Proteomic Changes in Methicillin-Resistant Staphylococcus aureus Exposed to Cannabinoids.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major human pathogen that causes a wide range of infections. Its resistance to ß-lactam antibiotics complicates treatment due to the limited number of antibiotics with activity against MRSA. To investigate development of alternative therapeutics, the mechanisms that mediate antibiotic resistance in MRSA need to be fully understood. In this study, MRSA cells were subjected to antibiotic stress from methicillin in combination with three cannabinoid compounds and analyzed using proteomics to assess the changes in physiology. Subjecting MRSA to nonlethal levels of methicillin resulted in an increased production of penicillin-binding protein 2 (PBP2). Exposure to cannabinoids showed antibiotic activity against MRSA, and differential proteomics revealed reduced levels of proteins involved in the energy production as well as PBP2 when used in combination with methicillin.

Impact of water quality parameters on geosmin levels and geosmin producers in European recirculating aquaculture systems.

AIMS: Geosmin is associated with off-flavour problems in recirculating aquaculture systems (RAS) and represents an economic problem for the aquaculture industry. This study aims at investigating factors influencing the composition of the bacterial microbiota, in particular the presence of geosmin producers and the environmental and farming factors favouring geosmin accumulation. METHODS AND RESULTS: Several water quality parameters were correlated to the composition of the microbiota with special emphasis on the presence of geosmin producers within 26 different RAS from four European countries. Three novel groups of geosmin-producing bacteria were quantified to identify potential correlations with geosmin concentration. CONCLUSIONS: The microbiome differed significantly between systems. However, phosphate levels, calcium levels and redox potential correlated to geosmin concentration in the water and the presence of the Actinomycetales geosmin-producers but not with the presence of other groups of geosmin-producing bacteria. Oxygen levels and conductivity were found to negatively correlate with geosmin concentration. A large proportion of the detected geosmin producers represented novel taxonomic groups not previously linked with this activity. SIGNIFICANCE AND IMPACT OF THE STUDY: These results improve our understanding of the diversity of microbiota in RAS and the water quality parameters favouring the populations of geosmin-producing bacteria and the production of geosmin.

A cohort study of cucumber greenhouse workers' exposure to microorganisms as measured using NGS and MALDI-TOF MS and biomarkers of systemic inflammation.

Work in greenhouses entails exposure to airborne fungi and bacteria. The aims of this study are to obtain knowledge about whether exposure to fungal and bacterial genera and species during work in a cucumber greenhouse is affected by work tasks, and whether a cohort of greenhouse workers' serum levels of serum amyloid A (SAA) and C-reactive protein (CRP), biomarkers of systemic inflammation, are associated with this. Data on personal exposure to airborne fungal and bacterial species measured over 4 years as well as serum levels of SAA and CRP sampled over two years were analyzed. For data analysis, the main work tasks were grouped into three different groups, called 'grouped work task'. Microorganisms were identified using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF MS) and next-generation sequencing (NGS). The 'daily exposure' of greenhouse workers' were as follows: 4.8 × 104 CFU bacteria/m3, 1.4 × 106 CFU fungi/m3, and 392 EU/m3 of endotoxin. Workers were exposed to many different microbial species including several species within the genera Acinetobacter, Bacillus, Microbacterium, Pseudomonas, Staphylococcus, and Streptomyces. The genera Ralstonia and Cladosporium were found in most samples. The exposure levels as well as the microbial composition were associated significantly with grouped work task and season with high exposures during tasks in close contact with mature and old plants and in the autumn. CRP and SAA levels were also associated with exposure level and grouped work tasks. The Shannon-Wiener indices were not different in the 3 'grouped work tasks'. Several specific species including e.g. Halomonas elongata, Stenotrophomonas maltophilia, Podosphaera fusca, and Wallemia spp. were found frequently or in high concentrations in the exposures associated with the highest levels of CRP and SAA. The microorganisms S. maltophilia, P. fusca, and Wallemia spp. were also found on the cucumber plant leaves. In conclusion, both exposure level and the species composition seem to have an effect on the serum levels of CRP and SAA of exposed workers. The greenhouse workers were exposed to only a few species characterized as human pathogens.

Integrated genome-wide investigations of the housefly, a global vector of diseases reveal unique dispersal patterns and bacterial communities across farms.

BACKGROUND: Houseflies (Musca domestica L.) live in intimate association with numerous microorganisms and is a vector of human pathogens. In temperate areas, houseflies will overwinter in environments constructed by humans and recolonize surrounding areas in early summer. However, the dispersal patterns and associated bacteria across season and location are unclear. We used genotyping-by-sequencing (GBS) for the simultaneous identification and genotyping of thousands of Single Nucleotide Polymorphisms (SNPs) to establish dispersal patterns of houseflies across farms. Secondly, we used 16S rRNA gene amplicon sequencing to establish the variation and association between bacterial communities and the housefly across farms. RESULTS: Using GBS we identified 18,000 SNPs across 400 individuals sampled within and between 11 dairy farms in Denmark. There was evidence for sub-structuring of Danish housefly populations and with genetic structure that differed across season and sex. Further, there was a strong isolation by distance (IBD) effect, but with large variation suggesting that other hidden geographic barriers are important. Large individual variations were observed in the community structure of the microbiome and it was found to be dependent on location, sex, and collection time. Furthermore, the relative prevalence of putative pathogens was highly dependent on location and collection time. CONCLUSION: We were able to identify SNPs for the determination of the spatiotemporal housefly genetic structure, and to establish the variation and association between bacterial communities and the housefly across farms using novel next-generation sequencing (NGS) techniques. These results are important for disease prevention given the fine-scale population structure and IBD for the housefly, and that individual houseflies carry location specific bacteria including putative pathogens.

Occurrence of Cyanobacteria and microcystins in hydroelectric reservoirs used for fish farming.

Fish farming can have a negative impact on water quality and aquatic organisms due to emerging blooms of Cyanobacteria and the production of cyanotoxins. In this study, the effect of aquaculture in hydroelectric reservoirs in Brazil was evaluated in six fish farms and in upstream and downstream water through analysis of the microbiome, Cyanobacteria and microcystin concentrations. Synechococcus and Microcystis were observed at all six locations, while Limnothrix was also observed abundantly at two locations. An increase in the relative abundance of Cyanobacteria inside the fish farms was observed at two locations, while an increase of Cyanobacteria was observed in downstream at five of the six locations. Microcystins were detected in significant and high values in all locations, with concentrations up to 1.59 µg/L. The trend in microcystin concentrations was mirrored in copy numbers of the mcyE gene (encodes microcystin synthetase) and presence of Microcystis, but not in any of the other observed cyanobacterial groups. In summary, the study shows that aquaculture production influenced the water microbiome inside and downstream the fish farms, and a direct correlation was found between mcyE gene copies, microcystin production and abundance of Microcystis, but not for the total abundance of Cyanobacteria.

A legume genetic framework controls infection of nodules by symbiotic and endophytic bacteria.

Legumes have an intrinsic capacity to accommodate both symbiotic and endophytic bacteria within root nodules. For the symbionts, a complex genetic mechanism that allows mutual recognition and plant infection has emerged from genetic studies under axenic conditions. In contrast, little is known about the mechanisms controlling the endophytic infection. Here we investigate the contribution of both the host and the symbiotic microbe to endophyte infection and development of mixed colonised nodules in Lotus japonicus. We found that infection threads initiated by Mesorhizobium loti, the natural symbiont of Lotus, can selectively guide endophytic bacteria towards nodule primordia, where competent strains multiply and colonise the nodule together with the nitrogen-fixing symbiotic partner. Further co-inoculation studies with the competent coloniser, Rhizobium mesosinicum strain KAW12, show that endophytic nodule infection depends on functional and efficient M. loti-driven Nod factor signalling. KAW12 exopolysaccharide (EPS) enabled endophyte nodule infection whilst compatible M. loti EPS restricted it. Analysis of plant mutants that control different stages of the symbiotic infection showed that both symbiont and endophyte accommodation within nodules is under host genetic control. This demonstrates that when legume plants are exposed to complex communities they selectively regulate access and accommodation of bacteria occupying this specialized environmental niche, the root nodule.

The microbial community of the gut differs between piglets fed sow milk, milk replacer or bovine colostrum.

The aim of this study was to characterise the gut microbiota composition of piglets fed bovine colostrum (BC), milk replacer (MR) or sow milk (SM) in the post-weaning period. Piglets (n 36), 23-d old, were randomly allocated to the three diets. Faecal samples were collected at 23, 25, 27 and 30 d of age. Digesta from the stomach, ileum, caecum and mid-colon was collected at 30 d of age. Bacterial DNA from all samples was subjected to amplicon sequencing of the 16S rRNA gene. Bacterial enumerations by culture and SCFA analysis were conducted as well. BC-piglets had the highest abundance of Lactococcus in the stomach (P<0·0001) and ileal (P<0·0001) digesta, whereas SM-piglets had the highest abundance of Lactobacillus in the stomach digesta (P<0·0001). MR-piglets had a high abundance of Enterobacteriaceae in the ileal digesta (P<0·0001) and a higher number of haemolytic bacteria in ileal (P=0·0002) and mid-colon (P=0·001) digesta than SM-piglets. BC-piglets showed the highest colonic concentration of iso-butyric and iso-valeric acid (P=0·02). Sequencing and culture showed that MR-piglets were colonised by a higher number of Enterobacteriaceae, whereas the gut microbiota of BC-piglets was characterised by a change in lactic acid bacteria genera when compared with SM-piglets. We conclude that especially the ileal microbiota of BC-piglets had a closer resemblance to that of SM-piglets in regard to the abundance of potential enteric pathogens than did MR-piglets. The results indicate that BC may be a useful substitute for regular milk replacers, and as a feeding supplement in the immediate post-weaning period.

Eat seldom is better than eat frequently: Pharmaceuticals degradation kinetics, enantiomeric profiling and microorganisms in moving bed biofilm reactors are affected by feast famine cycle times.

Feast-famine (FF) regimes improved the removal of recalcitrant pharmaceuticals in moving bed biofilm reactors (MBBRs), but the optimal FF cycle remained unresolved. The effects of FF cycle time on the removal of bulk substrates (organic carbon and nitrogen) and trace pharmaceuticals by MBBR are systematically evaluated in this study. The feast to famine ratio was fixed to 1:2 to keep the same loading rate, but the time for the FF cycles varied from 18 h to 288 h. The MBBR adapted to the longest FF cycle time (288 h equaling 48 × HRT) resulted in significantly higher degradation rates (up to +183%) for 12 out of 28 pharmaceuticals than a continuously fed (non-FF) reactor. However, other FF cycle times (18, 36, 72 and 144 h) only showed a significant up-regulation for 2-3 pharmaceuticals compared to the non-FF reactor. Enantioselective degradation of metoprolol and propranolol occurred in the second phase of a two phase degradation, which was different for the longer FF cycle time. N-oxidation and N-demethylation pathways of tramadol and venlafaxine differed across the FF cycle time suggestin the FF cycle time varied the predominant transformation pathways of pharmaceuticals. The abundance of bacteria in the biofilms varied considerably between different FF cycle times, which possibly caused the biofilm to remove more recalcitrant bulk organic C and pharmaceuticals under long cycle times.

Feeding Drosophila gut microbiomes from young and old flies modifies the microbiome.

It is becoming increasingly evident that the myriad of microbes in the gut, within cells and attached to body parts (or roots of plants), play crucial roles for the host. Although this has been known for decades, recent developments in molecular biology allow for expanded insight into the abundance and function of these microbes. Here we used the vinegar fly, Drosophila melanogaster, to investigate fitness measures across the lifetime of flies fed a suspension of gut microbes harvested from young or old flies, respectively. Our hypothesis was that flies constitutively enriched with a 'Young microbiome' would live longer and be more agile at old age (i.e. have increased healthspan) compared to flies enriched with an 'Old microbiome'. Three major take home messages came out of our study: (1) the gut microbiomes of young and old flies differ markedly; (2) feeding flies with Young and Old microbiomes altered the microbiome of recipient flies and (3) the two different microbial diets did not have any effect on locomotor activity nor lifespan of the recipient flies, contradicting our working hypothesis. Combined, these results provide novel insight into the interplay between hosts and their microbiomes and clearly highlight that the phenotypic effects of gut transplants and probiotics can be complex and unpredictable.

Anaerobic digestion of wastewater from hydrothermal liquefaction of sewage sludge and combined wheat straw-manure.

Hydrothermal liquefaction (HTL) shows promise for converting wet biomass waste into biofuel, but the resulting high-strength process water (PW) requires treatment. This study explored enhancing energy recovery by anaerobic digestion using semi-batch reactors. Co-digesting manure with HTL-PW from wheat straw-manure co-HTL yielded methane (43-49% of the chemical oxygen demand, COD) at concentrations up to 17.8 gCOD·L-1, whereas HTL-PW from sewage sludge yielded methane (43% of the COD) up to only 12.8 gCOD·L-1 and complete inhibition occurred at 17 gCOD·L-1. Microbial community shifts confirmed inhibition of methanogenic archaea, while hydrolytic-fermentative bacteria were resilient. Differences in chemical composition, particularly higher levels of N-containing heterocyclic compounds in PW of sewage sludge, likely caused the microbial inhibition. The considerable potential of combining HTL with anaerobic digestion for enhanced energy recovery from straw-manure in an agricultural context is demonstrated, yet sewage sludge HTL-PW requires more advanced approaches to deal with methanogenesis inhibitors.

Revealing the secrets of a 2900-year-old clay brick, discovering a time capsule of ancient DNA.

The recent development of techniques to sequence ancient DNA has provided valuable insights into the civilisations that came before us. However, the full potential of these methods has yet to be realised. We extracted ancient DNA from a recently exposed fracture surface of a clay brick deriving from the palace of king Ashurnasirpal II (883-859 BCE) in Nimrud, Iraq. We detected 34 unique taxonomic groups of plants. With this research we have made the pioneering discovery that ancient DNA, effectively protected from contamination inside a mass of clay, can successfully be extracted from a 2900-year-old clay brick. We encourage future research into this subject, as the scientific prospects for this approach are substantial, potentially leading to a deeper understanding of ancient and lost civilisations.

Unravelling gradient layers of microbial communities, proteins, and chemical structure in aerobic granules.

Most bacteria live in microbial assemblages like biofilms and granules, and each layer of these assemblages provides a niche for certain bacteria with specific metabolic functions. In this study, a gentle (non-destructive) extraction approach based on a cation exchange resin and defined shear was employed to gradually disintegrate biomass and collect single layers of aerobic granules from a full-scale municipal wastewater treatment plant. The microbial community composition of granule layers was characterized using next-generation sequencing (NGS) targeting the 16S rRNA gene, and protein composition was investigated using metaproteomics. The chemical composition of eroded layers was explored using Fourier Transformed Infrared Spectroscopy. On the surface of the granules, the microbial structure (flocculation-supporting Nannocystis sp.) as well as composition of extracellular polymers (extracellular DNA) and proteome (chaperonins and binding proteins) favored microbial aggregation. Extracellular polymeric substances in the granules were composed of mostly proteins and EPS-producers, such as Tetrasphaera sp. and Zoogloea sp., were evenly distributed throughout the granule structure. The interior of the granules harbored several denitrifiers (e.g., Thauera sp.), phosphate-accumulating denitrifiers (Candidatus Accumulibacter, Dechloromonas sp.) and nitrifiers (Candidatus Nitrotoga). Proteins associated with glycolytic activity were identified in the outer and middle granule layers, and proteins associated with phosphorus conversions, in the deeper layers. In conclusion, the use of an existing cation-exchange resin for gradual biomass disintegration, combined with NGS and metaproteomic analysis was demonstrated as a promising approach for simultaneously investigating the identity and functions of microbes in multilayered biofilm structures.

The Gut Microbiome of 54 Mammalian Species.

The gut microbiome plays a critical role in many aspects of host life, and the microbial community composition is heavily influenced by the prevailing conditions in the gut environment. Community composition has been suggested to have large implications for conservation efforts, and gut health has become of interest for optimizing animal care in captivity. In this study, we explore the gut microbiome of a wide range of animals in the context of conservation biology. The composition of the gut microbial community of 54 mammalian animal species was investigated using 16S rRNA gene amplicon sequencing. The composition of the gut microbiota clearly reflects diet and the structure of the gastrointestinal system, and it is to a certain degree more similar between closely related animals. Specific clusters of taxa were observed across animals of the same species, diet, and gut morphology. The microbiota retained regardless of captivity status is hypothesized to cover important symbiotic relationships with the host, while the remaining part reflects the artificial living conditions and can therefore be used as a future tool for conservation biologists. For five animal species (giraffes, horses, baboons, elephants, and zebras), it was possible to compare the microbiota of wild and captive individuals. Differences were observed in the proportion of microbiota detected between wild and captive specimens of the same animal species. We propose that the gut microbiota harbours important species, which can potentially serve as indicators for the well-being of the animal and the effect of living in captivity.

Wood-Ljungdahl pathway utilisation during in situ H2 biomethanation.

Biogas production from organic waste is a waste-to-energy technology with the potential to contribute significantly to sustainable energy production. Upgrading of biogas using in situ biomethanation with hydrogen has the potential for surplus electricity storage, and delivery of biogas with a methane content of >90%, allowing for easier integration into the natural gas grid, as well as conversion to other products. Microbial communities in biomethanation reactors undergo changes, however, these changes are largely unexplored. In the present study, metagenome-resolved protein stable isotope probing (Protein-SIP) was applied to laboratory scale batch incubations operating under anaerobic digestion, and (pre-adapted) biomethanation conditions, fed with 13C-labelled bicarbonate, in order to gain insight into the microbial activities during CO2-reduction. The strongest and most microbially diverse isotopic incorporation was observed in the pre-adapted biomethanation incubation. Furthermore, divergent incorporation of 13C-labelled bicarbonate was also observed in the Wood-Ljungdahl pathway, with the anaerobic digester incubations primarily showing labelled proteins in the peripheral pathways leading toward production of energy and biomass. The pre-adapted biomethanation incubations consumed H2 and CO2, but did not convert it to CH4, suggesting the production of acetate in these incubations, which was supported by heavy labelling of key enzymes in the Wood-Ljungdahl pathway. Twelve (ten high quality) metagenome-assembled genomes (MAGs) coding for 13C-incorporated proteins were extracted from the metagenome, eight of which contained one or more of the key genes in the Wood-Ljungdahl pathway, one of which was affiliated to Methanosarcina. Together, the findings in the present study deepen our knowledge surrounding microbial communities in biomethanation systems, and contribute to the development of better strategies for implementation of biogas upgrading and microbial management.

Characterisation of cellulose-degrading organisms in an anaerobic digester.

The recalcitrant nature of lignocellulosic biomass hinders efficient exploitation of this fraction for energy production. A better understanding of the microorganisms able to convert plant-based feedstocks is needed to improve anaerobic digestion of lignocellulosic biomass. In this study, active thermophilic cellulose-degrading microorganisms were identified from a full-scale anaerobic digester fed with maize by using metagenome-resolved protein stable isotope probing (protein-SIP). 13C-cellulose was converted into 13C-methane with a 13/12C isotope ratio of 0.127 after two days of incubation. Metagenomic analysis revealed 238 different genes coding for carbohydrate-active enzymes (CAZymes), six of which were directly associated with cellulose degradation. The protein-SIP analysis identified twenty heavily labelled peptides deriving from microorganisms actively assimilating labelled carbon from the degradation of 13C-cellulose, highlighting several members of the order Clostridiales. Corynebacterium was identified through CAZyme screening, amplicon analysis, and in the metagenome giving a strong identification of being a cellulose degrader.

Sulfate-reducing and methanogenic microbial community responses during anaerobic digestion of tannery effluent.

Microbial communities were monitored in terms of structure, function and response to physicochemical variables during anaerobic digestion of tannery and associated slaughterhouse effluent in: (i) 2 L biochemical methane potential batch reactors at different inoculum to substrate ratios (2-5) and initial sulfate concentrations (665-2000 mg/L), and (ii) 20 L anaerobic sequencing batch reactors with different mixing regimes (continuous vs. intermittent). Methanogenic and sulfidogenic community compositions in the 2 L reactors evolved initially, but stabilised after the start of biogas generation, although significant (ANOSIM p < 0.05) changes in the physicochemical parameters indicated continued metabolic activity. Both hydrogenotrophic and acetoclastic archaeal genera were present in high relative abundances. Continuous stirring preferentially selected the metabolically versatile genus Methanosarcina, suggesting that higher specific methane generation in the continuously stirred system (168 vs. 19.5 mL methane per gram volatile solids per week) was related to the metabolic activities of members of this genus.

Food origin influences microbiota and stable isotope enrichment profiles of cold-adapted Collembola (Desoria ruseki).

Collembola are a group of globally distributed microarthropods that can tolerate low temperature and are active in extremely cold environments. While it is well known that animal diets can shape their microbiota, the microbiota of soil animals is not well described, particularly for animals with limited food resources, such as Collembola active in winter at low temperatures. In this study, we explored the effects of three different food sources; corn litter (agriculture grain residuals), Mongolian oak litter (natural plant residuals), and yeast (common food for Collembola culture), on the microbiota of a winter-active Collembola species, Desoria ruseki. We found that microbial diversity and community composition of the Collembola were strongly altered after feeding with different food sources for 30 days. Collembola individuals fed on corn litter harbored the highest bacterial richness and were dominated by a representative of Microbacteriaceae. In contrast, those fed on yeast exhibited the lowest bacterial richness and were primarily colonized by Pseudomonas. The microbial communities associated with the winter-active Collembola differed significantly from those observed in the food. Collembola nutrient turnover also differed when cultured with different food sources, as indicated by the C and N stable isotopic signatures. Our study highlights microbial associations with stable isotopic enrichments of the host. Specifically, the Arthrobacter was positively correlated with δ13C enrichment in the host. Representatives of Microbacteriaceae, Micrococcaceae, TM7a, Devosia, and Rathayibacter were positively correlated with δ15N enrichment of the host. Our study indicates that food sources are major determinants for Collembola microbiota that simultaneously alter consumers' isotopic niches, thereby improving our understanding of the roles played by host-microbiota interactions in sustaining soil biodiversity during the winter.

Mastication of polyolefins alters the microbial composition in Galleria mellonella.

Recent studies have indicated that Galleria mellonella larvae ingest polyethylene films and the degradation mechanism could inspire biotechnological exploitation for degrading plastic to eliminate global pollution from plastic waste. In this study, we tested the chemical compositions of masticated and ingested different plastic types by G. mellonella. High throughput sequencing of 16S rRNA gene was used to characterize the alteration of the microbial communities derived from salivary glands, gut contents and whole G. mellonella larvae. Our results indicated that G. mellonella is able to masticate polyethylene (PE), expanded polystyrene (EPS) and polypropylene (PP) and convert it to small particles with very large and chemically modified surfaces. The characteristics of the polymer affect the rate of damage. Formation of functional carbonyl groups on the appearance of oxidized metabolic intermediates of polyolefins in the frass samples observed. We found that the mastication of EPS, PP or PE could significantly alter the microbial composition in the gut content while it did not appear to influence the salivary glands microbial community. Representatives of Desulfovibrio vulgaris and Enterobacter grew with the PE diet while mastication of polystyrene and polypropylene increased the abundance of Enterococcus. The evaluation of bacterial communities in whole larvae confirmed the obtained result and additionally showed that the abundance of Paenibacillus, Corynebacterium and Commamonadaceae increased by Styrofoam (EPS) consumption.

Cellulolytic and Xylanolytic Microbial Communities Associated With Lignocellulose-Rich Wheat Straw Degradation in Anaerobic Digestion.

The enzymatic hydrolysis of lignocellulosic polymers is generally considered the rate-limiting step to methane production in anaerobic digestion of lignocellulosic biomass. The present study aimed to investigate how the hydrolytic microbial communities of three different types of anaerobic digesters adapted to lignocellulose-rich wheat straw in continuous stirred tank reactors operated for 134 days. Cellulase and xylanase activities were monitored weekly using fluorescently-labeled model substrates and the enzymatic profiles were correlated with changes in microbial community compositions based on 16S rRNA gene amplicon sequencing to identify key species involved in lignocellulose degradation. The enzymatic activity profiles and microbial community changes revealed reactor-specific adaption of phylogenetically different hydrolytic communities. The enzymatic activities correlated significantly with changes in specific taxonomic groups, including representatives of Ruminiclostridium, Caldicoprobacter, Ruminofilibacter, Ruminococcaceae, Treponema, and Clostridia order MBA03, all of which have been linked to cellulolytic and xylanolytic activity in the literature. By identifying microorganisms with similar development as the cellulase and xylanase activities, the proposed correlation method constitutes a promising approach for deciphering essential cellulolytic and xylanolytic microbial groups for anaerobic digestion of lignocellulosic biomass.

Methanogenic archaea use a bacteria-like methyltransferase system to demethoxylate aromatic compounds.

Methane-generating archaea drive the final step in anaerobic organic compound mineralization and dictate the carbon flow of Earth's diverse anoxic ecosystems in the absence of inorganic electron acceptors. Although such Archaea were presumed to be restricted to life on simple compounds like hydrogen (H2), acetate or methanol, an archaeon, Methermicoccus shengliensis, was recently found to convert methoxylated aromatic compounds to methane. Methoxylated aromatic compounds are important components of lignin and coal, and are present in most subsurface sediments. Despite the novelty of such a methoxydotrophic archaeon its metabolism has not yet been explored. In this study, transcriptomics and proteomics reveal that under methoxydotrophic growth M. shengliensis expresses an O-demethylation/methyltransferase system related to the one used by acetogenic bacteria. Enzymatic assays provide evidence for a two step-mechanisms in which the methyl-group from the methoxy compound is (1) transferred on cobalamin and (2) further transferred on the C1-carrier tetrahydromethanopterin, a mechanism distinct from conventional methanogenic methyl-transfer systems which use coenzyme M as final acceptor. We further hypothesize that this likely leads to an atypical use of the methanogenesis pathway that derives cellular energy from methyl transfer (Mtr) rather than electron transfer (F420H2 re-oxidation) as found for methylotrophic methanogenesis.