperature mediate the assembly of the abundant and rare sub-communities respectively. A higher soil pH leads to deterministic assembly of the abundant sub-community. For the rare sub-community, the dominance of stochasticity in low-temperature regions indicates weaker niche-based exclusion and the arrival of more evolutionary lineages. These findings suggest that the community assembly processes for abundant and rare bacterial taxa are dependent on distinct environmental variables in agro-ecosystems.

It has been suggested that a few methanogens are capable of extracellular electron transfers. For instance, Methanosarcina barkeri can directly capture electrons from the coexisting microbial cells of other species. Methanothrix harundinacea and Methanosarcina horonobensis retrieve electrons from Geobacter metallireducens via direct interspecies electron transfer (DIET). Recently, Methanobacterium, designated strain YSL, has been found to grow via DIET in the co-culture with Geobacter metallireducens. Methanosarcina acetivorans can perform anaerobic methane oxidation and respiratory growth relying on Fe(III) reduction through the extracellular electron transfer. Methanosarcina mazei is capable of electromethanogenesis under the conditions where electron-transfer mediators like H2 or formate are limited. The membrane-bound multiheme c-type cytochromes (MHC) and electrically-conductive cellular appendages have been assumed to mediate the extracellular electron transfer in bacteria like Geobacter and Shewanella species. These molecules or structures are rare but have been recently identified in a few methanogens. Here, we review the current state of knowledge for the putative extracellular electron transfers in methanogens and highlight the opportunities and challenges for future research.

Additional file 1: Figure S1. Sampling and sequencing of methanogens. Abscissa: the number of sequences randomly selected from the sample; ordinate: the species diversity and the number of OTU that can be represented by the random sampling sequence. Each curve in the figure represents a sample, marked with a different color. Figure S2. Linear regression of methanogenic archaea α-diversity and MAT (A) and CH4 emissions (B). Solid lines denote significant relationship with 95% confidence interval (shadow area). Dotted line denotes P > 0.05. Figure S3. The co-occurrence network structure of methanogenic community of 39 plots. Percentage in the top right corner indicates the proportion of 33 key OTUs (Table S4) in total node numbers of each network. Figure S4. Contributions of climatic variables and soil properties to the network topological attributes based on correlation and random forest regression model. Circle size represents the variable importance (that is, proportion of explained variability calculated via forest regression analysis analysis). Colors represent Spearman’s correlation coefficients (node: node number; edge: edge number; modularity: modularity; positive: positive correlations; negative: negative correlations; Avg.C.C: average clustering coefficient; N.D: network diameter; C.P.L: characteristic path length). Figure S5. Spearman’s correlation analysis of network topological attributes and richness. Blue shaded area is a 95% confidence interval. Figure S6. Spearman’s correlation analysis of network topological attributes and CH4 emissions. Dotted lines denote P > 0.05. Solid lines indicate a significant correlation (P < 0.05), with the blue shaded area showing the 95% confidence interval. Figure S7. Relative abundance of OTUs of the five groups of methanogenic archaeal communities. Each column followed by different letters differed significantly at P < 0.05 (ANOVA, Duncan test). (A.endemic: always endemic group; C.endemic: conditionally endemic group; C.common: conditionally common group; A.common: always common group). Figure S8. Relative abundance frequency histogram of 33 keystone OTUs in 429 samples. The blue histogram indicates rare OTUs (average relative abundance < 1% in 429 samples). The red histogram indicates abundant OTUs (average relative abundance > 1% in 429 samples), and the inset represents low-frequency OTUs with average relative abundance < 1%. Figure S9. The relative abundance of keystone taxa (at the family level) changes with temperature. (HL: 1.5°C, CC: 4.5°C, SY:8.3°C, YY: 14.4°C, FQ:13.9°C, LA: 16.1°C, QZ:17.9°C, ZX:17°C, JO:18.8°C, CT:19°C, HY:18°C, QX:21.6°C, HK:23.8°C). Figure S10. The relationship between keystone species and microbial functional genes related to methane generation depicted as colored segments in a CIRCOS plot. Ribbons connecting two segments indicate the interaction between the two. The size of the ribbon is proportional to the number of links. Table S1. Geographic information of 13 sampling regions with 3 plots in each region. Table S2. The topological features of the co-occurrence network in each plot. Table S3. Partial mantel test between network topology attributes and variables. Table S4. Changes of network topology attributes with latitude. Table S5. Information of 33 keystone OTUs involved in common co-occurrence relationship.

Background Soil methanogens participate in complex interactions, which determine the community structures and functions. Studies continue to seek the coexistence patterns of soil methanogens, influencing factors and the contribution to methane (CH4) production, which are regulated primarily by species interactions, and the functional significance of these interactions. Here, methane emissions were measured in rice paddies across the Asian continent, and the complex interactions involved in coexistence patterns of methanogenic archaeal communities were represented as pairwise links in co-occurrence networks. Results The network topological properties, which were positively correlated with mean annual temperature, were the most important predictor of CH4 emissions among all the biotic and abiotic factors. The methanogenic groups involved in commonly co-occurring links among the 39 local networks contributed most to CH4 emission (53.3%), much higher than the contribution of methanogenic groups with endemic links (36.8%). The potential keystone taxa, belonging to Methanobacterium, Methanocella, Methanothrix, and Methanosarcina, possessed high linkages with the methane generation functional genes mcrA, fwdB, mtbA, and mtbC. Moreover, the commonly coexisting taxa showed a very different assembly pattern, with ~ 30% determinism and ~ 70% stochasticity. In contrast, a higher proportion of stochasticity (93~99%) characterized the assembly of endemically coexisting taxa. Conclusions These results suggest that the coexistence patterns of microbes are closely tied to their functional significance, and the potential importance of common coexistence further imply that complex networks of interactions may contribute more than species diversity to soil functions.

Electromethanogenesis refers to the process where methanogens utilize electrons derived from cathodes for the reduction of CO2 to CH4. Setting of low cathode potentials is essential for this process. In this study, we test if magnetite, an iron oxide mineral widespread in environment, can facilitate the adaption of methanogen community to the elevation of cathode potentials in electrochemical reactors. Two-chamber electrochemical reactors were constructed with inoculants obtained from a paddy field soil. We elevated cathode potentials stepwise from the initial −0.6 V vs standard hydrogen electrode (SHE) to −0.5 V and then to −0.4 V over the 120 days acclimation. Only weak current consumption and CH4 production were observed in the reactors without magnetite. But biocathodes were firmly developed and significant current consumption and CH4 production were recorded in the magnetite reactors. The robustness of electro-activity in the magnetite reactors was not affected with the elevation of cathode potentials from −0.6 V to −0.4 V. But, the current consumption and CH4 production were virtually halted in the reactors without magnetite when cathode potential was elevated to −0.4 V. Methanogens related to Methanospirillum were enriched on cathode surface of the magnetite reactors at −0.4 V, while Methanosarcina relatively dominated in the reactors without magnetite. Methanobacterium also increased in the magnetite reactors but stayed off electrodes in the culture medium at −0.4 V. Apparently, magnetite greatly facilitates the development of biocathodes, and it appears that with the aid of magnetite Methanospirillum spp. can adapt to high cathode potentials performing the efficient electromethanogenesis.IMPORTANCEConverting CO2 to CH4 through bioelectrochemistry is a promising approach for development of green energy biotechnology. This process however requires setting the low cathode potentials, which takes cost. In this study, we test if magnetite, a conductive iron mineral, can facilitate the adaption of methanogens to the elevation of cathode potentials. In the two-chamber reactors constructed using inoculants obtained from a paddy field soil, biocathodes were firmly developed in the presence of magnetite, whereas only weak electro-activity was observed in the reactors without magnetite. The elevation of cathode potentials did not affect the robustness of electro-activity in the magnetite reactors over the 120 days acclimation. Methanospirillum was identified as the key methanogens associated with cathode surface during the operation at relatively high potentials. The findings reported in this study shed a new light on the adaption of methanogen community to the elevated cathode potentials in the presence of magnetite.

Background: Belowground biodiversity supports multiple ecosystem functions and services that humans rely on. However, there is a dearth of studies conducted on a large spatial scale on the topic in intensely managed agricultural ecosystems. Existing studies have overlooked the fact that the functional diversity in other trophic groups within a food web could influence function of an individual in another trophic group. Here, we report significant and positive relationships between soil biodiversity (archaea, bacteria, fungi, protists, and invertebrates) and multiple ecosystem functions (nutrient provisioning, element cycling, and reduced pathogenicity potential) in 228 agricultural fields. Results: The relationships were influenced by (I) the types of organisms with significant relationships in archaea, bacteria, and fungi and not in protists and invertebrates, and (II) the connectedness of dominant phylotypes across soil food webs, which generate different ecological clusters within soil networks to maintain multiple functions. In addition, we highlight the role of soil food web complexity, reflected by ecological networks comprising diverse organisms, which promote the multiple functions and enhance the link between soil biodiversity and ecosystem functions. Conclusions: Overall, our results represent a significant advance in forecasting the impacts of belowground biodiversity within food webs on ecosystem functions in agricultural systems, and suggest that soil biodiversity, particularly soil food web complexity, should not be overlooked, but rather considered a key factor and integrated into policy and management activities aimed at enhancing and maintaining ecosystem productivity, stability, and sustainability under land-use intensification.

Background Global biogeochemical processes are not only gauged by dominant taxa of soil microbiome but also depend on the critical functions of “rare biosphere” members. Here we evaluated the biogeographical pattern of “rare biosphere” propionate-oxidizing syntrophs in 113 paddy soil samples collected across eastern China. Results The relative abundance, functioning capacity and growth potential of propionate-oxidizing syntrophs were analyzed to provide a panoramic view of syntroph biogeographical distribution at the continental scale. The relative abundances of four syntroph genera, Syntrophobacter, Pelotomaculum, Smithella and Syntrophomonas were significantly greater at the warm low latitudes than at the cool high latitudes. Correspondingly, the functioning potential of propionate degradation was greater in the low latitude soils compared with the high latitude soils. The slow rate of propionate degradation in high latitude soils resulted in a greater fold change in increase of the relative abundance, probably due to the growth rate-yield tradeoff relationship. The mean annual temperature (MAT) is the most important factor shaping the biogeographical pattern of propionate-oxidizing syntrophs, with the next factor to be the total S content (TS) in soil. Conclusions We suggest that the effect of MAT is related to the Gibbs free energy change, in which the endergonic tension of propionate oxidation is leveraged with the increase of MAT. The TS effect is likely due to that some propionate syntrophs can facultatively perform sulfate respiration.

Syntrophic propionate oxidation (SPO) coupled with methanogenesis is often inhibited under high ammonium concentrations in anaerobic digesters. However, the inhibitory mechanism remains poorly understood. We conducted two independent laboratory experiments with a swine manure digester sludge. In experiment I, RNA-based stable isotope probing (SIP) was applied to determine the active players of both bacteria and methanogens involved in SPO under different ammonium concentrations (0, 3 and 7 g NH4+ N L−1). In experiment II, the dynamics of the bacterial community under ammonia stress was monitored using the 16S rRNA pyrosequencing and quantitative PCR under similar conditions as in experiment I but without the addition of external propionate. An additional higher ammonium treatment (10 g NH4+ N L−1) was applied in experiment II to maximize the ammonia stress. We identified that the Smithella bacteria and the Methanosaetaceae and Methanospirillaceae archaea were the most active players involved in SPO and methanogenesis. We revealed that Smithella, Methanosaetaceae and Methanospirillaceae were moderately and severely inhibited at 3 and 7–10 g NH4+ N L−1, respectively. However, the fermentative bacteria appeared to be more tolerant to ammonia stress. The microbial responses were corroborated with the accumulation of VFAs and the repression of methanogenesis under high ammonium conditions.

It remains elusive if direct interspecies electron transfer (DIET) occurs in canonical syntrophy involving short-chain fatty acids oxidation. In the present study, we determined the effects of carbon nanomaterials on syntrophic oxidation of butyrate in two lake sediment enrichments and a defined coculture comprising Syntrophomonas wolfei and Methanococcus Maripaludis. After four continuous transfers of enrichment cultivation, Syntrophomonas dominated the bacterial populations in enrichments, and the dominated methanogens comprised Methanosarcina and Methanospirillum in one enrichment (from Weiming Lake) and Methanoregula and Methanospirillum in another (from Erhai Lake). Butyrate oxidation and CH4 production was significantly accelerated by carbon nanotubes (CNTs) in both enrichments. Replacement of CNTs by magnetite caused similar stimulating effect. For the defined coculture, two carbon nanomaterials, CNTs and reduced graphene oxide (rGO), were tested, both showed consistently stimulating effects on butyrate oxidation. Addition of kaolinite, an electric nonconductive clay mineral, however, revealed no effect. The test on M. maripaludis in pure culture showed no effect by rGO and a negative effect by CNTs (especially at a high concentration). Fluorescence in situ hybridization (FISH) and scanning electron microscopy (SEM) revealed that microbial cells were interwoven by CNTs forming cell-CNT mixture aggregates, and in case of rGO, cells were attached to surface or wrapped-up by rGO thin sheets. Collectively, our data suggest that the presence of conductive nanomaterials likely induces DIET in syntrophic butyrate oxidation.

Although specific soil properties controlling the arsenic (As) aging process have been studied extensively, few investigations have attempted to determine how soil types influence As bioavailability and fractionations in soils. Nine types of soil were selected from typical grain producing areas in China, and the bioavailability and fractionations of As during aging were measured. Results showed that available As in all soils rapidly decreased in the first 30 days and slowly declined thereafter. In spiked soils, As easily became less available and less toxic in low pH soils compared to high pH soils, demonstrating the importance of soil pH on As availability. Results from fitting kinetic equations revealed that the pseudo-second-order model described the As aging processes well in all soils (R2 = 0.945-0.999, P

The Earth surface contains various oxic and anoxic environments. The later include natural wetlands, river and lake sediments, paddy field soils and landfills. In the last few decades, the biogeochemical cycle of carbon in anoxic environments, which leads to the production and emission of methane, a potent greenhouse gas in the atmosphere, has drawn great attentions from both scientific and public sectors. New organisms and mechanisms involved in methanogenesis and carbon cycling have been uncovered. Interspecies electron transfer is considered as a crucial step in methanogenesis in anoxic environments. Electron-carrying mediators, like H2 and formate, are known to play the key role in electron transfer. Recently, it has been found that in addition to the conventional electron transfer via chemical mediators, direct interspecies electron transfer (DIET) can occur. In this Review, we describe the ecology and biogeochemistry of methanogenesis and highlight the effect of microbe-mineral interaction on microbial syntrophy. Recent advances in the study of DIET may pave the way towards a mechanistic understanding of methanogenesis and the influence of microbe-mineral interaction on this process.

About 50 years ago, most of the natural wetlands in northeast China, the Sanjiang plain, were converted to either flooded rice fields or to upland soybean fields. After the conversion, natural wetland soils were either managed as artificial wetland or as drained upland resulting in soil microbial community changes. The purpose of our study was to understand how methanogenic microbial communities and their functions had changed in the two different soils upon conversion, and whether these communities now exhibit different resistance/resilience to drying and rewetting. Therefore, we determined function, abundance and composition of the methanogenic archaeal and bacterial communities in two soils reclaimed from a Carex wetland 25 years ago. We incubated the soils under anoxic conditions and measured the rates and pathways of CH4 production by analyzing concentration and δ13C of CH4 and acetate in the presence and absence of methyl fluoride, an inhibitor of aceticlastic methanogenesis. We also analyzed the abundance of bacterial and archaeal 16S rRNA genes, and of mcrA (coding for a subunit of the methyl coenzyme M reductase) using qPCR. The composition of the archaeal and bacterial 16S rRNA genes was determined by using MiSeq illumina sequencing. Our results showed clear differences in structure and function of methanogenic archaeal communities in rice field soil versus upland soil. Furthermore, in both soils composition of bacteria and archaea changed after artificial drying and became less diverse. The archaeal and bacterial signature species in the two soils were also different. However, functional changes were similar, with rates of CH4 production and contribution of aceticlastic methanogenesis decreasing upon drying and rewetting in both soils.

Soil communities are intricately linked to ecosystem functioning, and a predictive understanding of how communities assemble in response to environmental change is of great ecological importance. Little is known about the assembly processes governing abundant and rare fungal communities across agro-ecosystems, particularly with regard to their environmental adaptation. By considering abundant and rare taxa, we tested the environmental thresholds and phylogenetic signals for ecological preferences of fungal communities across complex environmental gradients to reflect their environmental adaptation, and explored the factors influencing their assembly based on the large-scale soil survey in agricultural fields across eastern China. We found that the abundant taxa exhibited remarkably broader response thresholds and stronger phylogenetic signals for the ecological preferences across environmental gradients compared to the rare taxa. Neutral processes played a key role in shaping the abundant subcommunity compared to the rare subcommunity. Null model analysis revealed that the abundant subcommunity was less clustered phylogenetically and governed primarily by dispersal limitation, while homogeneous selection was the major assembly process in the rare subcommunity. Soil available sulfur was the major factor mediating the balance between stochastic and deterministic processes of both the abundant and rare subcommunities, as indicated by an increase in stochasticity with higher available sulfur concentration. Based on macroecological spatial scale datasets, our study revealed the potential broader environmental adaptation of abundant fungal taxa compared to rare fungal taxa, and identified the factors mediating their distinct community assembly processes in agricultural fields. These results contribute to our understanding of the mechanisms underlying the generation and maintenance of fungal diversity in response to global environmental change.

It has been recently shown that magnetite nanoparticles (nanoFe3 O4 ) can facilitate methanogenic syntrophy but the effect of magnetite on methanogenesis alone remains elusive. Here we show that aceticlastic methanogenesis by Methanosarcina mazei is accelerated by magnetite and is correlated with the redox cycling of structural Fe(II) and Fe(III) in the mineral. An enrichment and its closest pure culture relative, Ms. mazei zm-15, both obtained from a natural wetland of the Tibetan plateau were tested in this experiment. The Fe(II) to Fe(III) ratios in magnetite, as measured by multiple approaches, show an initial increase in both the methanogenic cultures and the blank preparations containing no microbes. The Fe(II)/Fe(III) ratio then displays a distinct decline followed by an increase towards the end of incubation only in the enrichment and pure culture cultivations. This redox cycling of magnetite is in accordance with the stimulation of aceticlastic methanogenesis. Microscopic observation reveals the precipitation of nanoFe3 O4 on methanogen cell surface. The genomic analysis predicts that in addition to electron transfer components essential for aceticlastic methanogenesis, Ms. mazei zm-15 contains an outer-surface multiheme c-type cytochrome (MHC) and a few function-unknown surface proteins that harbour monoheme motif. We hypothesize that the redox cycling of nanoFe3 O4 delivers a positive influence via the MHC to the membrane electron transfer chain and hence promote the aceticlastic methanogenesis.

Metatranscriptomics is a powerful tool for capturing gene expression patterns in microbial communities and investigating their responses to environmental conditions. Stable isotope probing (SIP) is a method to target specific functional groups of microorganisms in environmental samples. The combination of RNA-SIP with metatranscriptomic analysis enhances the detection and identification of mRNA from target microorganisms. In this chapter we provide a protocol for RNA-SIP, mRNA enrichment, and mRNA preparation for high-throughput sequencing using an example of targeting methanotrophs in rice field soil.

Magnetite nanoparticles (nanoFe3O4) have been reported to facilitate direct interspecies electron transfer (DIET) between syntrophic bacteria and methanogens thereby improving syntrophic methanogenesis. However, whether or how nanoFe3O4 affects acetotrophic methanogenesis remain unknown. Herein, we demonstrate the unique role of nanoFe3O4 in accelerating methane production from direct acetotrophic methanogenesis in Methanosarcina-enriched cultrures, which was further confirmed by pure cultures of Methanosarcina barkeri. Compared with other nanomaterials of higher electrical conductivity such as carbon nanotubes (CNTs) and graphite, nanoFe3O4 with mixed valence Fe(II) and Fe(III) had the most significant stimulatory effect on methane production, suggesting its redox activity rather than electrical conductivity led to enhanced methanogenesis by M. barkeri. Cell morphology and spectroscopy analysis revealed that nanoFe3O4 penetrated into the cell membrane and cytoplasm of M. barkeri. These results provide the unprecedented possibility that nanoFe3O4 in the cell membrane of methanogens serve as electron shuttles to facilitate intracellular electron transfer and thus enhance methane production. This work has important implications not only for understanding the mechanisms of mineral-methanogen interaction but also for optimizing engineered methanogenic processes.

Disentangling the roles of the core microbiota in community maintaining and soil nutrient cycling is an important yet poorly understood topic in microbial ecology. This study presents an exploratory effort to gain predictive understanding of the spatial atlas and ecological roles of the core microbiota. A systematic, continental-scale survey was conducted using agro-soils in adjacent pairs of maize (dryland) and rice (wetland) fields across eastern China. The results indicate that the core microbiota play major ecological roles in maintaining complex connections between bacterial taxa and are associated with belowground multinutrient cycling. A continental atlas was built for mapping the bacterial spatial distributions in agro-soils through identifying their habitat preferences. This study represents a significant advance in forecasting the responses of agricultural ecosystems to anthropogenic disturbance and thus helps manage soil bacterial communities for better provisioning of key ecosystem services—the ultimate goal of microbial ecology., Revealing the ecological roles of the core microbiota in community maintaining and soil nutrient cycling is crucial for understanding ecosystem function, yet there is a dearth of continental-scale studies on this fundamental topic in microbial ecology. Here, we collected 251 soil samples from adjacent pairs of maize and rice fields at a continental scale in eastern China. We revealed the major ecological roles of the core microbiota in maintaining complex connections between bacterial taxa and their associations with belowground multinutrient cycling. By identifying the habitat preferences of the core microbiota, we built a continental atlas for mapping the spatial distributions of bacteria in agro-soils, which helps forecast the responses of agricultural ecosystems to anthropogenic disturbance. The multinutrient cycling index for maize and rice soils was related to bacterial α-diversity and β-diversity, respectively. Rice soils exhibited higher bacterial diversity and closer bacterial cooccurrence relationships than maize soils. In contrast to the macro- or microecological latitudinal richness patterns in natural terrestrial ecosystems, the bacteria in maize soils showed higher richness at high latitudes; however, this trend was not observed in rice soils. This study provides a new perspective on the distinct bacterial biogeographic patterns to predict the ecological roles of the core microbiota in agro-soils and thus helps manage soil bacterial communities for better provisioning of key ecosystem services. IMPORTANCE Disentangling the roles of the core microbiota in community maintaining and soil nutrient cycling is an important yet poorly understood topic in microbial ecology. This study presents an exploratory effort to gain predictive understanding of the spatial atlas and ecological roles of the core microbiota. A systematic, continental-scale survey was conducted using agro-soils in adjacent pairs of maize (dryland) and rice (wetland) fields across eastern China. The results indicate that the core microbiota play major ecological roles in maintaining complex connections between bacterial taxa and are associated with belowground multinutrient cycling. A continental atlas was built for mapping the bacterial spatial distributions in agro-soils through identifying their habitat preferences. This study represents a significant advance in forecasting the responses of agricultural ecosystems to anthropogenic disturbance and thus helps manage soil bacterial communities for better provisioning of key ecosystem services—the ultimate goal of microbial ecology.

Revealing the linkages between community assembly and species coexistence, which is crucial for the understanding of ecosystem diversity and functioning, is a fundamental but rarely investigated subject in microbial ecology. Here we examined archaeal, bacterial, and fungal community assembly in adjacent pairs of maize (water-unsaturated) and rice (water-saturated) fields across different habitats and regions throughout Eastern China. The high-throughput sequencing dataset was analyzed by variation partitioning, null model, and neutral community model analyses. We demonstrated that microbial community assembly was governed more by species sorting than by dispersal limitation in maize fields, and to a lesser extent in rice fields. The relative importance of species sorting in maize soils was greater at low latitudes than at high latitudes, while rice soils exhibited an opposite trend. Microbial co-occurrence associations tended to be higher when communities were primarily driven by dispersal limitation relative to species sorting. There were greater community dissimilarities between maize and rice soils in low-latitude regions, which was consistent with the higher proportion of negative edges in the correlation networks. The results indicate that a balance between species sorting and dispersal limitation mediates species coexistence in soil microbiomes. This study enhances our understanding of contemporary coexistence theory in microbial ecosystems.

Metatranscriptomics is a powerful tool for capturing gene expression patterns in microbial communities and investigating their responses to environmental conditions. Stable isotope probing (SIP) is a method to target specific functional groups of microorganisms in environmental samples. The combination of RNA-SIP with metatranscriptomic analysis enhances the detection and identification of mRNA from target microorganisms. In this chapter we provide a protocol for RNA-SIP, mRNA enrichment, and mRNA preparation for high-throughput sequencing using an example of targeting methanotrophs in rice field soil.

Conductive materials like magnetite nanoparticles (nanoFe3O4) are known to stimulate methanogenesis from syntrophic oxidation of butyrate and propionate. Whether conductive materials increase the decomposition of complex organic substrates like plant residues is unknown. Here, we investigated the effects of biochar and nanoFe3O4 on anaerobic decomposition of rice straw and methanogenesis in a rice paddy soil. The straw decomposition and CH4 production were greatly stimulated by both treatments. Major bacterial populations were Bacteroidetes, Clostridia, Deltaproteobacteria, Actinobacteria and Anaerolineae, while methanogens were predominately Methanosarcina, Methanosaeta and Methanobacterium. Bacteroides were relatively enriched in the biochar treatment, while the Deltaproteobacteria and Anaerolineae were favored by nanoFe3O4. The relative abundance of typical syntrophs including Syntrophomonas, Syntrophus and Smithella were positively correlated with the degradation of intermediate short-chain fatty acids. Notably, the relative abundance of Geobacter, which increased during the incubation, was approximately tenfold greater than all other syntrophs. The co-occurrence of Geobacter together with Methanosarcina and Methanosaeta suggests that the enhanced decomposition of rice straw and methanogenesis by biochar and magnetite may be related to direct interspecies electron transfer involved in the syntrophic oxidation of intermediate products.

Microbial arsenic (As) methylation and demethylation are important components of the As biogeochemical cycle. Arsenic methylation is enhanced under flooded conditions in paddy soils, producing mainly phytotoxic dimethylarsenate (DMAs) that can cause rice straighthead disease, a physiological disorder occurring widely in some rice growing regions. The key microbial groups responsible for As methylation and demethylation in paddy soils are unknown. Three paddy soils were incubated under flooded conditions. DMAs initially accumulated in the soil porewater, followed by a rapid disappearance coinciding with the production of methane. The soil from a rice straighthead disease paddy field produced a much larger amount of DMAs than the other two soils. Using metabolic inhibition, quantification of functional gene transcripts, microbial enrichment cultures and 13C-labeled DMAs, we show that sulfate-reducing bacteria (SRB) and methanogenic archaea are involved in As methylation and demethylation, respectively, controlling the dynamics of DMAs in paddy soils. We present a model of As biogeochemical cycle in paddy soils, linking the dynamics of changing soil redox potential with arsenite mobilization, arsenite methylation and subsequent demethylation driven by different microbial groups. The model provides a basis for controlling DMAs accumulation and incidence of straighthead disease in rice.

Recent studies have shown that application of conductive materials including magnetite and carbon nanotubes (CNTs) can promote the methanogenic decomposition of short-chain fatty acids and even more complex organic matter in anaerobic digesters and natural habitats. The linkage to microbial identity and the mechanisms, however, remain poorly understood. Here, we evaluate the effects of nanoscale magnetite (nanoFe3 O4 ) and multiwalled CNTs on the syntrophic oxidation of propionate in an enrichment obtained from lake sediment. The microbial populations were composed mainly of Smithella, Syntrophomonas, Methanosaeta, Methanosarcina and Methanoregula. In addition to acetate, butyrate was transiently accumulated indicating that propionate was oxidized by Smithella via the dismutation pathway and part of the leaked butyrate was oxidized by Syntrophomonas. Propionate oxidation and CH4 production were significantly accelerated in the presence of nanoFe3 O4 and CNTs. While propionate oxidation was suppressed upon H2 application and suspended completely upon formate application in the control, this suppressive effect was substantially compromised in the presence of nanoFe3 O4 and CNTs. The tests on hydrogenotrophic methanogenesis of a pure culture methanogen and of the enrichment culture without propionate showed negative effect by both materials. The positive effect of nanoFe3 O4 disappeared when it was insulated by surface-coating with silica. Observations made with fluorescence in situ hybridization and scanning electron microscope indicated the extensive formation of microbial cell-conductive material mixture aggregates. Our results suggest that direct interspecies electron transfer is likely activated by the conductive materials and operates in concert with H2 /formate-dependent electron transfer for syntrophic propionate oxidation in the sediment enrichment.

Large variation of CH4 emissions from paddy and wetland ecosystems exists across different geographical locations in China. To obtain mechanistic understanding of this variation, we investigated the dynamics of methanogenesis over the course of glucose degradation in fourteen paddy field soils and five wetland soils collected from different regions of China. The results revealed that the maximal rate (2–3 mM per day) and the total amount (25–30 mM) of CH4 produced were similar across soil samples. The lag phase of methanogenesis, however, differed substantially with the shortest lag phase of 4 days in a paddy soil from north China and the longest of 32 days in a soil from south China, and this difference reflected a general geographical trend among all soils tested. Nitrate was reduced completely within 4 days in all soils. The reduction of Fe(III) and sulfate was completed after 21 days and 29 days, respectively. The depletion time of Fe(III) and sulfate were positively correlated with the lag phase of methanogenesis. Competition for common substrates between methanogens and iron and sulfate reducers, however, does not explain this coincidence because a slow production of CH4 was detected at the very beginning. It appears that the geographical variations in methanogenesis and the reduction of ferric iron and sulfate are related to the variation in soil pH but not to temperature, soil organic C and nutrient conditions in paddy and wetland soils across eastern China.

Microbial syntrophy is a thermodynamically-based cooperation between microbial partners that share the small amounts of free energy for anaerobic growth. To gain insights into the mechanism by which syntrophic microorganisms coordinate their metabolism, we constructed cocultures of propionate-oxidizing Pelotomaculum thermopropionicum and hydrogenotrophic Methanocella conradii and compared them to monocultures. Transcriptome analysis was performed on these cultures using strand-specific mRNA sequencing (RNA-Seq). The results showed that in coculture both P. thermopropionicum and M. conradii significantly upregulated the expression of genes involved in catabolism but downregulated those for anabolic biosynthesis. Specifically, genes coding for the methylmalonyl-CoA pathway in P. thermopropionicum and key genes for methanogenesis in M. conradii were substantially upregulated in coculture compared to monoculture. The putative flavin-based electron bifurcation/confurcation systems in both organisms were also upregulated in coculture. Formate dehydrogenase encoding genes in both organisms were markedly upregulated, indicating that formate was produced and utilized by P. thermopropionicum and M. conradii, respectively. The inhibition of syntrophic activity by formate and 2-bromoethanesulphonate (2-BES) but not H2/CO2 also suggested that formate production was used by P. thermopropionicum for the recycling of intracellular redox mediators. Finally, flagellum-induced signal transduction and amino acids exchange was upregulated for syntrophic interactions. Together, our study suggests that syntrophic organisms employ multiple strategies including global metabolic shift, utilization of electron bifurcation/confurcation and employing formate as an alternate electron carrier to optimize their metabolisms for syntrophic growth.

Females of most species live longer than do males. Furthermore, lifespan-extending interventions in laboratory model organisms are often more effective in females (Regan and Partridge 2013). For instance, genetic and pharmacological suppression of activity of the insulin/insulin-like signalling - target of rapamycin (IIS-TOR) network generally extends female lifespan more than that of males in both Drosophila and mice (Clancy et al. 2001; Selman et al. 2009). We previously showed that attenuation of Ras-dependent IIS signalling by treatment with the FDA-approved MEK inhibitor, trametinib extends lifespan in females (Slack et al. 2015). Here, we demonstrate that trametinib treatment has beneficial effects on female-specific, age-related gut pathologies, similar to those obtained through dietary restriction (Regan et al. 2016). Importantly, we identify Ras inhibition as an effective lifespan-extending manipulation in males as well as females, pointing to parallel mechanisms of lifespan extension by trametinib in both sexes.

It is tempting to develop green energy biotechnology to supplement the depletion of fossil fuel resources and to reduce the greenhouse gas emissions. It has been recently demonstrated that paddy soil can be used to construct the sediment-type microbial fuel cells (MFCs). The mechanisms involved in electricity generation and its effects on methanogenesis, however, remain poorly understood. In the present experiment, we constructed paddy soil MFCs to evaluate the effect of MFC on CH4 emission and to determine the bacterial and archaeal populations associated with MFC anodes. We found that electrical current in MFCs increased rapidly in three weeks, while only minor current was detected in MFCs without rice plants. Methane emission decreased in closed circuit MFCs compared with open circuit MFCs that did not have electricity output. Molecular approaches revealed that the upstream fermentation bacterial community was not significantly affected by MFCs. However, the relative abundances of Geobacteraceae increased markedly in MFCs; and concurrently the classical syntrophs including Syntrophaceae, Syntrophorhabdaceae, Syntrophobacteraceae and Syntrophomonadaceae increased. Among methanogenic archaea, the relative abundance of Methanosaetaceae increased significantly in closed circuit MFCs compared with open-circuit MFCs. Collectively, our study suggests that the activities of electron-transferring bacteria and archaea are significantly promoted in paddy soil MFCs.