Application of a real-time qPCR method to measure the methanogen concentration during anaerobic digestion as an indicator of biogas production capacity

Traversi, Deborah; Villa, Silvia; Lorenzi, Eugenio; Degan, Raffaella; Gilli, Giorgio

doi:10.1016/j.jenvman.2012.07.021

Cited by 76 publications

(38 citation statements)

References 35 publications

(34 reference statements)

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“…). This finding complements previous studies which linked mcrA gene copy number to methane flux (Traversi et al ., ). However, the qRT‐PCR results did not demonstrate a correlation between mcrA transcripts and SMA values.…”

Section: Discussionmentioning

confidence: 97%

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H₂/CO₂‐enriched anaerobic biomass

Morris

Schauer‐Gimenez

Bhattad

et al. 2013

Microbial Biotechnology

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Biologically produced methane (CH4) from anaerobic digesters is a renewable alternative to fossil fuels, but digester failure can be a serious problem. Monitoring the microbial community within the digester could provide valuable information about process stability because this technology is dependent upon the metabolic processes of microorganisms. A healthy methanogenic community is critical for digester function and CH4 production. Methanogens can be surveyed and monitored using genes and transcripts of mcrA, which encodes the α subunit of methyl coenzyme M reductase – the enzyme that catalyses the final step in methanogenesis. Using clone libraries and quantitative polymerase chain reaction, we compared the diversity and abundance of mcrA genes and transcripts in four different methanogenic hydrogen/CO2 enrichment cultures to function, as measured by specific methanogenic activity (SMA) assays using H2/CO2. The mcrA gene copy number significantly correlated with CH4 production rates using H2/CO2, while correlations between mcrA transcript number and SMA were not significant. The DNA and cDNA clone libraries from all enrichments were distinctive but community diversity also did not correlate with SMA. Although hydrogenotrophic methanogens dominated these enrichments, the results indicate that this methodology should be applicable to monitoring other methanogenic communities in anaerobic digesters. Ultimately, this could lead to the engineering of digester microbial communities to produce more CH4 for use as renewable fuel.

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Section: Discussionmentioning

confidence: 97%

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H₂/CO₂‐enriched anaerobic biomass

Morris

Schauer‐Gimenez

Bhattad

et al. 2013

Microbial Biotechnology

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“…Using qPCR with broad-specificity mcrA primers, the mcrA gene copy number has been found to correlate with methanogen abundance in sludge from anaerobic digesters and in methane seep sediments (61) and with methane production rates in a biogas reactor (17). However, gene copy number is not a reliable predictor of metabolic activity and is unable to capture the real-time responses of methanogens to changing operating conditions.…”

Section: Discussionmentioning

confidence: 99%

“…mcrA encodes the enzyme catalyzing the terminal step in methanogenesis and is ubiquitous among known methanogens (15). Methanogen phylogenies determined using the mcrA and 16S rRNA genes are largely congruent (8,15), and both genes have been used to elucidate the diversity and phylogeny of methanogens in anaerobic digesters (14)(15)(16)(17). Methanogens in mixed communities have traditionally been investigated either by cultivation-dependent methods (18,19) or Sanger sequencing of 16S rRNA or mcrA gene clone libraries (14,20).…”

mentioning

confidence: 99%

Pyrosequencing of mcrA and Archaeal 16S rRNA Genes Reveals Diversity and Substrate Preferences of Methanogen Communities in Anaerobic Digesters

Wilkins

Lü

Shen

et al. 2015

Appl Environ Microbiol

101

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Methanogenic archaea play a key role in biogas-producing anaerobic digestion and yet remain poorly taxonomically characterized. This is in part due to the limitations of low-throughput Sanger sequencing of a single (16S rRNA) gene, which in the past may have undersampled methanogen diversity. In this study, archaeal communities from three sludge digesters in Hong Kong and one wastewater digester in China were examined using high-throughput pyrosequencing of the methyl coenzyme M reductase (mcrA) and 16S rRNA genes. Methanobacteriales, Methanomicrobiales, and Methanosarcinales were detected in each digester, indicating that both hydrogenotrophic and acetoclastic methanogenesis was occurring. Two sludge digesters had similar community structures, likely due to their similar design and feedstock. Taxonomic classification of the mcrA genes suggested that these digesters were dominated by acetoclastic methanogens, particularly Methanosarcinales, while the other digesters were dominated by hydrogenotrophic Methanomicrobiales. The proposed euryarchaeotal order Methanomassiliicoccales and the uncultured WSA2 group were detected with the 16S rRNA gene, and potential mcrA genes for these groups were identified. 16S rRNA gene sequencing also recovered several crenarchaeotal groups potentially involved in the initial anaerobic digestion processes. Overall, the two genes produced different taxonomic profiles for the digesters, while greater methanogen richness was detected using the mcrA gene, supporting the use of this functional gene as a complement to the 16S rRNA gene to better assess methanogen diversity. A significant positive correlation was detected between methane production and the abundance of mcrA transcripts in digesters treating sludge and wastewater samples, supporting the mcrA gene as a biomarker for methane yield. Methane from anaerobic digestion is an important source of renewable energy that can circumvent the problems of dwindling fossil fuels and of atmospheric carbon dioxide (CO 2 ) emissions due to fossil fuel combustion (1). Anaerobic digestion is becoming a key part of municipal wastewater treatment, as it allows recovery of energy (biogas) from waste streams to offset onsite energy consumption. The anaerobic digestion of heterogeneous organic substrates for methane production is a complex process involving four major sequential phases: hydrolysis, fermentation, acetogenesis, and methanogenesis (2). Methanogens are strictly anaerobic archaea that produce methane from a limited number of substrates, including hydrogen (H 2 ), acetate, and some C 1 compounds (2). Phylogenetically, methanogens belong to the Euryarchaeota with six established (Methanobacteriales, Methanococcales, Methanomicrobiales, Methanocellales, Methanopyrales, and Methanosarcinales) and one proposed (Methanomassiliicoccales) order(s) (3) and at least 31 genera (4). As well as being major functional components of anaerobic digester communities (5, 6), methanogens are found in other anoxic environments such as peatlands (7), landfills...

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“…Firmicutes and Bacteroidetes members possess cellulose and hydrogenase activity (Wirth et al, ), while Actinobacteria produce lignin‐degrading enzymes that break down complex organic materials (Wirth et al, ; Yang et al, ). In addition, Firmicutes and Bacteroidetes participate in the fermentation of the generated products into organic acids, CO 2 and H 2 (Traversi et al, ; Wirth et al, ). Also, Firmicutes can proceed with the consumption of butyrate and various VFAs (Ariesyady et al, ).…”

Section: Discussionmentioning

confidence: 99%

“…Methanogenesis as the last crucial step in anaerobic digestion is where the stability of the process is more susceptible (Traversi et al, ). Our results showed a high representation of Archaea in the microbial community associated to the biodigester and leachate (8%–17%, Table ).…”

Section: Discussionmentioning

confidence: 99%

Biogas‐producing microbial composition of an anaerobic digester and associated bovine residues

et al. 2019

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Influenced by feedstock type and microbial inoculum, different microbial groups must precisely interact for high‐quality biogas yields. As a first approach for optimization, this study aimed to identify through time the biogas‐producing microbial community in a 10‐ton dry anaerobic digester treating cattle manure by denaturing gradient gel electrophoresis (DGGE) and metagenomics. Moreover, the associated bovine residues or feedstocks (leachate, manure, oxidation lagoon water, rumen) were also characterized to determine their contribution. A diverse and dynamic community characterized by Bacteria (82%–88%) and a considerable amount of Archaea (8%–15%) presented profiles particular to each stage of biogas production. Eukaryotes (2.6%–3.6%), mainly fungi, were a minor but stable component. Proteobacteria represented 47% of the community at the start of the run but only 18% at the end, opposite to the Bacteroidetes/Chlorobi group (8% and 20%, respectively), while Firmicutes (12%–18%) and Actinobacteria (12%–32%) remained relatively constant. Methanogens of the order Methanomicrobiales represented by several species of Methanoculleus were abundant at the end of the run (77%) contrary to Methanosarcinales (11%) and Methanobacteriales (0.7%). Therefore, methanogenesis mainly occurred by the hydrogenotrophic pathway. Manure and oxidation lagoon water seemed to contribute key microorganisms, while rumen dominated by Methanobrevibacter (72%) did not proliferate in the digester. Manure particularly possessed Methanoculleus (24%) and uncultured methanogens identified by DGGE, whereas oxidation lagoon was exclusively abundant in Methanolinea (18%) and Methanosaeta (19%). Leachate, as the microbial inoculum from a previous run, adequately preserved the biogas‐producing community. These results could lead to higher biogas yields through bioaugmentation strategies by incorporating higher proportions or an enriched inoculum from the relevant feedstocks.

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Application of a real-time qPCR method to measure the methanogen concentration during anaerobic digestion as an indicator of biogas production capacity

Cited by 76 publications

References 35 publications

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H₂/CO₂‐enriched anaerobic biomass

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H₂/CO₂‐enriched anaerobic biomass

Pyrosequencing of mcrA and Archaeal 16S rRNA Genes Reveals Diversity and Substrate Preferences of Methanogen Communities in Anaerobic Digesters

Biogas‐producing microbial composition of an anaerobic digester and associated bovine residues

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Application of a real-time qPCR method to measure the methanogen concentration during anaerobic digestion as an indicator of biogas production capacity

Cited by 76 publications

References 35 publications

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H2/CO2‐enriched anaerobic biomass

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H2/CO2‐enriched anaerobic biomass

Pyrosequencing of mcrA and Archaeal 16S rRNA Genes Reveals Diversity and Substrate Preferences of Methanogen Communities in Anaerobic Digesters

Biogas‐producing microbial composition of an anaerobic digester and associated bovine residues

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Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H₂/CO₂‐enriched anaerobic biomass

Methyl coenzyme M reductase (mcrA) gene abundance correlates with activity measurements of methanogenic H₂/CO₂‐enriched anaerobic biomass