Proteotyping of biogas plant microbiomes separates biogas plants according to process temperature and reactor type.

BACKGROUND: Methane yield and biogas productivity of biogas plants (BGPs) depend on microbial community structure and function, substrate supply, and general biogas process parameters. So far, however, relatively little is known about correlations between microbial community function and process parameters. To close this knowledge gap, microbial communities of 40 samples from 35 different industrial biogas plants were evaluated by a metaproteomics approach in this study. RESULTS: Liquid chromatography coupled to tandem mass spectrometry (Orbitrap Elite™ Hybrid Ion Trap-Orbitrap Mass Spectrometer) of all 40 samples as triplicate enabled the identification of 3138 different metaproteins belonging to 162 biological processes and 75 different taxonomic orders. The respective database searches were performed against UniProtKB/Swiss-Prot and seven metagenome databases. Subsequent clustering and principal component analysis of these data allowed for the identification of four main clusters associated with mesophile and thermophile process conditions, the use of upflow anaerobic sludge blanket reactors and BGP feeding with sewage sludge. Observations confirm a previous phylogenetic study of the same BGP samples that was based on 16S rRNA gene sequencing by De Vrieze et al. (Water Res 75:312-323, 2015). In particular, we identified similar microbial key players of biogas processes, namely Bacillales, Enterobacteriales, Bacteriodales, Clostridiales, Rhizobiales and Thermoanaerobacteriales as well as Methanobacteriales, Methanosarcinales and Methanococcales. For the elucidation of the main biomass degradation pathways, the most abundant 1 % of metaproteins was assigned to the KEGG map 1200 representing the central carbon metabolism. Additionally, the effect of the process parameters (i) temperature, (ii) organic loading rate (OLR), (iii) total ammonia nitrogen (TAN), and (iv) sludge retention time (SRT) on these pathways was investigated. For example, high TAN correlated with hydrogenotrophic methanogens and bacterial one-carbon metabolism, indicating syntrophic acetate oxidation. CONCLUSIONS: This is the first large-scale metaproteome study of BGPs. Proteotyping of BGPs reveals general correlations between the microbial community structure and its function with process parameters. The monitoring of changes on the level of microbial key functions or even of the microbial community represents a well-directed tool for the identification of process problems and disturbances.Graphical abstractCorrelation between the different orders and process parameter, as well as principle component analysis of all investigated biogas plants based on the identified metaproteins.

Hygienization of sludge through anaerobic digestion at 35, 55 and 60 °C.

Legislation in Sweden and the European Union concerning the use of sewage sludge in agriculture is under revision and future concentration limits for pathogens in treated sludge are likely to be expected. The aim of this study was to evaluate the hygienization of Salmonella, Escherichia coli, Enterococcus and Clostridium perfringens through continuous anaerobic digestion at 35, 55 or 60 °C, as well as to investigate process stability and methane production at 60 °C. The results indicated that digestion at 55 or 60 °C with a minimum exposure time of 2 h resulted in good reduction of Salmonella, E. coli and Enterococcus and that anaerobic digestion could thus be used to reach the concentration limits suggested for the EU, as well as Sweden. Furthermore, stable continuous anaerobic digestion of sludge was achieved at 60 °C, albeit with 10% less methane production compared to digestion at 35 and 55 °C.

Triplet properties and interactions of the primary electron donor and antenna chromophores in membranes of Heliobacterium chlorum, studied with ADMR spectroscopy.

The triplet states of antenna and reaction center bacteriochlorophyll (BChl) g in membranes of Heliobacterium chlorum were studied by optically detected magnetic resonance in zero magnetic field, using absorbance detection. A variety of triplet states was detected, which were all localized on single BChl g chromophores as concluded from a comparison with the triplet state of monomeric BChl g in organic solvents. With the aid of the microwave-induced absorbance difference spectra, we assign a triplet state with zero-field splitting parameters |D| = 727.5 and |E| = 254. 5 MHz to that of the primary donor. The low |E| value indicates that the BChls of the primary donor are monoligated. The intensities of the zero-field transitions were strongly dependent on the redox state of the secondary electron acceptors. A triplet state with |D| = 690-705 MHz and |E| =230 MHz, present under all redox conditions, is associated with antenna BChl g absorbing at 814 nm. Its triplet yield was independent of the redox conditions; we conclude therefore that the antenna chromophores absorbing at 814 nm are not connected with the reaction center at cryogenic temperatures (1.2 K). In addition, relatively strong signals were detected belonging to triplet states with |D| and |E| of 663-680 and 220-227 MHz, respectively, whose amplitudes were dependent on the redox conditions. Triplet states with these zero-field splitting parameters are located on antenna chromophores absorbing between 798-814 nm; their zero-field transitions and absorbance difference spectra indicate a considerable heterogeneity. The concentration of triplet states of antenna chromophores absorbing around 800 nm decreased markedly upon prolonged excitation at 1.2 K. This phenomenon is attributed to quenching of excitations on antenna pigments by stable charge separation in the closely connected reaction center, possibly involving a low-quantum yield menaquinone electron acceptor.

The triplet state of the FMO complex of the green sulfur bacterium Prosthecochloris aestuarii studied with single-crystal EPR.

Triplet-electron paramagnetic resonance (EPR) spectra were obtained of single crystals of the FMO complex of the green sulfur bacterium Prostecochloris aestuarii. The experiments support the results presented in a previous paper (Louwe et al., J. Phys. Chem. 101 (1997) 11280), which showed that the experimental optical spectra of this pigment-protein complex are best reproduced by assuming that one bacteriochlorophyll (BChl 3) is energetically isolated and that this BChl is the triplet-carrying BChl of the FMO complex at cryogenic temperatures for low excitation density. When comparing the experimental and simulated data sets of the triplet-EPR spectra in single crystals, the best fit is obtained for two triplet states, one localized at BChl 3 and the other at BChl 1. The existence of two different triplet states is traced to the relatively high excitation power necessary to observe the small triplet-EPR signal of the FMO single crystals.

Spectroscopic characterization of reaction centers of the (M)Y210W mutant of the photosynthetic bacterium Rhodobacter sphaeroides.

The tyrosine-(M)210 of the reaction center of Rhodobacter sphaeroides 2.4.1 has been changed to a tryptophan using site-directed mutagenesis. The reaction center of this mutant has been characterized by low-temperature absorption and fluorescence spectroscopy, time-resolved sub-picosecond spectroscopy, and magnetic resonance spectroscopy. The charge separation process showed bi-exponential kinetics at room temperature, with a main time constant of 36 ps and an additional fast time constant of 5.1 ps. Temperature dependent fluorescence measurements predict that the lifetime of P(*) becomes 4-5 times slower at cryogenic temperatures. From EPR and absorbance-detected magnetic resonance (ADMR, LD-ADMR) we conclude that the dimeric structure of P is not significantly changed upon mutation. In contrast, the interaction of the accessory bacteriochlorophyll BA with its environment appears to be altered, possibly because of a change in its position.