Microbial and viral communities transform the chemistry of Earth's ecosystems, yet the specific reactions catalyzed by these biological engines are hard to decode due to the absence of a scalable, metabolically resolved, annotation software. Here, we present DRAM (Distilled and Refined Annotation of Metabolism), a framework to translate the deluge of microbiome-based genomic information into a catalog of microbial traits. To demonstrate the applicability of DRAM across metabolically diverse genomes, we evaluated DRAM performance on a defined, in silico soil community and previously published human gut metagenomes. We show that DRAM accurately assigned microbial contributions to geochemical cycles and automated the partitioning of gut microbial carbohydrate metabolism at substrate levels. DRAM-v, the viral mode of DRAM, established rules to identify virally-encoded auxiliary metabolic genes (AMGs), resulting in the metabolic categorization of thousands of putative AMGs from soils and guts. Together DRAM and DRAM-v provide critical metabolic profiling capabilities that decipher mechanisms underpinning microbiome function.

Syntrophic interaction occurs during anaerobic fermentation of organic substances forming methane as the final product. H2 and formate are known to serve as the electron carriers in this process. Recently, it has been shown that direct interspecies electron transfer (DIET) occurs for syntrophic CH4 production from ethanol and acetate. Here, we constructed paddy soil enrichments to determine the involvement of DIET in syntrophic butyrate oxidation and CH4 production. The results showed that CH4 production was significantly accelerated in the presence of nanoFe3 O4 in all continuous transfers. This acceleration increased with the increase of nanoFe3 O4 concentration but was dismissed when Fe3 O4 was coated with silica that insulated the mineral from electrical conduction. NanoFe3 O4 particles were found closely attached to the cell surfaces of different morphology, thus bridging cell connections. Molecular approaches, including DNA-based stable isotope probing, revealed that the bacterial Syntrophomonadaceae and Geobacteraceae, and the archaeal Methanosarcinaceae, Methanocellales and Methanobacteriales, were involved in the syntrophic butyrate oxidation and CH4 production. Among them, the growth of Geobacteraceae strictly relied on the presence of nanoFe3 O4 and its electrical conductivity in particular. Other organisms, except Methanobacteriales, were present in enrichments regardless of nanoFe3 O4 amendment. Collectively, our study demonstrated that the nanoFe3 O4 -facilitated DIET occurred in syntrophic CH4 production from butyrate, and Geobacter species played the key role in this process in the paddy soil enrichments.

ABSTRACTPropionate is one of the major intermediary products in the anaerobic decomposition of organic matter in wetlands and paddy fields. Under methanogenic conditions, propionate is decomposed through syntrophic interaction between proton-reducing and propionate-oxidizing bacteria and H2-consuming methanogens. Temperature is an important environmental regulator; yet its effect on syntrophic propionate oxidation has been poorly understood. In the present study, we investigated the syntrophic oxidation of propionate in a rice field soil at 15°C and 30°C. [U-13C]propionate (99 atom%) was applied to anoxic soil slurries, and the bacteria and archaea assimilating13C were traced by DNA-based stable isotope probing.Syntrophobacterspp.,Pelotomaculumspp., andSmithellaspp. were found significantly incorporating13C into their nucleic acids after [13C]propionate incubation at 30°C. The activity ofSmithellaspp. increased in the later stage, and concurrently that ofSyntrophomonasspp. increased. AceticlasticMethanosaetaceaeand hydrogenotrophicMethanomicrobialesandMethanocellalesacted as methanogenic partners at 30°C. Syntrophic oxidation of propionate also occurred actively at 15°C.Syntrophobacterspp. were significantly labeled with13C, whereasPelotomaculumspp. were less active at this temperature. In addition,Methanomicrobiales,Methanocellales, andMethanosarcinaceaedominated the methanogenic community, whileMethanosaetaceaedecreased. Collectively, temperature markedly influenced the activity and community structure of syntrophic guilds degrading propionate in the rice field soil. Interestingly,Geobacterspp. and some other anaerobic organisms likeRhodocyclaceae,Acidobacteria,Actinobacteria, andThermomicrobiaprobably also assimilated propionate-derived13C. The mechanisms for the involvement of these organisms remain unclear.

The methanogenic degradation of oil hydrocarbons can proceed through syntrophic partnerships of hydrocarbon-degrading bacteria and methanogenic archaea [1][2][3] . However, recent culture-independent studies have suggested that the archaeon 'Candidatus Methanoliparum' alone can combine the degradation of long-chain alkanes with methanogenesis 4,5 . Here we cultured Ca. Methanoliparum from a subsurface oil reservoir. Molecular analyses revealed that Ca. Methanoliparum contains and overexpresses genes encoding alkyl-coenzyme M reductases and methyl-coenzyme M reductases, the marker genes for archaeal multicarbon alkane and methane metabolism. Incubation experiments with different substrates and mass spectrometric detection of coenzyme-M-bound intermediates confirm that Ca. Methanoliparum thrives not only on a variety of long-chain alkanes, but also on n-alkylcyclohexanes and n-alkylbenzenes with long n-alkyl (C ≥13 ) moieties. By contrast, short-chain alkanes (such as ethane to octane) or aromatics with short alkyl chains (C ≤12 ) were not consumed. The wide distribution of Ca. Methanoliparum 4-6 in oil-rich environments indicates that this alkylotrophic methanogen may have a crucial role in the transformation of hydrocarbons into methane.In subsurface oil reservoirs and marine oil seep sediments, microorganisms use hydrocarbons as a source of energy and carbon 7,8 . The microorganisms preferentially consume alkanes, cyclic and aromatic compounds, leaving an unresolved complex mixture as residue and thereby altering the quality of the oil 7,8 . In the absence of sulfate, microorganisms couple anaerobic hydrocarbon degradation to methane formation 1,9,10 . This reaction was originally demonstrated by Zengler et al 2 as methanogenic 'microbial alkane cracking', and a large number of studies have shown that it can be performed in syntrophic interactions of bacteria and archaea 11 . In this syntrophy, the bacteria ferment the oil to acetate, carbon dioxide and hydrogen, while hydrogenotrophic and/or acetotrophic methanogenic archaea use the products for methanogenesis 1,2,11 .Diverse anaerobic hydrocarbon activation mechanisms exist, including the well-studied fumarate addition pathway catalysed by glycyl radical enzymes 12 . This mechanism is widespread among bacteria that thrive on alkanes of various chain lengths and other hydrocarbons 12,13 . By contrast, several archaeal lineages activate gaseous alkanes with the help of a specific type of methyl-coenzyme M reductase (MCR), an enzyme that was originally described to catalyse the reduction of methyl-coenzyme M (methyl-CoM) to methane in methanogens 14 . Anaerobic methanotrophic archaea use canonical MCRs to activate methane into methyl-CoM, which is then oxidized to CO 2 . Short-chain alkane-oxidizing archaea contain divergent variants of this enzyme, which are known as alkyl-CoM reductases (ACRs). Analogous to the methane-activating MCR, ACRs activate multicarbon alkanes to form CoM-bound alkyl units [15][16][17] . The cultured alkane-oxidizing archaea oxidize sho...

DNA-based stable-isotope probing was applied to identify the active microorganisms involved in syntrophic butyrate oxidation in paddy field soil. After 14 and 21 days of incubation with [U-13 C]butyrate, the bacterial Syntrophomonadaceae and the archaeal Methanosarcinaceae and Methanocellales incorporated substantial amounts of 13 C label into their nucleic acids. Unexpectedly, members of the Planctomycetes and Chloroflexi were also labeled with 13 C by yet-unclear mechanisms.Butyrate is one of the important intermediates in the degradation of organic matter in anoxic environments (3)(4)(5)22). The degradation of butyrate to H 2 , formate, and acetate is endergonic under standard conditions. This thermodynamic barrier, however, can be overcome by the syntrophic interaction between butyrate-oxidizing bacteria and methanogenic archaea, which keep the products H 2 , formate, and acetate at low concentrations (23). A few strains involved in syntrophic butyrate oxidation have been isolated into pure cultures (e.g., see references 15, 16, and 30). These organisms represent a thermodynamically extreme lifestyle, since even in the optimum syntrophic association with methanogens, the Gibbs free energy available for syntrophic butyrate oxidizers is still close to the thermodynamic limit (⌬G 0Ј ϭ Ϫ20 kJ per reaction). Recently, genome sequences of two syntrophic butyrate oxidizers (Syntrophomonas wolfei and Syntrophus aciditrophicus) (17, 24) revealed that they have limited fermentation and respiration mechanisms but possess multiple copies of ␤-oxidation genes and sets of reverse electron transfer machineries which are essential for the syntrophic oxidation of butyrate.Studies on natural environments, however, are very scarce (1, 5). Acetate, propionate, and butyrate are the most important intermediates during the degradation of organic residues in paddy field soils (4, 22). Only two studies known to us so far, however, have been conducted to determine the syntrophic degradation of propionate (12) and acetate (7), and none have been done on syntrophic butyrate oxidation in paddy field soil. Therefore, our objectives were to investigate syntrophic butyrate oxidation and identify the active organisms responsible for this process in a Chinese paddy field soil using nucleic acid-based stable-isotope probing (SIP), which has been proven to be powerful in linking the identities of microorganisms in environments with their specific functions (2).Soil sample and anoxic incubation. Paddy field soil was collected from an experimental station of the China National Rice Research Institute in Hangzhou, China (30 o 04Ј37ЉN, 119 o 54Ј37ЉE). The soil was a clay loam and had the following properties as measured by standard methods (18): pH 6.7, a cation exchange capacity of 14.4 cmol kg Ϫ1 , an organic C content of 24.2 g kg Ϫ1 , and a total N content of 2.3 g kg Ϫ1 . Soil was air dried and passed through 2-mm sieves. Three-gram soil samples were weighed in 15-ml serum bottles and mixed with 4.5 ml distilled anoxic water. The vials were c...

Methane is an important greenhouse gas and propionate is next to acetate the main intermediate (average 23%) of the carbon flow to CH in paddy fields. Sulfate (e.g., gypsum) application can reduce CH emissions up to 70%. However, the effect of gypsum application on propionate degradation and the microbial communities involved are not well understood. Therefore, we studied propionate-dependent sulfate reduction in anoxic microcosms of paddy soils from Italy and the Philippines, combining 16S rRNA and dissimilatory sulfite reductase (dsrB) gene profiling and co-occurrence network analysis. Sulfate was stoichiometrically reduced in treatments with propionate addition, while CH production was partially suppressed. Methane production but not sulfate reduction were suppressed and acetate accumulated after addition of methyl fluoride or fluoroacetate. With methyl fluoride in the presence of sulfate, the accumulated acetate was consumed after the depletion of propionate. Simultaneously, the relative abundances of Syntrophobacteraceae and Desulfovibrionaceae were significantly enhanced, while fluoroacetate repressed Desulfobulbaceae in both soils. Syntrophobacter 16S rRNA and dsrB gene copy numbers were also remarkably increased with gypsum amendment. Network analysis of both 16S rRNA and dsrB genes illustrated a strong co-occurrence of operational taxonomic units belonging to Syntrophobacteraceae, Desulfovibrionaceae and Desulfobulbaceae. In summary, Syntrophobacteraceae affiliated species were identified as the major propionate-dependent sulfate reducers in paddy soil. They (together with Desulfobulbaceae) oxidized propionate directly to acetate and CO , or coupled the oxidation syntrophically to H /formate-utilizing Desulfovibrionaceae. The transiently accumulating acetate was preferentially consumed by acetoclastic Methanosarcinaceae.

To investigate the effect of ultrasonic treatment on the properties of sweet potato starch and sweet potato starch-based films, the complexing index, thermograms and diffractograms of the sweet potato starch-lauric acid composite were tested, and light transmission, microstructure, and mechanical and moisture barrier properties of the films were measured. The results indicated that the low power density ultrasound was beneficial to the formation of an inclusion complex. In thermograms, the gelatinization enthalpies of the ultrasonically treated starches were lower than those of the untreated sample. With the ultrasonic amplitude increased from 40% to 70%, the melting enthalpy (ΔH) of the inclusion complex gradually decreased. X-ray diffraction revealed that the diffraction intensity of the untreated samples was weaker than that of the ultrasonically treated samples. When the ultrasonic amplitude was above 40%, the diffraction intensity and relative crystallinity of inclusion complex gradually decreased. The scanning electronic microscope showed that the surface of the composite films became smooth after being treated by ultrasonication. Ultrasonication led to a reduction in film surface roughness under atomic force microscopy analysis. The films with ultrasonic treatment exhibited higher light transmission, lower elongation at break, higher tensile strength and better moisture barrier property than those without ultrasonic treatment.

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