Aims: The objective of this work was to provide functional evidence of key metabolic pathways important for anaerobic digestion processes through the identification of highly expressed proteins in a mixed anaerobic microbial consortium.
Methods and Results: The microbial communities from an anaerobic industrial‐like wastewater treatment bioreactor were characterized using phylogenetic analyses and metaproteomics. Clone libraries indicated that the bacterial community in the bioreactor was diverse while the archaeal population was mainly composed of Methanocorpusculum‐like (76%) micro‐organisms. Three hundred and eighty‐eight reproducible protein spots were obtained on 2‐D gels, of which 70 were excised and 33 were identified. The putative functions of the proteins detected in the anaerobic bioreactor were related to cellular processes, including methanogenesis from CO2 and acetate, glycolysis and the pentose phosphate pathway. Metaproteomics also indicated, by protein assignment, the presence of specific micro‐organisms in the bioreactor. However, only a limited overlap was observed between the phylogenetic and metaproteomic analyses.
Conclusions: This study provides some direct evidence of the microbial activities taking place during anaerobic digestion.
Significance and Impact of Study: This study demonstrates metaproteomics as a useful tool to uncover key biochemical pathways underpinning specific anaerobic bioprocesses.
Granular biomass was temporally sampled from a cold (4-15 1C) anaerobic bioreactor, which was inoculated with mesophilic biomass and used to treat industrial wastewater in a long-term (3.4 year) study. Data from 16S rRNA gene clone libraries, quantitative PCR and terminal restriction fragment length polymorphism analyses indicated that microbial community structure was dynamic, with shifts in the archaeal and bacterial communities' structures observed following start-up and during temperature decreases from 15 to 9.5 1C (phase 1). Specifically, the relative abundance of architecturally important Methanosaeta-like (acetoclastic) methanogens decreased, which was concomitant with granule disintegration and the development of a putatively psychrophilic hydrogenotrophic methanogenic community. Genetic fingerprinting suggested the development of a psychroactive methanogenic community between 4 and 10 1C (phase 2), which was dominated by acetogenic bacteria and Methanocorpusculum-like (hydrogenotrophic) methanogens. High levels of Methanosaeta-like acetoclastic methanogens and granular biofilm integrity were maintained during phase 2. Overall, decreasing temperature resulted in distinctly altered microbial community structure during phase 1, and the development of a less dynamic psychroactive methanogenic consortium during phase 2. Moreover, psychrophilic H 2 -oxidizing methanogens emerged as important members of the psychroactive consortia after 41200 days of low-temperature cultivation. The data suggest that prolonged psychrophilic cultivation of mesophilic biomass can establish a well-functioning psychroactive methanogenic consortium, thus highlighting the potential of low-temperature anaerobic digestion technology.
Whole-cell immobilization of selenate-respiring Sulfurospirillum barnesii in polyacrylamide gels was investigated to allow the treatment of selenate contaminated (790µg Se×L −1 ) synthetic wastewater with a high molar excess of nitrate (1,500 times) and sulfate (200 times). Gelimmobilized S. barnesii cells were used to inoculate a mesophilic (30°C) bioreactor fed with lactate as electron donor at an organic loading rate of 5 g chemical oxygen demand (COD)×L −1 day −1 . Selenate was reduced efficiently (>97%) in the nitrate and sulfate fed bioreactor, and a minimal effluent concentration of 39µg Se×L −1 was obtained. Scanning electron microscopy with energy dispersive X-ray (SEM-EDX) analysis revealed spherical bioprecipitates of ≤2µm diameter mostly on the gel surface, consisting of selenium with a minor contribution of sulfur. To validate the bioaugmentation success under microbial competition, gel cubes with immobilized S. barnesii cells were added to an Upflow Anaerobic Sludge Bed (UASB) reactor, resulting in earlier selenate (24 hydraulic retention times (HRTs)) and sulfate (44 HRTs) removal and higher nitrate/nitrite removal efficiencies compared to a nonbioaugmented control reactor. S. barnesii was efficiently immobilized inside the UASB bioreactors as the selenatereducing activity was maintained during long-term operation (58 days), and molecular analysis showed that S. barnesii was present in both the sludge bed and the effluent. This demonstrates that gel immobilization of specialized bacterial strains can supersede wash-out and out-competition of newly introduced strains in continuous bioaugmented systems. Eventually, proliferation of a selenium-respiring specialist occurred in the non-bioaugmented control reactor, resulting in simultaneous nitrate and selenate removal during a later phase of operation.
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