Fractionation of stable carbon isotopes during acetate consumption by methanogenic and sulfidogenic microbial communities in rice paddy soils and lake sediments

Acetate is an important intermediate during the degradation of organic matter in anoxic flooded soils and sediments. Acetate is disproportionated to CH4 and CO2 by methanogenic or is oxidized to CO2 by sulfate-reducing microorganisms. These reactions result in carbon isotope fractionation, depending on the microbial species and their particular carbon metabolism. To learn more about the magnitude of the isotopic enrichment factors (ε) involved, acetate conversion to CH4 and CO2 was measured in anoxic paddy soils from Vercelli (Italy) and the International Rice Research Institute (IRRI, the Philippines) and in anoxic lake sediments from the northeastern and the southwestern basins of Lake Fuchskuhle (Germany). Acetate consumption was measured using samples of paddy soil or lake sediment suspended in water or in phosphate buffer (pH 7.0), both in the absence and presence of sulfate (gypsum), and of methyl fluoride (CH3F), an inhibitor of aceticlastic methanogenesis. Under methanogenic conditions, values of εac for acetate consumption were always in a range of −21 ‰ to −17 ‰ but higher in the lake sediment from the southwestern basin (−11 ‰). Under sulfidogenic conditions εac values tended to be slightly lower (−26 ‰ to −19 ‰), especially when aceticlastic methanogenesis was inhibited. Again, εac in the lake sediment of the southwestern basin was higher (−18 ‰ to −14 ‰). Determination of εCH4 from the accumulation of 13C in CH4 resulted in much lower values (−37 ‰ to −27 ‰) than from the depletion of 13C in acetate (−21 ‰ to −17 ‰ ), especially when acetate degradation was measured in buffer suspensions. The microbial communities were characterized by sequencing the bacterial 16S rRNA (ribosomal ribonucleic acid) genes as well as the methanogenic mcrA and sulfidogenic dsrB genes. The microbial communities were quite different between lake sediments and paddy soils but were similar in the sediments of the two lake basins and in the soils from Vercelli and the IRRI, and they were similar after preincubation without and with addition of sulfate (gypsum). The different microbial compositions could hardly serve for the prediction of the magnitude of enrichment factors.

Fractionation of stable carbon isotopes during acetate consumption by methanogenic and sulfidogenic microbial communities in rice paddy soils and lake sediments

Acetate is an important intermediate during the degradation of organic matter in anoxic flooded soils and sediments. Acetate is disproportionated to CH4 and CO2 by methanogenic or is oxidized to CO2 by sulfate-reducing microorganisms. These reactions result in carbon isotope fractionation, depending on the microbial species and their particular carbon metabolism. To learn more about the magnitude of the isotopic enrichment factors (ε) involved, acetate conversion to CH4 and CO2 was measured in anoxic paddy soils from Vercelli (Italy) and the International Rice Research Institute (IRRI, the Philippines) and in anoxic lake sediments from the north east (NE) and the south west (SW) basins of Fuchskuhle (Germany). Acetate consumption was measured using samples of paddy soil or lake sediment suspended in water or in phosphate buffer (pH 7.0), both in the absence and presence of sulfate (gypsum), and of methyl fluoride (CH3F), an inhibitor of aceticlastic methanogenesis. Under methanogenic conditions, values of εac for acetate consumption were always in a range of -21 ‰ to -17 ‰, but higher in the lake sediment from the SW basin (-11 ‰). Under sulfidogenic conditions εac values tended to be slightly lower (-26 ‰ to -19 ‰) especially when aceticlastic methanogenesis was inhibited. Again, εac in the lake sediment of the SW basin was higher (-18 ‰ to -14 ‰). Determination of εCH4 from the accumulation of 13C in CH4 resulted in much lower values (-37 ‰ to -27 ‰) than from the depletion of 13C in acetate (-21 ‰ to -17 ‰), especially when acetate degradation was measured in buffer suspensions. The microbial communities were characterized by sequencing the bacterial 16S rRNA genes as well as the methanogenic mcrA and sulfidogenic dsrB genes. The microbial communities were quite different between lake sediments and paddy soils, but were similar in the sediments of the two lake basins and in the soils from Vercelli and IRR, and were similar after preincubation without and with addition of sulfate (gypsum). The different microbial compositions could hardly serve for the prediction of the magnitude of enrichment factors.

Biogas and volatile fatty acid production during anaerobic digestion of straw, cellulose, and hemicellulose with analysis of microbial communities and functions

Background Anaerobic digestion (AD) is a promising method for straw treatment, but the complex composition and structure of straw limit AD efficiency and methane production. The main biodegradable components of straw are cellulose and hemicellulose. Because of the different chemical structures and physicochemical properties, the performance of AD of cellulose and hemicellulose is different, thus it’s also different from that of straw. Research on the similarities and differences of AD of straw, cellulose and hemicellulose is helpful to clarify the law of anaerobic digestion of straw and provide theoretical basis for further improving the efficiency of anaerobic digestion. However, there are very few studies on AD using cellulose and hemicellulose as raw materials. Results Rice straw (RS), cellulose, and hemicellulose were used as raw materials to study biogas production performance and changes in the volatile fatty acids (VFAs). Further, microbial communities and genetic functions were analyzed separately for each material. The biogas production potential of RS, cellulose, and hemicellulose was different, with cumulative biogas production of 620.64, 412.50, and 283.75 mL/g·VS− 1, respectively. The methane content of the biogas produced from cellulose and hemicellulose was approximately 10% higher than that produced from RS after the methane content stabilized. Biogas production and the methane content of RS stabilized more quickly than that of cellulose and hemicellulose. The accumulation of VFAs occurred in the early stage of anaerobic digestion in all the three materials, and the main volatile fatty acid component of RS was acetic acid, whereas that of cellulose and hemicellulose was propionic acid. The cumulative amount of VFAs in both cellulose and hemicellulose was relatively higher than that in RS, and the accumulation time was 12 and 14 days longer, respectively. When anaerobic digestion progressed to a stable stage, Clostridium was the dominant bacterial genus in all three AD systems, and the abundance of Ruminofilibacter was higher during anaerobic digestion of RS. Genetically, AD of all the three materials proceeded mainly via aceticlastic methanogenesis, with similar functional components. Conclusion The biogas and VFAs production during AD of RS, cellulose, and hemicellulose showed marked differences. But when the AD progressed to the stable stage, there was no significant difference in microbial community and genetic function. Specifically, the biogas production potential of cellulose and hemicellulose was greater than that of RS. The accumulation of VFAs in the three AD systems occurred in the early stages. The main component of VFA that accumulated in RS was acetic acid, while the major component of VFAs accumulated in cellulose and hemicellulose digestions was propionic acid. At the stable stage, Clostridium was the dominant bacterial genus in all three AD systems. The AD of all the three materials proceeded mainly via aceticlastic methanogenesis, with similar components of gene functions.

Acetate turnover and methanogenic pathways in Amazonian lake sediments

Lake sediments in Amazonia are a significant source of CH4, a potential greenhouse gas. Previous studies of sediments using 13C analysis found that the contribution of hydrogenotrophic versus aceticlastic methanogenesis to CH4 production was relatively high. Here, we determined the methanogenic pathway in the same sediments (n = 6) by applying [14C]bicarbonate or [2-14C]acetate, and confirmed the high relative contribution (50–80 %) of hydrogenotrophic methanogenesis. The respiratory index (RI) of [2-14C]acetate, which is 14CO2 relative to 14CH4 + 14CO2, divided the sediments into two categories, i.e., those with an RI 0.4 showing that a large percentage of the acetate-methyl was oxidized to CO2 rather than reduced to CH4. Hence, part of the acetate was probably converted to CO2 plus H2 via syntrophic oxidation, thus enhancing hydrogenotrophic methanogenesis. This happened despite the presence of potentially aceticlastic Methanosaetaceae in all the sediments. Alternatively, acetate may have been oxidized with a constituent of the sediment organic matter (humic acid) serving as oxidant. Indeed, apparent acetate turnover rates were larger than CH4 production rates except in those sediments with a R

Mathematical modeling of primary sludge anaerobic hydrolysis

<p>The aim of this work was the application and evaluation of a mathematical model for the simulation of anaerobic hydrolysis and acid production processes. For the description of the processes involved, the Anaerobic Digestion Model ADM1 was employed. The ADM1 implementation in prefermenters is relatively easy, but the estimation of its components concentrations, kinetic parameters and stoichiometric coefficients, remains a problem to be solved. This study provides useful results for the ADM1 implementation in acid digesters design and operation. Model calibration and verification was performed using experimental data from two bench scale acid digesters operating at different temperatures (12, 20, 27 and 34 °C) and retention times (1,2, 4 and 6 days). Model sensitivity analysis illustrated that the values of pH, disintegration kinetic parameter and aceticlastic methanogenesis specific rate, as well as biomass and inert soluble COD concentrations in the primary sludge exert a significant influence on soluble COD production. Model parameters (disintegration and hydrolysis kinetic parameters and specific rate of methane production) as well as the temperature dependency of these parameters are given in this paper.</p>

Metabolic marker gene mining provides insight in globalmcrAdiversity and, coupled with targeted genome reconstruction, sheds further light on metabolic potential of theMethanomassiliicoccales

Over the past years, metagenomics has revolutionized our view of microbial diversity. Moreover, extracting near-complete genomes from metagenomes has led to the discovery of known metabolic traits in unsuspected lineages. Genome-resolved metagenomics relies on assembly of the sequencing reads and subsequent binning of assembled contigs, which might be hampered by strain heterogeneity or low abundance of a target organism. Here we present a complementary approach, metagenome marker gene mining, and use it to assess the global diversity of archaeal methane metabolism through themcrAgene. To this end, we have screened 18,465 metagenomes for the presence of reads matching a database representative of all known mcrA proteins and reconstructed gene sequences from the matching reads. We use our mcrA dataset to assess the environmental distribution of theMethanomassiliicoccalesand reconstruct and analyze a draft genome belonging to the ‘Lake Pavin cluster’, an uncultivated environmental clade of theMethanomassiliicoccales. Analysis of the ‘Lake Pavin cluster’ draft genome suggests that this organism has a more restricted capacity for hydrogenotrophic methylotrophic methanogenesis than previously studiedMethanomassiliicoccales, with only genes for growth on methanol present. However, the presence of the soluble subunits of methyltetrahydromethanopterin:CoM methyltransferase (mtrAH)provide hypothetical pathways for methanol fermentation, and aceticlastic methanogenesis that await experimental verification. Thus, we show that marker gene mining can enhance the discovery power of metagenomics, by identifying novel lineages and aiding selection of targets for in-depth analyses. Marker gene mining is less sensitive to strain heterogeneity and has a lower abundance threshold than genome-resolved metagenomics, as it only requires short contigs and there is no binning step. Additionally, it is computationally cheaper than genome resolved metagenomics, since only a small subset of reads needs to be assembled. It is therefore a suitable approach to extract knowledge from the many publicly available sequencing projects.

Transcriptomic and Physiological Insights into the Robustness of Long Filamentous Cells of Methanosaeta harundinacea, Prevalent in Upflow Anaerobic Sludge Blanket Granules

ABSTRACTMethanosaetaspp. are widely distributed in natural environments, and their filamentous cells contribute significantly to sludge granulation and the good performance of anaerobic reactors. A previous study indicated thatMethanosaeta harundinacea6Ac displays a quorum sensing-regulated morphological transition from short to long filaments, and more acetate is channeled into methane production in long filaments, whereas more is channeled into biomass synthesis in short filaments. Here, we performed transcriptomic and physiological analysis to gain insights into active methanogenesis in long filaments ofM. harundinacea6Ac. Both RNA sequencing (RNA-seq) and quantitative reverse transcription-PCR indicated that transcription of the genes involved in aceticlastic methanogenesis and energy metabolism was upregulated 1.2- to 10.3-fold in long filaments, while transcription of the genes for the methyl oxidative shunt was upregulated in short filaments. [2-13C]acetate trace experiments demonstrated that a relatively higher portion of the acetate methyl group was oxidized to CO2in short filaments than in long filaments. The long filaments exhibited higher catalase activity and oxygen tolerance than the short ones, which is consistent with increased transcription of the oxidant-scavenging genes. Moreover, transcription of genes for cell surface structures was upregulated in the long filaments, and transmission electron microscopy revealed a thicker cell envelope in the filaments. RNA-seq determined a >2-fold upregulation of a variety of antistress genes in short filaments, like those encoding chaperones and DNA repair systems, which implies that the short filaments can be stressed. This study reveals the genetic basis for the prevalence of the long filamentous morphology ofM. harundinaceacells in upflow anaerobic sludge blanket granules.