Biogeography of microbial communities in high-latitude ecosystems: Contrasting drivers for methanogens, methanotrophs and global prokaryotes.

Methane-cycling is becoming more important in high-latitude ecosystems as global warming makes permafrost organic carbon increasingly available. We explored 387 samples from three high-latitudes regions (Siberia, Alaska and Patagonia) focusing on mineral/organic soils (wetlands, peatlands, forest), lake/pond sediment and water. Physicochemical, climatic and geographic variables were integrated with 16S rDNA amplicon sequences to determine the structure of the overall microbial communities and of specific methanogenic and methanotrophic guilds. Physicochemistry (especially pH) explained the largest proportion of variation in guild composition, confirming species sorting (i.e., environmental filtering) as a key mechanism in microbial assembly. Geographic distance impacted more strongly beta diversity for (i) methanogens and methanotrophs than the overall prokaryotes and, (ii) the sediment habitat, suggesting that dispersal limitation contributed to shape the communities of methane-cycling microorganisms. Bioindicator taxa characterising different ecological niches (i.e., specific combinations of geographic, climatic and physicochemical variables) were identified, highlighting the importance of Methanoregula as generalist methanogens. Methylocystis and Methylocapsa were key methanotrophs in low pH niches while Methylobacter and Methylomonadaceae in neutral environments. This work gives insight into the present and projected distribution of methane-cycling microbes at high latitudes under climate change predictions, which is crucial for constraining their impact on greenhouse gas budgets.

Effect of Inoculum Size and Age, and Sucrose Concentration on Cell Growth to Promote Metabolites Production in Cultured Taraxacum officinale (Weber) Cells.

Pentacyclic triterpenes, including lupeol, α- amyrin, and ß-amyrin, present a large range of biological activities including anti-inflammatory, anti-cancer, and gastroprotective properties. The phytochemistry of dandelion (Taraxacum officinale) tissues has been widely described. Plant biotechnology offers an alternative for secondary metabolite production and several active plant ingredients are already synthesized through in vitro cultures. This study aimed to establish a suitable protocol for cell growth and to determine the accumulation of α-amyrin and lupeol in cell suspension cultures of T. officinale under different culture conditions. To this end, inoculum density (0.2% to 8% (w/v)), inoculum age (2- to 10-week-old), and carbon source concentration (1%, 2.3%, 3.2%, and 5.5% (w/v)) were investigated. Hypocotyl explants of T. officinale were used for callus induction. Age, size, and sucrose concentrations were statistically significant in cell growth (fresh and dry weight), cell quality (aggregation, differentiation, viability), and triterpenes yield. The best conditions for establishing a suspension culture were achieved by using a 6-week-old callus at 4% (w/v) and 1% (w/v) of sucrose concentration. Results indicate that 0.04 (±0.02) α-amyrin and 0.03 (±0.01) mg/g lupeol can be obtained in suspension culture under these starting conditions at the 8th week of culture. The results of the present study provide a backdrop for future studies in which an elicitor could be incorporated to increase the large-scale production of α-amyrin and lupeol from T. officinale.

A combined microbial and biogeochemical dataset from high-latitude ecosystems with respect to methane cycle.

High latitudes are experiencing intense ecosystem changes with climate warming. The underlying methane (CH4) cycling dynamics remain unresolved, despite its crucial climatic feedback. Atmospheric CH4 emissions are heterogeneous, resulting from local geochemical drivers, global climatic factors, and microbial production/consumption balance. Holistic studies are mandatory to capture CH4 cycling complexity. Here, we report a large set of integrated microbial and biogeochemical data from 387 samples, using a concerted sampling strategy and experimental protocols. The study followed international standards to ensure inter-comparisons of data amongst three high-latitude regions: Alaska, Siberia, and Patagonia. The dataset encompasses different representative environmental features (e.g. lake, wetland, tundra, forest soil) of these high-latitude sites and their respective heterogeneity (e.g. characteristic microtopographic patterns). The data included physicochemical parameters, greenhouse gas concentrations and emissions, organic matter characterization, trace elements and nutrients, isotopes, microbial quantification and composition. This dataset addresses the need for a robust physicochemical framework to conduct and contextualize future research on the interactions between climate change, biogeochemical cycles and microbial communities at high-latitudes.

Formation of Recalcitrant Compounds during Anaerobic Digestion of Thermally Pre-Treated Sludge: A Critical Macromolecular and Structural Study.

Thermal hydrolysis, when used as pre-treatment, enhances the anaerobic digestion of sewage sludge; moreover, due to the high temperature normally applied, undesirable recalcitrant compounds via Maillard reactions may also be formed. However, although the appearance of these recalcitrant compounds is widely reported, more information on the formation, structure, and fate of these compounds is still needed. This study was focused on understanding the amount and whereabouts of such compounds during the anaerobic digestion process with thermal pre-treatment in soluble and total phase and advance in its structural identification by analyzing their infrared (IR) spectra. It was found that, even with the improved methane production and COD degradation, at 165 °C for 30 min, humic-like compounds are formed which could not be degraded at the anaerobic digestion step. These compounds account for 25% of the original sludge. Infrared spectroscopy proved to be a powerful technique, permitting their differentiation from the natural humic-like compounds. This research provides new information about the structure of melanoidins at every stage of the thermal hydrolysis pre-treatment and how they contribute to the dissolved organic nitrogen.

Valorization of Artichoke Industrial By-Products Using Green Extraction Technologies: Formulation of Hydrogels in Combination with Paulownia Extracts.

The integral valorization of artichoke bracts generated during industrial canning of artichoke was assessed. The extraction of bioactive compounds was addressed with pressurized hot water under subcritical conditions. The performance of this stage on the extraction of phenolics with antioxidant properties and the saccharidic fraction using conventional and microwave heating was compared. The microwave assisted process was more efficient than the conventional one regarding extraction yields of total solubles, and glucose and fructose oligomers and phenolics, because lower operational temperatures and shorter times were needed. Degradation of fructose oligomers was observed at temperatures higher than 160 °C, whereas the maximal phenolic content occurred at 220 °C. Both the extracts and the residual solids, obtained at conditions leading to maximum phenolics yields, were evaluated for the production of starch-based hydrogels, supplemented with Paulownia leaves' aqueous extracts.

Temperature differently affected methanogenic pathways and microbial communities in sub-Antarctic freshwater ecosystems.

Freshwater ecosystems are responsible for an important part of the methane (CH4) emissions which are likely to change with global warming. This study aims to evaluate temperature-induced (from 5 to 20 °C) changes on microbial community structure and methanogenic pathways in five sub-Antarctic lake sediments from Magallanes strait to Cape Horn, Chile. We combined in situ CH4 flux measurements, CH4 production rates (MPRs), gene abundance quantification and microbial community structure analysis (metabarcoding of the 16S rRNA gene). Under unamended conditions, a temperature increase of 5 °C doubled MPR while microbial community structure was not affected. Stimulation of methanogenesis by methanogenic precursors as acetate and H2/CO2, resulted in an increase of MPRs up to 127-fold and 19-fold, respectively, as well as an enrichment of mcrA-carriers strikingly stronger under acetate amendment. At low temperatures, H2/CO2-derived MPRs were considerably lower (down to 160-fold lower) than the acetate-derived MPRs, but the contribution of hydrogenotrophic methanogenesis increased with temperature. Temperature dependence of MPRs was significantly higher in incubations spiked with H2/CO2 (c. 1.9 eV) compared to incubations spiked with acetate or unamended (c. 0.8 eV). Temperature was not found to shape the total microbial community structure, that rather exhibited a site-specific variability among the studied lakes. However, the methanogenic archaeal community structure was driven by amended methanogenic precursors with a dominance of Methanobacterium in H2/CO2-based incubations and Methanosarcina in acetate-based incubations. We also suggested the importance of acetogenic H2-production outcompeting hydrogenotrohic methanogenesis especially at low temperatures, further supported by homoacetogen proportion in the microcosm communities. The combination of in situ-, and laboratory-based measurements and molecular approaches indicates that the hydrogenotrophic pathway may become more important with increasing temperatures than the acetoclastic pathway. In a continuously warming environment driven by climate change, such issues are crucial and may receive more attention.

Effect of growth conditions on cell wall composition and cadmium adsorption in Chlorella vulgaris: A new approach to biosorption research.

Biosorption of toxic metals in microalgae is a process relying on the presence of cell wall reactive groups acting as binding sites. This work studied the effect of culture conditions on the outer cell wall composition of C. vulgaris and cadmium biosorption. The experiments were conducted in continuous culture under light and nitrogen limitation at two growth rates (0.4 and 0.2 d-1). Functional groups were profiled using ATR-FTIR spectrometry, and total cadmium biosorption was assayed. Significant differences in composition were attested the most salient being the absence of carboxyl groups in the light deprived states and a larger number of carbohydrates and amino groups in the nitrogen deprived cultures, particularly amino groups from deacetylated D-glucosamine polysaccharides. Higher biosorption was obtained with the nitrogen-restricted biomass, reaching a maximum of 11.9 mgCd/gbiomass, as compared to a minimum of 8.0 mgCd/gbiomass achieved in the light-restricted states. The increased biosorption exhibited by nitrogen-restricted strains was attributed to the deacetylated amino groups that have enhanced cation affinity. This work has shown that the characteristics of the outer cell wall can be engineered by culture conditions to improve biosorption, providing a new approach that opens up new research frontiers for the biosorption of hazardous metals.

Correlations between microbial population dynamics, bamA gene abundance and performance of anaerobic sequencing batch reactor (ASBR) treating increasing concentrations of phenol.

The relevant microorganims driving efficiency changes in anaerobic digestion of phenol remains uncertain. In this study correlations were established between microbial population and the process performance in an anaerobic sequencing batch reactor (ASBR) treating increasing concentrations of phenol (from 120 to 1200 mg L-1). Sludge samples were taken at different operational stages and microbial community dynamics was analyzed by 16S rRNA sequencing. In addition, bamA gene was quantified in order to evaluate the dynamics of anaerobic aromatic degraders. The microbial community was dominated by Anaerolineae, Bacteroidia, Clostridia, and Methanobacteria classes. Correlation analysis between bamA gene copy number and phenol concentration were highly significant, suggesting that the increase of aromatic degraders targeted by bamA assay was due to an increase in the amount of phenol degraded over time. The incremental phenol concentration affected hydrogenotrophic archaea triggering a linear decrease of Methanobacterium and the growth of Methanobrevibacter. The best performance in the reactor was at 800 mg L-1 of phenol. At this stage, the highest relative abundances of Syntrophorhabdus, Chloroflexus, Smithella, Methanolinea and Methanosaeta were observed and correlated positively with initial degradation rate, suggesting that these microorganisms are relevant players to maintain a good performance in the ASBR.

Enzymatic Pretreatment of Microalgae: Cell Wall Disruption, Biomass Solubilisation and Methane Yield Increase.

Anaerobic digestion of microalgal biomass for biogas production may be limited due to the cell wall resulting in an inefficient bioconversion. Enzymatic pretreatments are applied for inducing cell damage/lysis and organic matter solubilisation and this way increasing biogas production. We evaluated enzymatic pretreatments in different conditions for comparing in relation to cell wall rupture, increase of soluble material and increase in biogas production through anaerobic digestion performance in BMP assay. Chlorella sorokiniana cultures were subjected to three different enzymatic pretreatments, each under four different conditions of enzyme/substrate ratio, pH and application time. The results showed increases over 21% in biogas productions for all enzymatic pretreatments. Enzymatic pretreatment was effective at damaging microalgae cell wall, releasing organic compounds and increasing the rate and final methane yield in BMP tests. We observed a synergistic activity between the mixtures enzymes, which would depend on operational conditions used for each pretreatment.

A VFA-based controller for anaerobic digestion of industrial winery wastewater.

A variable-gain controller for anaerobic digestion of industrial winery wastewater is presented. A control law using both volatile fatty acids (VFA) and methane production rate as controlled variables and organic loading rate (OLR) as manipulated variable is defined. The process state is quantitatively estimated by an empirical function comparing VFA measurements against a setpoint value; then, it is modified with a second empirical function that compares the methane flow rate with a maximum capacity reference, and finally it is adjusted with a third factor considering the actual hydraulic retention time. The variable-gain function determines the extent of the OLR change applied to the system. The controller was successfully validated in a 95 L upflow-anaerobic-sludge-blanket (UASB) reactor, treating industrial wine wastewater at OLR ranged between 2.0 and 39.2 g COD/L d for 120 days at mesophilic conditions. Higher performance was achieved contrasted with a conventional strategy carried out in a parallel UASB unit.

Kinetics and modeling of cell growth for potential anthocyanin induction in cultures of Taraxacum officinale G.H. Weber ex Wiggers (Dandelion) in vitro

Background: Taraxacum officinale G.H. Weber ex Wiggers is a wild plant used in folk medicine to treat several diseases owing to bioactive secondary metabolites present in its tissue. The accumulation of such molecules in plant cells can occur as a response against abiotic stress, but these metabolites are often deposited in low concentrations. For this reason, the use of a biotechnological approach to improve the yields of technologically interesting bioactive compounds such as anthocyanins is a compelling option. This work focuses on investigating the potential of in vitro T. officinale cultures as an anthocyanin source. Results: To demonstrate the suitability of anthocyanin induction and accumulation in calluses under specific conditions, anthocyanin was induced in the T. officinale callus. A specific medium of 5.5% sucrose supplemented with 6-benzylaminopurine /1-naphthaleneacetic acid in a 10:1 ratio was used to produce an anthocyanin yield of 1.23 mg g-1 fw. An in vitro dandelion callus line was established from this experiment. Five mathematical models were then used to objectively and predictably explain the growth of anthocyanin-induced calluses from T. officinale. Of these models, the Richards model offered the most suitable representation of anthocyanin callus growth in a solid medium and permitted the calculation of the corresponding kinetic parameters. Conclusions: The findings demonstrate the potential of an in vitro anthocyanin-induced callus line from T. officinale as an industrial anthocyanin source.

Key microbial populations involved in anaerobic degradation of phenol and p-cresol using different inocula

Background: Anaerobic digestion is an alternative bioprocess used to treat effluents containing toxic compounds such as phenol and p-cresol. Selection of an adequate sludge as inoculum containing an adapted microbial consortium is a relevant factor to improve the removal of these pollutants. The objective of this study is to identify the key microorganisms involved in the anaerobic digestion of phenol and p-cresol and elucidate the relevance of the bamA gene abundance (a marker gene for aromatic degraders) in the process, in order to establish new strategies for inocula selection and improve the system's performance. Results: Successive batch anaerobic digestion of phenol and p-cresol was performed using granular or suspended sludge. Granular sludge in comparison to suspended sludge showed higher degradation rates both for phenol (11.3 ± 0.7 vs 8.1 ± 1.1 mg l-1 d-1) and p-cresol (7.8 ± 0.4 vs 3.7 ± 1.0 mg l-1 d-1). After three and four re-feedings of phenol and p-cresol, respectively, the microbial structure from both sludges was clearly different from the original sludges. Anaerobic digestion of phenol and p-cresol generated an abundance increase in Syntrophorhabdus genus and bamA gene, together with hydrogenotrophic and aceticlastic archaea. Analysis of results indicates that differences in methanogenic pathways and levels of Syntrophorhabdus and bamA gene in the inocula, could be the causes of dissimilar degradation rates between each sludge. Conclusions: Syntrophorhabdus and bamA gene play relevant roles in anaerobic degradation of phenolics. Estimation of these components could serve as a fast screening tool to find the most acclimatized sludge to efficiently degrade mono-aromatic compounds.

Assessing the Effect of Pretreatments on the Structure and Functionality of Microbial Communities for the Bioconversion of Microalgae to Biogas.

Microalgae biomethanization is driven by anaerobic sludge associated microorganisms and is generally limited by the incomplete hydrolysis of the microalgae cell wall, which results in a low availability of microalgal biomass for the methanogenic community. The application of enzymatic pretreatments, e.g., with hydrolytic enzymes, is among the strategies used to work around the incomplete hydrolysis of the microalgae cell wall. Despite the proven efficacy of these pretreatments in increasing biomethanization, the changes that a given pretreatment may cause to the anaerobic sludge associated microorganisms during biomethanization are still unknown. This study evaluated the changes in the expression of the metatranscriptome of anaerobic sludge associated microorganisms during Chlorella sorokiniana biomethanization without pretreatment (WP) (control) and pretreated with commercial cellulase in order to increase the solubilization of the microalgal organic matter. Pretreated microalgal biomass experienced significant increases in biogas the production. The metatranscriptomic analysis of control samples showed functionally active microalgae cells, a bacterial community dominated by γ- and δ-proteobacteria, and a methanogenic community dominated by Methanospirillum hungatei. In contrast, pretreated samples were characterized by the absence of active microalgae cells and a bacteria population dominated by species of the Clostridia class. These differences are also related to the differential activation of metabolic pathways e.g., those associated with the degradation of organic matter during its biomethanization.

Active and total microbial community dynamics and the role of functional genes bamA and mcrA during anaerobic digestion of phenol and p-cresol.

The aim of the present work was to investigate the dynamics of microbial community at DNA and RNA level and the role of bamA and mcrA gene during anaerobic digestion of phenol and p-cresol. Anaerobic digestion was conducted in batch reactors and microbial community dynamics was analysed. Results showed that active microbial community was quite dissimilar in comparison to the total microbial community. Syntrophorhabdus and Bacillus were the dominant active bacterial genera whereas Methanosaeta together with Methanobacterium showed the highest potential activity in the Archaea domain indicating a relevant role of these microorganisms in the anaerobic process. Ecological Networks revealed dissimilar interactions at DNA and RNA level, being the latter a better descriptor of the known roles of dominant OTUs. QRT-PCR results showed that expression of bamA gene correlated positively with instantaneous degradation rate proving for first time its functionality and its relationship with the kinetics of the process.

A need for a standardization in anaerobic digestion experiments? Let's get some insight from meta-analysis and multivariate analysis.

An important variability in the experimental results in anaerobic digestion lab test has been reported. This study presents a meta-analysis coupled with multivariate analysis aiming to assess the impact of this experimental variability in batch and continuous operation at mesophilic and thermophilic anaerobic digestion of waste activated sludge. An analysis of variance showed that there was no significant difference between mesophilic and thermophilic conditions in both continuous and batch conditions. Concerning the operation mode, the values of methane yield were significantly higher in batch experiment than in continuous reactors. According to the PCA, for both cases, the methane yield is positive correlated to the temperature rises. Interestingly, in the batch experiments, the higher the volatile solids in the substrate was, the lowest was the methane production, which is correlated to experimental flaws when setting up those tests. In continuous mode, unlike the batch test, the methane yield is strongly (positively) correlated to the organic content of the substrate. Experimental standardization, above all, in batch conditions are urgently necessary or move to continuous experiments for reporting results. The modeling can also be a source of disturbance in batch test.

Isolation and Identification of Compounds from Bioactive Extracts of Taraxacum officinale Weber ex F. H. Wigg. (Dandelion) as a Potential Source of Antibacterial Agents.

Currently, the most effective treatment for recurrent urinary tract infections in women is antibiotics. However, the limitation for this treatment is the duration and dosage of antibiotics and the resistance that bacteria develop after a long period of administration. With the aim of identifying mainly novel natural agents with antibacterial activity, the present study was undertaken to investigate the biological and phytochemical properties of extracts from the leaves Taraxacum officinale. The structural identification of compounds present in hexane (Hex) and ethyl acetate (AcOEt) extracts was performed by mass spectrometry (GC-MS) spectroscopic techniques and nuclear magnetic resonance (NMR) with the major compounds corresponding to different sesquiterpene lactones (α-santonin, glabellin, arborescin, and estafiatin), monoterpene (9,10-dimethyltricycle [4.2.1.1 (2,5)]decane-9,10-diol), phytosterol (Stigmasta-5,22-dien-3ß-ol acetate), terpenes (lupeol acetate, pregn-5-en-20-one-3ß-acetyloxy-17-hydroxy, 2-hydroxy-4-methoxy benzaldehyde), and coumarin (benzofuranone 5,6,7,7-a-tetraaldehyde-4,4,7a-trimethyl). The results obtained show that the Hex extract was highly active against Staphylococcus aureus showing a MIC of 200 µg/mL and moderately active against Escherichia coli and Klebsiella pneumoniae with MIC values of 400 µg/mL and 800 µg/mL for the other Gram-negative strains tested with Proteus mirabilis as uropathogens in vitro. Therefore, the effective dandelion extracts could be used in the development of future products with industrial application.

On the derivation of a simple dynamic model of anaerobic digestion including the evolution of hydrogen.

Hydrogen has been found to be an important intermediate during anaerobic digestion (AD) and a key variable for process monitoring as it gives valuable information about the stability of the reactor. However, simple dynamic models describing the evolution of hydrogen are not commonplace. In this work, such a dynamic model is derived using a systematic data driven-approach, which consists of a principal component analysis to deduce the dimension of the minimal reaction subspace explaining the data, followed by an identification of the kinetic parameters in the least-squares sense. The procedure requires the availability of informative data sets. When the available data does not fulfill this condition, the model can still be built from simulated data, obtained using a detailed model such as ADM1. This dynamic model could be exploited in monitoring and control applications after a re-identification of the parameters using actual process data. As an example, the model is used in the framework of a control strategy, and is also fitted to experimental data from raw industrial wine processing wastewater.

Physical Pretreatment Methods for Improving Microalgae Anaerobic Biodegradability.

Microalgae may be a potential feedstock for biogas production through anaerobic digestion. However, this process is limited by the hydrolytic stage, due to the complex and resistant microalgae cell wall components. This fact hinders biomass conversion into biogas, demanding the application of pretreatment techniques for inducing cell damage and/or lysis and organic matter solubilisation. In this study, sonication, thermal, ultrasound, homogeneizer, hydrothermal and steam explosion pretreatments were evaluated in different conditions for comparing their effects on anaerobic digestion performance in batch reactors. The results showed that the highest biomass solubilisation values were reached for steam explosion (65-73%) and ultrasound (33-57%). In fact, only applied energies higher than 220 W or temperatures higher than 80 °C induced cell wall lysis in C. sorokiniana. Nonetheless, the highest methane yields were not correlated to biogas production. Thermal hydrolysis and steam explosion showed lower methane yields in respect to non-pretreated biomass, suggesting the presence of toxic compounds that inhibited the biological process. Accordingly, these pretreatment techniques led to a negative energy balance. The best pretreatment method among the ones evaluated was thermal pretreatment, with four times more energy produced that demanded.

Influence of phenylacetic acid pulses on anaerobic digestion performance and archaeal community structure in WWTP sewage sludge digesters.

The effect of phenylacetic acid (PAA) pulses on anaerobic digestion (AD) performance and archaeal community structure was evaluated in anaerobic digesters treating sewage sludge from a wastewater treatment plant (WWTP). Four pilot-scale continuous stirred tank reactors were set up at a full-scale municipal WWTP in Santiago de Chile, and fed with either primary or mixed sewage sludge. AD performance was evaluated by volatile fatty acid (VFA) and biogas production monitoring. Archaeal community structure was characterized by 16S rRNA denaturing gradient gel electrophoresis and band sequencing. In the primary sludge digester, a single PAA pulse at 200 mg L(-1) was sufficient to affect AD performance and archaeal community structure, resulting in long-term VFA accumulation, reduced biogas production and community shift from dominant acetoclastic (Methanosaeta concilii) to hydrogenotrophic (Methanospirillum hungatei) methanogens. By contrast, AD performance and archaeal community structure in the mixed sludge digester were stable and resistant to repeated PAA pulses at 200 and 600 mg L(-1). This work demonstrated that the effect of PAA pulses on methanogenic activity and archaeal community structure differed according to AD substrate, and suggests that better insights of the correlations between archaeal population dynamics and functional performance could help to better face toxic shocks in AD.