Eiko Kuramae

Phosphate nanomaterials, such as hydroxyapatite/β-tricalcium nanoparticles (nHAs) derived from food industry by-products, offer a sustainable alternative to enhance P-use efficiency in agriculture. However, their limited solubility remains a challenge. This study first investigated the mechanisms of P solubilization of salmon and tuna bones (SnHAs and TnHAs) in fifteen strains of phosphate-solubilizing bacteria (PSB) by an in vitro system. Then, best-performing strains were assembled in a consortium and tested in vivo on maize. We hypothesized that combining nHAs and the PSB consortium inoculated as seed coating (SC) outperforms single treatments alone in promoting plant growth and P cycling, and ensures the establishment in plant-soil system without a bacterial reinforcement (BR) by an additional inoculum suspension. The synergistic effect of nHAs and PSB was proved, improving maize root (+22 %) and total plant biomass (+29 %), as well as P (+32 %) and K (66 %) uptake compared to single treatments. With nHAs and SC, P-use efficiency and recovery increased by 25 % and three-fold, respectively, compared to nHAs alone or with bacterial reinforcement. Consistently, root and substrate bacterial biomass were associated with nHAs plus SC, while nHAs alone or with PSB upregulated PHT1;1 and PHT1;2 transporter genes in maize. Finally, linking the in vitro and in vivo system, we demonstrated that propionic acid production and P-solubilization efficiency of PSB co-applied with nHAs are key drivers of maize growth and P uptake. Our findings indicated that co-applying nHAs and PSB through SC offers a sustainable strategy to improve maize P-use efficiency.

Soil pH is one of the most important factors influencing microbial activity and function. In agricultural land, soil acidification results in decline of microbial abundance and diversity, and is associated with increased N₂O emissions. Moreover, soil pH is a crucial factor determining the effect of fertilizer on soil microbial abundance and functionality. However, the interaction effect of soil pH and the application of fertilizer on the abundance of key metabolic pathways involved in nitrogen, carbon, and sulfur cycles remains unclear. Therefore, we modified the original pH of soil (pH 6.0) to pH 4.5 by adding elemental sulfur to the soil and compared the effects of chemical (urea), organic fertilizer (concentrated vinasse), and the combination of both (urea and concentrated vinasse) on soil microbial functions under low soil pH (pH 4.5) and high soil pH (pH 6.0) conditions. Shotgun metagenome sequencing was conducted to obtain microbial functional gene abundance, and the data were analyzed using a model-based statistical approach to determine potential interactions among energy metabolic pathways (i.e., nitrogen, sulfur, methane metabolisms, and carbon fixation). Our result showed the strong effect of soil pH rather than the interaction effect of soil pH and fertilizer treatments on functional gene composition involved in microbial energy metabolisms. Furthermore, we determined nitrogen metabolism most affected by soil pH followed by the sulfur metabolism, methane metabolism, and carbon fixation. When relative abundances of microbial genes were averaged among fertilizer treatments, high soil pH increased gene abundance potentially associated with amino acid synthesis, while low pH increased gene abundance related to denitrifiers denitrification. N₂O emissions were higher under high soil pH conditions, suggesting a small contribution of denitrifiers to N₂O emissions. Among the fertilizer treatments, urea differently impacted gene abundance associated with N₂O and CH₄ emission depending on soil pH. This study provides an overview of microbial metabolic pathways influenced by soil pH and fertilizer treatments, specifically focusing on the microbial functions associated with soil environmental processes.

Nanomaterials are increasingly recognized for their potential in soil remediation. However, their impact on soil microbial communities in contaminated soil remains poorly understood. In this study, we investigated the dynamic effects of sulfonated graphene (SG) following one-time or repeated applications on heavy metal availability and soil microbial communities in long-term heavy metal-contaminated soil over 180 days. Our findings revealed that one-time SG application at 30 mg kg⁻¹ significantly increased the bioavailable cadmium (Cd) and copper (Cu) contents by approximately 30 %–40 % after 2 and 180 days. Repeated SG applications, however, displayed no significant influence on heavy metal availability. One-time SG application, coupled with the increased available Cd, induced significant enrichment of some specific functional bacterial genera involved in glycan biosynthesis metabolism and biosynthesis of other secondary metabolites, thereby decreasing the available contents of heavy metals after 90 days. However, the shifts in bacterial community structure and function were subsequently partially recovered after 180 days. Conversely, repeated SG treatments led to minimal alterations after 90 days while leading to similar shifts in the bacterial community at 60 mg kg⁻¹ after 180 days. The fungal community structure remained largely unaltered across all SG treatments. Intriguingly, SG treatments substantially stimulated fungal biomass, with the stimulation degree dependent on SG dosage. These results provide valuable insights for developing phytoremediation strategies, suggesting tailored SG applications during specific growth phases to optimize remediation efficiency.

Asbestos poses a substantial environmental health risk, and biological treatment offers a promising approach to mitigate its impact by altering its chemical composition. However, the dynamics of microbial co-inoculation in asbestos bioremediation remain poorly understood. This study investigates the effect of microbial single cultures and co-cultures on modifying crocidolite and chrysotile fibers, focusing on the extraction of iron and magnesium. Seventy bacterial and eighty-three fungal strains were isolated from five diverse sites, characterized phylogenetically using the 16S rRNA gene and ITS region, respectively, and assessed for siderophore and organic acid production. Most bacterial strains were identified as Pseudomonas, while Penicillium predominated among fungal strains. Ten bacterial and 25 fungal strains were found to produce both organic compounds. Four microbial co-cultures (one bacterium-bacterium, two fungus-bacterium, and one fungus-fungus) exhibiting synergistic effects in plate assays, alongside their respective single cultures, were incubated with crocidolite and chrysotile. ICP-OES analysis revealed that in crocidolite, the co-culture HRF19–HRB12 removed more iron than their single cultures, while Penicillium TPF36 showed the highest iron removal. The co-culture of two Pseudomonas strains (HRB12–RB5) exhibited the highest magnesium concentration in the supernatant. In chrysotile, the co-culture HRB12–RB5 removed more iron than their individual cultures, with Penicillium TFSF27 exhibiting the highest iron concentration in the solution. Penicillium TFSF27 and the co-culture TFSF27–TPF36 demonstrated the highest magnesium removal. SEM-XRMA analysis showed a significant reduction in iron and magnesium content, confirming elemental extraction from the fibers' structure. This study significantly broadens the range of microbial strains capable of modifying asbestos fibers and underscores the potential of microbial co-cultures in asbestos remediation.

Biostimulants emerged as a versatile tool to modify plant biological processes, by enhancing growth, improving nutrition, increasing stress tolerance, and enhancing crop quality. Among various biostimulant compounds, chitin and gelatin have shown promise in promoting plant growth and enhancing microbial communities. In this study, we investigated the biostimulant effects of chitin, gelatin, and their mixture on cucumber plantlets and associated rhizosphere microbial communities during plantlet production. Cucumber seeds were sown in seedling substrate amended with gelatin, chitin, or a mixture of both biostimulants. Plants were grown at 25°C/21°C with a 16/8 h photoperiod and 75 % humidity. Unamended samples served as controls, while urea was used as a mineral fertilizer control. After 8, 11 and 15 days, rhizosphere samples were collected, DNA was extracted, and the bacterial and fungal communities were assessed by high-throughput sequencing of the 16 S rRNA gene and the ITS region, respectively. Our findings revealed that the application of these biostimulants significantly improved cucumber plantlet growth, with the most pronounced effects 15 days after germination. Gelatin had significantly superior performance compared to chitin. The microbial communities with those amendments were enriched with microbes of genera Cellvibrio, Catenulispora, Arthrobacter, Mortierella, and Penicillium, all known for their production of hydrolytic enzymes such as chitinases, cellulases, and proteases. Overall, this research contributes to a deeper understanding of the biostimulant-mediated interactions between plants and their associated microbial communities, offering potential applications to enhance crop productivity, especially at the plantlet stage while promoting circular economy and environmental sustainability in agriculture.

In the context of global efforts to reduce carbon (C) emissions, several studies have examined the effects of agricultural practices such as straw returning and fertilization on C sequestration by microorganisms. However, our understanding of the specific microbial groups and their roles in long-term C increase remains limited. In this study, a 36-year (1984–2020) farmland experiment was conducted to investigate the impact of bacterial C metabolism on the augmentation of organic C in a Typic Hapludoll (Mollisol) in the black soil region of Jilin Province, Northeast China. Our results demonstrated a noteworthy increase in the diversity of microorganisms in the farmland as a result of long-term straw returning and application of mixed chemical fertilizers. However, by examining the functions of microorganisms involved in C metabolism, it was observed that the effects of fertilization on C metabolism were relatively consistent. This consistency was attributed to a deterministic competitive exclusion process, which minimized the differences between treatment groups. On the other hand, the influence of straw addition on C metabolism appeared to follow a more random pattern. These changes in microbial activity were closely linked to the downregulation of core metabolic pathways related to C metabolism. Notably, long-term fertilization had a negative impact on soil organic C levels, while long-term straw returning plus fertilization resulted in a positive increase in soil organic C. These findings have important implications for enhancing soil organic C and grain yield in the regions with typical black soil.

Farm dairy effluents (FDE) from washing the milking parlor contain manure, urine, and chemicals and constitute a large amount of wastewater. Applying FDE as soil fertilizers to pastures can enhance forage yield and improve soil nutrient status. Since the dairy industry is increasingly attempting to maximize returns through better utilization of forage with lesser inputs, there is demand for a supply of FDE as fertilizers. Nevertheless, the impact of this practice on soil microbiota remains largely unexplored. It must be studied before large-scale soil disposal to avoid diminishing microbial diversity or enhancing pathogen abundance. This study evaluated the effects of applying lagoon-stored (Lagoon) and raw dairy effluents (Raw) at a rate of 50 kg N ha⁻¹ in four equal doses, in comparison to urea fertilization, on soil fertility and the activity, abundance, and community structure of soil microbiota. Raw was obtained after solid separation, and Lagoon corresponds to the Raw stationed in a two-lagoon system. Microbial activity was assessed as basal respiration, potentially mineralizable N, potential nitrification activity, and enzymatic activities. The catabolic activity of the microbial community was evaluated using Biolog Ecoplates™. Bacterial and fungal community composition and diversity were analyzed through amplicon sequencing of 16S rRNA and ITS2. The application of FDE benefited soil fertility and microbial activity. Lagoon had the most potent effects on soil available P and extractable K⁺, Na⁺, Mg²⁺ and Ca²⁺. Soil treated with Raw displayed higher microbial activities, such as dehydrogenase, basal respiration, urease, and potentially mineralizable N, than the other treatments. FDE did not significantly alter the microbial composition, abundance, or functional diversity. In conclusion, in this short-term trial, despite changes in soil chemical properties and microbial activity, the composition and diversity of the bacterial and fungal communities remained unaffected by FDE irrigation.

With the expected growth of insect mass rearing industry, residual streams are set to become increasingly available. However, before these residues can be used as organic amendments, more knowledge of the decomposition dynamics and the associated microbial communities is needed. This study aimed to investigate the decomposition, N-mineralization, and fungal/bacterial community composition over a 16-week incubation period of exuviae and frass from BSF, mealworm, and house cricket in pots containing arable soil. The decomposition of insect residues in litterbags was rapid, with over 50% weight loss observed within two weeks. The release of mineral nitrogen from dispersed insect materials was highest during the initial two weeks, particularly evident in soils treated with exuviae compared to their frass counterparts. Moreover, soil amendment with insect residues enriched chitinolytic soil microbial inhabitants belonging to Gammaproteobacteria, Bacilli, Actinobacteria, and Mortierellomycetes. A comparison of soil amendments with sterilized and non-sterilized mealworm exuviae indicated minimal influence of microbial propagules on the composition of bacterial decomposers, though a notable impact on the fungal community was observed. These findings suggest that amendments with insect residues show promise in enhancing natural biocontrol by stimulating microbial antagonists of plant-pathogenic fungi, thereby presenting a potential tool for integration into soil-borne disease management strategies.

In this study, we adopt an interdisciplinary approach, integrating agronomic field experiments with soil chemistry, molecular biology techniques, and statistics to investigate the impact of organic residue amendments, such as vinasse (a by-product of sugarcane ethanol production), on soil microbiome and greenhouse gas (GHG) production. The research investigates the effects of distinct disturbances, including organic residue application alone or combined with inorganic N fertilizer on the environment. The methods assess soil microbiome dynamics (composition and function), GHG emissions, and plant productivity. Detailed steps for field experimental setup, soil sampling, soil chemical analyses, determination of bacterial and fungal community diversity, quantification of genes related to nitrification and denitrification pathways, measurement and analysis of gas fluxes (N₂O, CH₄, and CO₂), and determination of plant productivity are provided. The outcomes of the methods are detailed in our publications (Lourenço et al., 2018a; Lourenço et al., 2018b; Lourenço et al., 2019; Lourenço et al., 2020). Additionally, the statistical methods and scripts used for analyzing large datasets are outlined. The aim is to assist researchers by addressing common challenges in large-scale field experiments, offering practical recommendations to avoid common pitfalls, and proposing potential analyses, thereby encouraging collaboration among diverse research groups. • Interdisciplinary methods and scientific questions allow for exploring broader interconnected environmental problems. • The proposed method can serve as a model and protocol for evaluating the impact of soil amendments on soil microbiome, GHG emissions, and plant productivity, promoting more sustainable management practices. • Time-series data can offer detailed insights into specific ecosystems, particularly concerning soil microbiota (taxonomy and functions).

Plant Growth-Promoting Microbes (PGPM) have the potential to enhance sustainable agriculture, but there is still a limited understanding of how the complex interplay between plant genetic variability, the native soil community, and soil nutrients affects PGPM recruitment. To address this challenge, we investigated the impact of bacteria isolates and arbuscular mycorrhizal fungi (AMF) along with their accompany microbiome (AMFc) derived from a wild chrysanthemum on the growth of five different commercial chrysanthemum cultivars (Chic, Chic 45, Chic Cream, Haydar and Barolo), as well as their rhizosphere microbiomes, within a nutrient-rich complex substrate environment. We found 23 bacterial strains capable of producing siderophore, 14 strains capable of producing Indole-3-acetic acid, and 18 strains capable of solubilizing phosphate. The AMFc had six AMF species, and the bacterial and fungal communities associated with AMF belonged to different phyla. Using generalized joint models, we investigated the impact of the three most effective bacterial strains and the AMFc on plant growth (shoot and root dry mass) while integrating information on plant genotype, environment, and microbes. The impact of PGPM inoculation varied from positive to negative effects depending on the cultivar, with Chic Cream showing the highest increase in root biomass after inoculation with both bacterial strain SMF006 (57 %) and AMFc inoculation (79 %). Our study demonstrates that PGPM from wild relative can impact the growth and assembly of the chrysanthemum root microbiome, but this impact is cultivar-dependent. Furthermore, inoculation with a complex AMF containing community (AMFc) induced greater changes in the rhizosphere microbiome than with a single bacterial isolate. Our study shows that inoculation of a complex community of beneficial microbes results in more effective plant growth promotion.

Diversity of the soil microbial community is an important factor affecting its stability against disturbance. However, the impact of the decline in soil microbial diversity on the stability of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) is not known, particularly considering the repeated soil nutrient disturbances occurring in modern agricultural systems. Here, we conducted a microcosm experiment and modified the soil microbial diversity using the dilution-to-extinction approach to determine the stability and population dynamics of AOB and AOA communities with repeated nitrogen (N) fertilizer application. Our results demonstrated that the AOB community became more abundant and stable against repeated disturbances by N in the treatments with the highest microbial diversity. In contrast, the abundance of AOA decreased following repeated N fertilizer application, regardless of the microbial diversity. Notably, during the initial application phase, AOA displayed a potential for increased abundance in treatments with high soil microbial diversity. These findings highlight that the soil microbial diversity controls the stability of ammonia oxidizers during short-interval repeated N disturbances.

Periodic inoculations of soil-beneficial microbes can increase their populations, but they also act as recurring biotic disturbances on the native microbial community. Soil rare and abundant microorganisms disproportionally shape the community diversity and stability. Uncovering their dynamic responses to recurring biotic disturbances and the underlying driving factors helps improve our understanding of the inoculation effects. Here, we imposed temporally recurring biotic disturbances by inoculating soils with phosphate-solubilizing bacteria, nitrogen-fixing bacteria, and the combination of both, with the overall aim of studying the successive responses of bacterial and fungal subcommunities along a rarity index. Our results showed that, in both bacterial and fungal communities, the relatively rare taxa exhibited higher diversity than the abundant taxa, and the relative abundance of rare taxa increased with recurring disturbances. However, the responses of rare and abundant taxa to inoculations were different between bacteria and fungi and were related to time and inoculation frequency. The rarer bacteria and the more abundant fungi explained most of the effects of inoculations on the resident microbial community. About 20 percent of the microbes changed their rarity categories over time, and most of the changes and interactions occurred within the rarer taxa during the first 45 days. Modeling analyses and co-occurrence networks indicated that microbial interactions, soil biochemical factors, and inoculation time drove the shifts of subcommunities. In summary, relatively rare bacteria and relatively abundant fungi play major roles in understanding the impacts of recurring biotic disturbances, while the conditionality of microbial rarity is dependent on both biotic and abiotic factors.

The bacteria that dominate and become enriched in the rhizosphere during continuous cropping are of increasing interest, as they can greatly adapt to the rhizosphere. However, there are still little knowledge about the general composition and function of these bacteria. In this study, we planted tomatoes in three different soils for three planting cycles and used both high-throughput sequencing and culture-dependent workflows. Despite significant differences in bacterial communities from the initial soils, we observed a similar succession in the rhizosphere bacterial community compositions. We identified certain bacteria that were gradually enriched and potentially beneficial, such as Rhizobium and Flavobacterium. However, some other potentially beneficial bacteria, such as Massilia and Lysobacter, were gradually depleted. Additionally, we found that predicted functions related to xenobiotic biodegradation, nutrient metabolism and antibiotic biosynthesis were enriched in different rhizosphere soils. Beijerinckia fluminensis GR2, which was gradually enriched in all tested soils, significantly inhibited the growth of Ralstonia solanacearum and protected the host from infection. Our study provides new insights into the assembly mechanism of gradually enriched bacteria and their role as plant-beneficial microbes that adapt well to the rhizosphere.

Introducing probiotics to soil is a sustainable way to stimulate the production of plant metabolites. However, the soil-resident microbes may compromise the efficiency of probiotics. To date, it remains challenging to integrate the effects of probiotics on plant performance with soil microbiome changes. Using Cyclocarya paliurus (Batal.) Iljinsk as a model medicinal plant and two types of probiotic consortia combined with organic fertilizer at three levels (low: 0.5, medium: 1.0, and high: 1.5 kg·plant⁻¹), we examined the impacts of three fertilization regimes (O: organic fertilizer, OMF: O coupled with Bacillus megaterium and Pseudomonas fluorescens, OCB: O coupled with Azotobacter chroococcum and Azospirillum brasilense) on plant metabolites and nutrient stoichiometry after three-year applications and identified the key soil microbes relating to the accumulation of plant metabolites via generalized joint attribute model (GJAM) analysis. Our results indicated that the concentration of flavonoids reached 36.9 mg·g⁻¹ in OCB treatment at a low level, and 30.0 mg·g⁻¹ in OMF treatment at a medium level, both were significantly higher than that in O treatment (25.8 mg·g⁻¹ on average). Furthermore, the accumulations of metabolites were associated with plant nutrient acquisition and C: N: P stoichiometry. GJAM analysis showed that higher fertilizer levels restricted the influence of probiotic consortia on the variance of plant-soil-microbe system, with fewer differences observed between fertilizer types. Specific soil microbes were predicted as potential indicators that may assist or impede the effects of probiotics on plant metabolite production. The predictions were further tested in a comparative pot experiment, and the effects of common indicators in both pot and field experiments were consistently associated with probiotics’ addition. This study reveals that the effects of probiotics on plant metabolites are associated with fertilization regimes and soil-indigenous microbes. Identifying microbial indicators will help to understand the probiotics' effects and further improve plant productivity.

Cover crops are a potential pathway for ecological cultivation in agricultural systems. In tropical no-till agricultural systems, the maintenance of residues on the soil surface and the addition of nitrogen (N) benefit the growth and grain yield of cash crops as well as the chemical and physical properties of the soil. However, the effects of these management practices on the soil microbiota are largely unknown. Here, we evaluated the effects of the timing of N application as a pulse disturbance and the growth of different cover crop species before maize in rotation on soil properties, maize productivity, and soil bacterial and fungal community diversity and composition. N fertilizer was applied either on live cover crops (palisade grass or ruzigrass), on cover crop straw just before maize seeding or in the maize V₄ growth stage. Soils previously cultivated with palisade grass established similar microbial communities regardless of N application timing, with increases in total bacteria, total archaea, nutrients, and the C:N ratio. The soil microbial alpha diversity in treatments with palisade grass did not vary with N application timing, whereas the bacterial and fungal diversities in the treatments with ruzigrass decreased when N was applied to live ruzigrass or maize in the V₄ growth stage. We conclude that palisade grass is a more suitable cover crop than ruzigrass, as palisade grass enhanced soil microbial diversity and maize productivity regardless of N application timing. Ruzigrass could be used as an alternative to palisade grass when N is applied during the straw phase. However, considering the entire agricultural system (soil–plant–microbe), ruzigrass is not as efficient as palisade grass in tropical no-till cover crop–maize rotation systems. Palisade grass is a suitable cover crop alternative for enhancing maize productivity, soil chemical properties and nutrient cycling, regardless of the timing of N application. Additionally, this study demonstrates that a holistic approach is valuable for evaluating soil diversity and crop productivity in agricultural systems.

Nitrous oxide (N₂O) production in tropical soils cultivated with sugarcane is associated with ammonia-oxidizing bacteria (AOB) and fungal denitrifiers. However, the taxonomic identities and the community diversities, compositions, and structures of AOB and fungal denitrifiers in these soils are not known. Here, we examined the effects of applying different concentrations of an organic recycled residue (vinasse: regular non-concentrated or 5.8-fold concentrated) on the dynamics of AOB and fungal denitrifier community diversity and composition and greenhouse gas emissions during the sugarcane cycle in two different seasons, rainy and dry. DNA was extracted from soil samples collected at six timepoints to determine the dynamics of amoA-AOB and nirK-fungal community diversity and composition by amplicon sequencing with gene-specific primers. Bacterial and archaeal amoA, fungal and bacterial nirK, bacterial nirS and nosZ, total bacteria (16S rRNA) and total fungi (18S rRNA) were quantified by real-time PCR, and N₂O and CO₂ emissions were measured. The genes amoA-AOB and bacterial nirK clade II correlated with N₂O emissions, followed by fungal nirK. The application of inorganic nitrogen fertilizer combined with organic residue, regardless of concentration, did not affect the diversity and structure of the AOB and fungal denitrifier communities but increased their abundances and N₂O emissions. Nitrosospira sp. was the dominant AOB, while unclassified fungi were the dominant fungal denitrifiers. Furthermore, the community structures of AOB and fungal denitrifiers were affected by season, with dominance of uncultured Nitrosospira and unclassified fungi in the rainy season and the genera Nitrosospira and Chaetomium in the dry season. Nitrosospira, Chaetomium, Talaromyces purpureogenus, and Fusarium seemed to be the main genera governing N₂O production in the studied tropical soils. These results highlight the importance of deciphering the main players in N₂O production and demonstrate the impact of fertilization on soil microbial N functions.

Soil microbial communities are essential components of agroecological ecosystems that influence soil fertility, nutrient turnover, and plant productivity. Metagenomics data are increasingly easy to obtain, but studies of soil metagenomics face three key challenges: (1) accounting for soil physicochemical properties; (2) incorporating untreated controls; and (3) sharing data. Accounting for soil physicochemical properties is crucial for better understanding the changes in soil microbial community composition, mechanisms, and abundance. Untreated controls provide a good baseline to measure changes in soil microbial communities and separate treatment effects from random effects. Sharing data increases reproducibility and enables meta-analyses, which are important for investigating overall effects. To overcome these challenges, we suggest establishing standard guidelines for the design of experiments for studying soil metagenomics. Addressing these challenges will promote a better understanding of soil microbial community composition and function, which we can exploit to enhance soil quality, health, and fertility.

Chemolitho-autotrophic microorganisms like the nitrite-oxidizing Nitrobacter winogradskyi create an environment for heterotrophic microorganisms that profit from the production of organic compounds. It was hypothesized that the assembly of a community of heterotrophic microorganisms around N. winogradskyi depends on the ecosystem from which the heterotrophs are picked. To test this hypothesis, pure cultures of N. winogradskyi were grown in continuously nitrite-fed bioreactors in a mineral medium free of added organic carbon that had been inoculated with diluted sewage sludge or with a suspension from a grassland soil. Samples for chemical and 16S rRNA gene amplicon analyses were taken after each volume change in the bioreactor. At the end of the enrichment runs, samples for shotgun metagenomics were also collected. Already after two volume changes, the transformations in community structure became less dynamic. The enrichment of heterotrophs from both sewage and soil was highly stochastic and yielded different dominant genera in most of the enrichment runs that were independent of the origin of the inoculum. Hence, the hypothesis had to be refuted. Notwithstanding the large variation in taxonomic community structure among the enrichments, the functional compositions of the communities were statistically not different between soil- and sludge-based enrichments. IMPORTANCE In the process of aerobic nitrification, nitrite-oxidizing bacteria together with ammonia-oxidizing microorganisms convert mineral nitrogen from its most reduced appearance, i.e., ammonium, into its most oxidized form, i.e., nitrate. Because the form of mineral nitrogen has large environmental implications, nitrite-oxidizing bacteria such as Nitrobacter winogradskyi play a central role in the global biogeochemical nitrogen cycle. In addition to this central role, the autotrophic nitrite-oxidizing bacteria also play a fundamental role in the global carbon cycle. They form the basis of heterotrophic food webs, in which the assimilated carbon is recycled. Little is known about the heterotrophic microorganisms that participate in these food webs, let alone their assembly in different ecosystems. This study showed that the assembly of microbial food webs by N. winogradskyi was a highly stochastic process and independent of the origin of the heterotrophic microorganisms, but the functional characteristics of the different food webs were similar.

The increase in sequencing capacity has amplified the number of taxonomically unclassified sequences in most databases. The classification of such sequences demands phylogenetic tree construction and comparison to currently classified sequences, a process that demands the processing of large amounts of data and use of several different software. Here, we present PhyloFunDB, a pipeline for extracting, processing, and inferring phylogenetic trees from specific functional genes. The goal of our work is to decrease processing time and facilitate the grouping of sequences that can be used for improved taxonomic classification of functional gene datasets.

Soil microbiota plays fundamental roles in maintaining ecosystem functions and services, including biogeochemical processes and plant productivity. Despite the ubiquity of soil microorganisms from the topsoil to deeper layers, their vertical distribution and contribution to element cycling in subsoils remain poorly understood. Here, nine soil profiles (0 to 135 cm) were collected at the local scale (within 300 km) from two canonical paddy soil types (Fe-accumuli and Hapli stagnic anthrosols), representing redoximorphic and oxidative soil types, respectively. Variations with depth in edaphic characteristics and soil bacterial and diazotrophic community assemblies and their associations with element cycling were explored. The results revealed that nitrogen and iron status were the most distinguishing edaphic characteristics of the two soil types throughout the soil profile. The acidic Fe-accumuli stagnic anthrosols were characterized by lower concentrations of free iron oxides and total iron in topsoil and ammonia in deeper layers compared with the Hapli stagnic anthrosols. The bacterial and diazotrophic community assemblies were mainly shaped by soil depth, followed by soil type. Random forest analysis revealed that nitrogen and iron cycling were strongly correlated in Fe-accumuli stagnic anthrosol, whereas in Hapli soil, available sulfur was the most important variable predicting both nitrogen and iron cycling. The distinctive biogeochemical processes could be explained by the differences in enrichment of microbial taxa between the two soil types. The main discriminant clades were the iron-oxidizing denitrifier Rhodanobacter, Actinobacteria, and diazotrophic taxa (iron-reducing Geobacter, Nitrospirillum, and Burkholderia) in Fe-accumuli stagnic anthrosol and the sulfur-reducing diazotroph Desulfobacca in Hapli stagnic anthrosol. IMPORTANCE Rice paddy ecosystems support nearly half of the global population and harbor remarkably diverse microbiomes and functions in a variety of soil types. Diazotrophs provide significant bioavailable nitrogen in paddy soil, priming nitrogen transformation and other biogeochemical processes. This study provides a novel perspective on the vertical distribution of bacterial and diazotrophic communities in two hydragric anthrosols. Microbiome analysis revealed divergent biogeochemical processes in the two paddy soil types, with a dominance of nitrogen-iron cycling processes in Fe-accumuli stagnic anthrosol and sulfur-nitrogen-iron coupling in Hapli stagnic anthrosol. This study advances our understanding of the multiple significant roles played by soil microorganisms, especially diazotrophs, in biogeochemical element cycles, which have important ecological and biogeochemical ramifications.

BACKGROUND: The assembly of the rhizomicrobiome, i.e., the microbiome in the soil adhering to the root, is influenced by soil conditions. Here, we investigated the core rhizomicrobiome of a wild plant species transplanted to an identical soil type with small differences in chemical factors and the impact of these soil chemistry differences on the core microbiome after long-term cultivation. We sampled three natural reserve populations of wild rice (i.e., in situ) and three populations of transplanted in situ wild rice grown ex situ for more than 40 years to determine the core wild rice rhizomicrobiome.

RESULTS: Generalized joint attribute modeling (GJAM) identified a total of 44 amplicon sequence variants (ASVs) composing the core wild rice rhizomicrobiome, including 35 bacterial ASVs belonging to the phyla Actinobacteria, Chloroflexi, Firmicutes, and Nitrospirae and 9 fungal ASVs belonging to the phyla Ascomycota, Basidiomycota, and Rozellomycota. Nine core bacterial ASVs belonging to the genera Haliangium, Anaeromyxobacter, Bradyrhizobium, and Bacillus were more abundant in the rhizosphere of ex situ wild rice than in the rhizosphere of in situ wild rice. The main ecological functions of the core microbiome were nitrogen fixation, manganese oxidation, aerobic chemoheterotrophy, chemoheterotrophy, and iron respiration, suggesting roles of the core rhizomicrobiome in improving nutrient resource acquisition for rice growth. The function of the core rhizosphere bacterial community was significantly (p < 0.05) shaped by electrical conductivity, total nitrogen, and available phosphorus present in the soil adhering to the roots.

CONCLUSION: We discovered that nitrogen, manganese, iron, and carbon resource acquisition are potential functions of the core rhizomicrobiome of the wild rice Oryza rufipogon. Our findings suggest that further potential utilization of the core rhizomicrobiome should consider the effects of soil properties on the abundances of different genera. Video Abstract.

Slash-and-burn clearing of forest typically results in increase in soil nutrient availability.However, the impact of these nutrients on the soil microbiome is not known. Using next generation sequencing of 16S rRNA gene and shotgun metagenomic DNA,we compared the structure and the potential functions of bacterial community in forest soils to deforested  soils in the Amazon region and related the differences to soil chemical factors.
Deforestation decreased soil organic matter content and factors linked to soil acidity and raised soil pH, base saturation and exchangeable bases. Concomitant to expected changes in soil chemical factors, we observed an increase in the alpha diversity of the bacterial microbiota and relative abundances of putative copiotrophic bacteria such as Actinomycetales and a decrease in the relative abundances of bacterial taxa such as Chlamydiae, Planctomycetes and Verrucomicrobia in the deforested soils. We did not observe an increase in genes related to microbial nutrient metabolism in deforested soils. However,
we did observe changes in community functions such as increases in DNA repair, protein processing, modification, degradation and folding functions, and these functions might reflect adaptation to changes in soil characteristics due to forest clear-cutting and burning. In addition, there were changes in the composition of the bacterial groups associated with metabolism-related functions. Co-occurrence microbial network analysis identified distinct phylogenetic patterns for forest and deforested soils and suggested relationships between Planctomycetes and aluminium content, and Actinobacteria and nitrogen sources in Amazon soils. The results support taxonomic and functional adaptations in the soil bacterial community following deforestation. We hypothesize that these microbial adaptations may serve as a buffer to drastic changes in soil fertility after slashand-burning deforestation in the Amazon region.

To contribute to our understanding of the genome complexity of sugarcane, we undertook a large-scale expressed sequence tag (EST) program. More than 260,000 cDNA clones were partially sequenced from 26 standard cDNA libraries generated from different sugarcane tissues. After the processing of the sequences, 237,954 high-quality ESTs were identified. These ESTs were assembled into 43,141 putative transcripts. Of the assembled sequences, 35.6% presented no matches with existing sequences in public databases. A global analysis of the whole SUCEST data set indicated that 14,409 assembled sequences (33% of the total) contained at least one cDNA clone with a full-length insert. Annotation of the 43,141 assembled sequences associated almost 50% of the putative identified sugarcane genes with protein metabolism, cellular communication/signal transduction, bioenergetics, and stress responses. Inspection of the translated assembled sequences for conserved protein domains revealed 40,821 amino acid sequences with 1415 Pfam domains. Reassembling the consensus sequences of the 43,141 transcripts revealed a 22% redundancy in the first assembling. This indicated that possibly 33,620 unique genes had been identified and indicated that >90% of the sugarcane expressed genes were tagged.

Xylella fastidiosa is a xylem-dwelling, insect-transmitted, gamma-proteobacterium that causes diseases in many plants, including grapevine, citrus, periwinkle, almond, oleander, and coffee. X. fastidiosa has an unusually broad host range, has an extensive geographical distribution throughout the American continent, and induces diverse disease phenotypes. Previous molecular analyses indicated three distinct groups of X. fastidiosa isolates that were expected to be genetically divergent. Here we report the genome sequence of X. fastidiosa (Temecula strain), isolated from a naturally infected grapevine with Pierce's disease (PD) in a wine-grapegrowing region of California. Comparative analyses with a previously sequenced X. fastidiosa strain responsible for citrus variegated chlorosis (CVC) revealed that 98% of the PD X. fastidiosa Temecula genes are shared with the CVC X. fastidiosa strain 9a5c genes. Furthermore, the average amino acid identity of the open reading frames in the strains is 95.7%. Genomic differences are limited to phage-associated chromosomal rearrangements and deletions that also account for the strain-specific genes present in each genome. Genomic islands, one in each genome, were identified, and their presence in other X. fastidiosa strains was analyzed. We conclude that these two organisms have identical metabolic functions and are likely to use a common set of genes in plant colonization and pathogenesis, permitting convergence of functional genomic strategies.

In a search of the sugarcane expressed sequence tag (SUCEST) database we located three full-length cDNAs (SCCCLR1022D05.g, SCCCRZ1001D02.g and SCBFLR1026E02.g) encoding the 14-3-3 proteins from sugarcane (Saccharum officinarum). The encoded proteins were identified based on the clustering of the expressed sequence tags and were shown to encode proteins similar to 14-3-3 proteins of other monocotyledonous plants. Cluster SCCCLR1022D05.g was 99% similar to the maize 14-3-3-like protein (gi1345587) while cluster SCCCRZ1001D02.g shared 96% and SCBFLR1026E02.g 94% similarity with the 14-3-3 protein of rice (gi7435022). Although 14-3-3 proteins have been reported to be specific to particular species, tissue or organ from which they were isolated, all three sugarcane clusters were found to be expressed in several tissues.