Construction of Xylose-Utilizing Cyanobacterial Chassis for Bioproduction Under Photomixotrophic Conditions.

Xylose is a major component of lignocellulose and the second most abundant sugar present in nature after glucose; it, therefore, has been considered to be a promising renewable resource for the production of biofuels and chemicals. However, no natural cyanobacterial strain is known capable of utilizing xylose. Here, we take the fast-growing cyanobacteria Synechococcus elongatus UTEX 2973 as an example to develop the synthetic biology-based methodology of constructing a new xylose-utilizing cyanobacterial chassis with increased acetyl-CoA for bioproduction.

Appraising the phycoremediation potential of cyanobacterial strains Phormidium and Oscillatoria for nutrient removal from textile wastewater (TWW) and synchronized biodiesel production from TWW-tolerant biomass.

In this study, phycoremediation of textile wastewater (TWW) by freshwater cyanobacterial strains such as sp., Oscillatoria sp. F01 and Oscillatoria sp. F02 was evaluated, and lipids were simultaneously extracted from biomass for biodiesel production. Onset of the study, Phormidium sp. and Oscillatoria sp. F01 has better growth rates, increased biomass production, high chlorophyll content, and efficient nutrient utilization in TWW compared to Oscillatoria sp. F02. Phormidium sp. showed 1.41 g/L dry weight, followed by Oscillatoria sp. F01 with 1.39 g/L and Oscillatoria sp. F02 with 1.02 g/L biomass. Both strains demonstrated their capability to elevate the pH level while reducing TDS and eliminating/reducing several nutrients such as nitrates, nitrites, phosphates, sulphates, sulphides, chlorides, calcium, sodium, and magnesium. Further, the total lipids extracted from the TWW-grown Phormidium sp., Oscillatoria sp. F01 and Oscillatoria sp. F02 was estimated to be 8.20, 13.70 and 11.20 %, respectively, on day 21, which was higher than the lipid content obtained from control cultures. Further, biodiesel produced from the lipids of all strains showed higher levels of C12:0, C16:0, C16:1, C18:1, C18:2, and C18:3 among all the fatty acids. Therefore, they can potentially offer a valuable source of lipids and diverse fatty acids for high-quality biodiesel production. This integrated system not only offers a solution for TWW treatment but also provides a feedstock for renewable fuel production simultaneously.

Enhancing biogas generation from lignocellulosic biomass through biological pretreatment: Exploring the role of ruminant microbes and anaerobic fungi.

Biogas production from Lignocellulosic Biomass (LB) via anaerobic digestion (AD) has gained attention for its potential in self-sustainability. However, the recalcitrance of LB cell walls pose a challenge to its degradability and biogas generation. Therefore, pretreatment of LB is necessary to enhance lignin removal and increase degradability. Among the different approaches, environmentally friendly biological pretreatment ispromising as it avoids the production of inhibitors. The ruminal microbial community, including anaerobic fungi, bacteria, and protozoa, has shown an ability to effectively degrade LB through biomechanical and microbial penetration of refractory cell structures. In this review, we provide an overview of ruminant microbes dominating LB's AD, their degradation mechanism, and the bioaugmentation of the rumen. We also explore the potential cultivation of anaerobic fungi from the rumen, their enzyme potential, and their role in AD. The rumen ecosystem, comprising both bacteria and fungi, plays a crucial role in enhancing AD. This comprehensive review delves into the intricacies of ruminant microorganisms' adhesion to plant cells, elucidates degradation mechanisms, and explores integrated pretreatment approaches for the effective utilization of LB, minimizing the impact of inhibitors. The discussion underscores the considerable potential of ruminant microbes in pretreating LB, paving the way for sustainable biogas production. Optimizing fungal colonization and ligninolytic enzyme production, such as manganese peroxidase and laccase, significantly enhances the efficiency of fungal pretreatment. Integrating anaerobic fungi through bioaugmentation during mainstream processing demonstrably increases methane production. This study opens promising avenues for further research and development of these microorganisms for bioenergy production.

Nitrogen and phosphorus stress as a tool to induce lipid production in microalgae.

Microalgae, capable of accumulating large amounts of lipids, are of great value for biodiesel production. The high cost of such production stimulates the search for cultivation conditions that ensure their highest productivity. Reducing the content of nitrogen and phosphorus in the culture medium is widely used to change the content and productivity of lipids in microalgae. Achieving the right balance between maximum growth and maximum lipid content and productivity is the primary goal of many experimental works to ensure cost-effective biodiesel production from microalgae. The content of nitrogen and phosphorus in nutrient media for algal cultivation after converted to nitrogen (-N) and phosphorus (-P) lies in an extensive range: from 0.007 g L- 1 to 0.417 g L- 1 and from 0.0003 g L- 1 to 0.227 g L- 1 and N:P ratio from 0.12:1 to 823.33:1. When studying nutritional stress in microalgae, no single approach is used to determine the experimental concentrations of nitrogen and phosphorus. This precludes the possibility of correct interpretation of the data and may lead to erroneous conclusions. This work results from the systematisation of information on using nitrogen and phosphorus restriction to increase the lipid productivity of microalgae of different taxonomic and ecological groups to identify future research directions. The results of 301 experiments were included in the analysis using the principal components method. The investigation considered various divisions and classes: Cyanobacteria, Rhodophyta, Dinophyta, Haptophyta, Cryptophyta, Heterokontophyta/Ochrophyta (Bacillariophyceae, Eustigmatophyceae, Xanthophyceae), Chlorophyta, and also the ratio N:P, the time of the experiment, the light intensity during cultivation. Based on the concentrations of nitrogen and phosphorus existing in various nutrient media, a general scheme for designating the supply of nutrient media for nitrogen (as NO3- or NH4+, N g L- 1) and phosphorus (as РO4-, P g L- 1) has been proposed: replete -N (˃0.4 g L- 1), moderate -N (0.4-0.2), moderate N-limitation (0.19-0.1), strong N-limitation (˂0.1), without nitrogen (0), replete -Р (˃0.2), moderate -P (0.2-0.02), moderate P-limitation (0.019-0.01), strong P-limitation (˂0.01), without phosphorus (0).

Overview of microalgae and cyanobacteria-based biostimulants produced from wastewater and CO2 streams towards sustainable agriculture: A review.

For a long time, marine macroalgae (seaweeds) have been used to produce commercial biostimulants in order to ensure both productivity and quality of agricultural crops under abiotic stress. With similar biological properties, microalgae have slowly attracted the scientific community and the biostimulant industry, in particular because of their ability to be cultivated on non-arable lands with high biomass productivity all year long. Moreover, the recent strategies of culturing these photosynthetic microorganisms using wastewater and CO2 opens the possibility to produce large quantity of biomass at moderate costs while integrating local and circular economy approaches. This paper aims to provide a state of the art review on the development of microalgae and cyanobacteria based biostimulants, focusing on the different cultivation, extraction and application techniques available in the literature. Emphasis will be placed on microalgae and cyanobacteria cultivation using liquid and gaseous effluents as well as emerging green-extraction approaches, taking in consideration the actual European regulatory framework.

Rhizopus stolonifer biomass catalytic transesterification capability: optimization of cultivation conditions.

BACKGROUND: Using fungal biomass for biocatalysis is a potential solution for the expensive cost of the use o enzymes. Production of fungal biomass with effective activity requires optimizing the cultivation conditions. RESULTS: Rhizopus stolonifer biomass was optimized for transesterification and hydrolysis of waste frying oil (WFO). Growth and biomass lipolytic activities of R. stolonifer improved under shaking conditions compared to static conditions, and 200 rpm was optimum. As biomass lipase and transesterification activities inducer, olive oil was superior to soybean, rapeseed, and waste frying oils. Biomass produced in culture media containing fishmeal as an N-source feedstock had higher lipolytic capabilities than corn-steep liquor and urea. Plackett Burman screening of 9 factors showed that pH (5-9), fishmeal (0.25-1.7%, w/v), and KH2PO4 (0.1-0.9%, w/v) were significant factors with the highest main effect estimates 11.46, 10.42, 14.90, respectively. These factors were selected for response surface methodology (RSM) optimization using central composite design (CCD). CCD models for growth, biomass lipase activity, and transesterification capability were significant. The optimum conditions for growth and lipid modification catalytic activities were pH 7.4, fishmeal (2.62%, w/v), and KH2PO4 (2.99%, w/v). CONCLUSION: Optimized culture conditions improved the whole cell transesterification capability of Rhizopus stolonifer biomass in terms of fatty acid methyl ester (FAME) concentration by 67.65% to a final FAME concentration of 85.5%, w/w.

Quantification of indicator and pathogenic bacteria in manures and digestates from three agricultural biogas plants over a one-year period.

Interest in the conversion of manure in biogas via anaerobic digestion (AD) is growing, but questions remain about the biosafety of digestates. For a period of one year, we monitored the impact of three mesophilic agricultural biogas plants (BPs) mainly fed with pig manure (BP1, BP3) or bovine manure (BP2) on the physicochemical parameters, the composition of the microbial community and the concentration of bacteria (E. coli, enterococci, Salmonella, Campylobacter, Listeria monocytogenes, Clostridium perfringens, Clostridium botulinum and Clostridioides difficile). The BP2 digestate differed from those of the two other BPs with a higher nitrogen content, more total solids and greater abundance of Clostridia MBA03 and Disgonomonadacea. Persistence during digestion ranked from least to most, was: Campylobacter (1.6 to >2.9 log10 reduction, according to the BP) < E. coli (1.8 to 2.2 log10) < Salmonella (1.1 to 1.4 log10) < enterococci (0.2 to 1.2 log10) and C. perfringens (0.2 to 1 log10) < L. monocytogenes (-1.2 to 1.6 log10) < C. difficile and C. botulinum (≤0.5 log10). No statistical link was found between the reduction in the concentration of the targeted bacteria and the physicochemical and operational parameters likely to have an effect (NH3, volatile fatty acids and total solids contents, hydraulic retention time, presence of co-substrates), underlining the fact that the fate of the bacteria during mesophilic digestion depends on many interacting factors. The reduction in concentrations varied significantly over the sampling period, underlining the need for longitudinal studies to estimate the impact of AD on pathogenic microorganisms.

Inter-kingdom interactions and stability of methanogens revealed by machine-learning guided multi-omics analysis of industrial-scale biogas plants.

Multi-omics analysis is a powerful tool for the detection and study of inter-kingdom interactions, such as those between bacterial and archaeal members of complex biogas-producing microbial communities. In the present study, the microbiomes of three industrial-scale biogas digesters, each fed with different substrates, were analysed using a machine-learning guided genome-centric metagenomics framework complemented with metatranscriptome data. This data permitted us to elucidate the relationship between abundant core methanogenic communities and their syntrophic bacterial partners. In total, we detected 297 high-quality, non-redundant metagenome-assembled genomes (nrMAGs). Moreover, the assembled 16 S rRNA gene profiles of these nrMAGs showed that the phylum Firmicutes possessed the highest copy number, while the representatives of the archaeal domain had the lowest. Further investigation of the three anaerobic microbial communities showed characteristic alterations over time but remained specific to each industrial-scale biogas plant. The relative abundance of various microorganisms as revealed by metagenome data was independent from corresponding metatranscriptome activity data. Archaea showed considerably higher activity than was expected from their abundance. We detected 51 nrMAGs that were present in all three biogas plant microbiomes with different abundances. The core microbiome correlated with the main chemical fermentation parameters, and no individual parameter emerged as a predominant shaper of community composition. Various interspecies H2/electron transfer mechanisms were assigned to hydrogenotrophic methanogens in the biogas plants that ran on agricultural biomass and wastewater. Analysis of metatranscriptome data revealed that methanogenesis pathways were the most active of all main metabolic pathways.

Performance of co-culture strategy on nutrient removal and biogas upgrading by strigolactone induction.

In this study, we investigated the performance and elucidated the synergistic effects of microalgae-fungi symbionts co-cultured with 10-7 and 10-9  mol L-1 of GR24 and supplemented with endophytic bacteria, multi-walled carbon nanotubes (MWCNTs) or vitamin B12 (VB12), on nutrient removal and biogas upgrading. The results showed that the microalgae-fungi-bacteria symbiotic system co-cultured with 10-9  mol L-1 GR24 presented the optimal growth performance of 0.368 ± 0.04 d-1 , chlorophyll a of 249.36 ± 22.31 µg L-1 , and extracellular carbonic anhydrase activity of 42.55 ± 3.755 enzyme units. In this co-culture system, the organic matter, nutrients, and CO2 purification obtained the highest removal efficiency, with 81.35 ± 7.96% for chemical oxygen demand, 83.56 ± 7.91% total nitrogen, 84.17 ± 7.95% total phosphorus, and 63.72 ± 6.06% CO2 . The symbiont system also greatly increased the methane content in the biogas by 30.67%. The remarkable performance of the microalgae-fungi-bacteria symbiotic system shows its ability to be broadly applied in simultaneous biogas upgrading and wastewater treatment. PRACTITIONER POINTS: The optimal GR24 concentration for microalgae-fungi consortia was 10-9  M. Endophytic bacteria were superior to MWCNTs and VB12. Fungi-algae-bacteria consortia presented excellent growth and removal performance. Removal efficiencies of COD, TN, and TP were about 81% under optimum treatment.

Impact of carbon monoxide on performance and microbial community of extreme-thermophilic hydrogenotrophic methanation in horizontal rotary bioreactor.

A novel horizontal rotary bioreactor was developed for upgrading biogas from coke oven gas at extreme-thermophilic condition. The introduction of CO decreased the outlet methane content from 80% to 50% due to insufficient H2. This hindrance was overcome by increasing the proportion of incoming hydrogen, coupled with a prolonged gas retention time from 24 to 72 h, leading to a restoration of methane content to 91.6%. Notably, CO and CO2 exhibited a competitive relationship to hydrogen, which was determined by their contents. The substitution of Methanothermobacter for Methanobacterium as the dominant genus was observed at 70 °C, with relative abundance exceeding 98%. Incorporation of CO increased bacteria diversity and fostered a syntrophic relationship between the bacterial community and M. thermautotrophicus. This study provides both theoretical basis and practical support for biogas upgrading from coke oven gas using a biofilm reactor, thus aiding its future industrialization prospects.

Unraveling prevalence of homoacetogenesis and methanogenesis pathways due to inhibitors addition.

Three inhibitors targeting different microorganisms, both from Archaea and Bacteria domains, were evaluated for their effect on CO2 biomethanation: sodium ionophore III (ETH2120), carbon monoxide (CO), and sodium 2-bromoethanesulfonate (BES). This study examines how these compounds affect the anaerobic digestion microbiome in a biogas upgrading process. While archaea were observed in all experiments, methane was produced only when adding ETH2120 or CO, not when adding BES, suggesting archaea were in an inactivated state. Methane was produced mainly via methylotrophic methanogenesis from methylamines. Acetate was produced at all conditions, but a slight reduction on acetate production (along with an enhancement on CH4 production) was observed when applying 20 kPa of CO. Effects on CO2 biomethanation were difficult to observe since the inoculum used was from a real biogas upgrading reactor, being this a complex environmental sample. Nevertheless, it must be mentioned that all compounds had effects on the microbial community composition.

Taxonomic and enzymatic basis of the cellulolytic microbial consortium KKU-MC1 and its application in enhancing biomethane production.

Lignocellulosic biomass is a promising substrate for biogas production. However, its recalcitrant structure limits conversion efficiency. This study aims to design a microbial consortium (MC) capable of producing the cellulolytic enzyme and exploring the taxonomic and genetic aspects of lignocellulose degradation. A diverse range of lignocellulolytic bacteria and degrading enzymes from various habitats were enriched for a known KKU-MC1. The KKU-MC1 was found to be abundant in Bacteroidetes (51%), Proteobacteria (29%), Firmicutes (10%), and other phyla (8% unknown, 0.4% unclassified, 0.6% archaea, and the remaining 1% other bacteria with low predominance). Carbohydrate-active enzyme (CAZyme) annotation revealed that the genera Bacteroides, Ruminiclostridium, Enterococcus, and Parabacteroides encoded a diverse set of cellulose and hemicellulose degradation enzymes. Furthermore, the gene families associated with lignin deconstruction were more abundant in the Pseudomonas genera. Subsequently, the effects of MC on methane production from various biomasses were studied in two ways: bioaugmentation and pre-hydrolysis. Methane yield (MY) of pre-hydrolysis cassava bagasse (CB), Napier grass (NG), and sugarcane bagasse (SB) with KKU-MC1 for 5 days improved by 38-56% compared to non-prehydrolysis substrates, while MY of prehydrolysed filter cake (FC) for 15 days improved by 56% compared to raw FC. The MY of CB, NG, and SB (at 4% initial volatile solid concentration (IVC)) with KKU-MC1 augmentation improved by 29-42% compared to the non-augmentation treatment. FC (1% IVC) had 17% higher MY than the non-augmentation treatment. These findings demonstrated that KKU-MC1 released the cellulolytic enzyme capable of decomposing various lignocellulosic biomasses, resulting in increased biogas production.

A state of the art review on the co-cultivation of microalgae-fungi in wastewater for biofuel production.

The microalgae have a great potential as the fourth generation biofuel feedstock to deal with energy crisis, but the cost of production and biomass harvest are the major hurdles in terms of large scale production and applications. Using filamentous fungi to culture targeted alga for biomass accumulation and eventually harvesting is a sustainable way to mitigate environmental impacts. Microalgal co-culture method could be an alternative to overcome limitations and increase biomass yield and lipid accumulation. It was found to be the high feasibility for the production of biofuels from fungi and microalgae using wastewater. This article aimed to state the synergistic approaches, their culture protocols, harvesting procedure and their potential biotechnological applications. Additionally, algal-fungal consortia could digest cellulosic biomass, potentially reducing operating costs as part of industrial need. As a result of co-cultivation, biofuel production could be economically feasible owing to its excellent ability to treat wastewater and be eco-friendly. The implications of the innovative co-cultivation technology have demonstrated the potential for further development based on the policies that have been supported and implemented.

Bacterial metabolic engineering for the production of second-generation (2 G) bioethanol and biobutanol; a review.

The second generation (2 G) biofuels were introduced to solve the issues associated with first-generation biofuel (dependency on food materials) and fossil fuels, such as reservoirs diminution, high demand, price fluctuation, and lethal greenhouse gases emission. Butanol and ethanol are the main 2 G biofuels. They are used as a disinfectant, antiseptic, and chemical solvent in the pharmaceutical, plastic, textiles, cosmetics, and fuel industries. Currently, their bacterial biological production from lignocellulosic material at the industrial level with primitive microorganisms is under development and not economical and qualitative compatible as compared to that of fossil origin, due to the slow growth rate, low titer, recalcitrant nature of lignocellulose, strain intolerance to a higher amount of butanol and ethanol, and strain inability to tolerate inhibitors accumulated during pretreatment of lignocellulosic materials. Therefore, metabolic engineering strategies such as redirection of carbon flux, knocking out competing pathways, enhancing strain robustness and wide range of substrate utilization ability, and overexpression of enzymes involved in their biological synthesis have been applied to bacteria for enhancing their ability for 2 G ethanol and butanol production in a highly cost-effective amount from lignocellulosic materials. Herein, we summarized and reviewed the progress in metabolic engineering of bacterial species such as Clostridium spp,Escherichia coli, and Zymomonas mobilis for the synthesis of 2 G butanol and ethanol, especially from lignocellulosic materials.

Effect of GR24 concentrations on tetracycline and nutrient removal from biogas slurry by different microalgae-based technologies.

A biogas slurry composed of carbon, nitrogen, phosphorus, and antibiotics was generated. Investigations into the nutrient and tetracycline removal performance of four microalgae-based contaminant removal technologies, including Chlorella vulgaris, C. vulgaris co-cultured with endophytic bacteria, C. vulgaris co-cultured with Ganoderma lucidum, and C. vulgaris co-cultured with G. lucidum and endophytic bacteria, were conducted. The algal-bacterial-fungal consortium with 10-9 M strigolactone (GR24) yielded the maximum growth rate and average daily yield for algae at 0.325 ± 0.03 d-1 and 0.192 ± 0.02 g L-1 d-1, respectively. The highest nutrient/ tetracycline removal efficiencies were 83.28 ± 7.95 % for chemical oxygen demand (COD), 82.62 ± 7.97 % for total nitrogen (TN), 85.15 ± 8.26 % for total phosphorus (TP) and 83.92 ± 7.65 % for tetracycline. Adding an algal-bacterial-fungal consortium with an optimal synthetic analog GR24 concentration is seemingly an encouraging strategy for enhancing pollutant removal by algae, possibly overcoming the challenges of eutrophication and antibiotic pollution.

Nutrient removal from biogas slurry and biogas upgrading by microalgae-fungi-bacteria co-cultivation under different carbon nanotubes concentration.

In this study, Chlorella vulgaris, Ganoderma lucidum, and endophytic bacteria were co-cultivated with the stimulation of strigolactone analogs GR24 to prepare pellets. During the purification of biogas slurry and biogas, multi-walled carbon nanotubes (MWCNTs) were introduced to enhance the removal efficiencies of nutrients and CO2. The results showed that both GR24 and MWCNTs affected the purification of biogas slurry and biogas. The maximum chemical oxygen demand, total nitrogen, total phosphorus, and CO2 removal efficiencies of the Chlorella vulgaris-Ganoderma lucidum-endophytic bacterial symbionts were 82.57 ± 7.96% (P < 0.05), 82.14 ± 7.87% (P < 0.05), 84.27 ± 7.96% (P < 0.05), and 63.93 ± 6.22% (P < 0.05), respectively, with the induction of 10-9 M GR24 and 1 mg L-1 MWCNTs. Moreover, the growth and photosynthetic performance of the symbionts were consistent with the removal effects. The Chlorella vulgaris-Ganoderma lucidum-endophytic bacterial symbionts obtained high growth rates and enzyme activity with the maximum growth rate of 0.365 ± 0.03 d-1, mean daily productivity of 0.182 ± 0.016 g L-1 d-1, and carbonic anhydrase activity of 31.07 ± 2.75 units, respectively. These results indicated that an appropriate concentration of GR24 and MWCNTs could promote the growth of symbionts, reinforce the purification effects of biogas slurry and biogas, and provide a new idea for the simultaneous purification of wastewater and biogas.

Biogas digestate as a sustainable phytosterol source for biotechnological cascade valorization.

Every year, several million tonnes of anaerobic digestate are produced worldwide as a by-product of the biogas industry, most of which is applied as agricultural fertilizer. However, in the context of a circular bioeconomy, more sustainable uses of residual digestate biomass would be desirable. This study investigates the fate of the sterol lipids ß-sitosterol and cholesterol from the feedstocks to the final digestates of three agricultural and one biowaste biogas plants to assess if sterols are degraded during anaerobic digestion or if they remain in the digestate, which could provide a novel opportunity for digestate cascade valorization. Gas chromatographic analyses showed that feedstock sterols were not degraded during anaerobic digestion, resulting in their accumulation in the digestates to up to 0.15% of the dry weight. The highest concentrations of around 1440 mg ß-sitosterol and 185 mg cholesterol per kg dry weight were found in liquid digestate fractions, suggesting partial sterol solubilization. Methanogenic batch cultures spiked with ß-sitosterol, cholesterol, testosterone and ß-oestradiol confirmed that steroids persist during anaerobic digestion. Mycobacterium neoaurum was able to transform digestate sterols quantitatively into androstadienedione, a platform chemical for steroid hormones, without prior sterol extraction or purification. These results suggest that digestate from agricultural and municipal biowaste is an untapped resource for natural sterols for biotechnological applications, providing a new strategy for digestate cascade valorization beyond land application.

The effects of influent chemical oxygen demand and strigolactone analog concentration on integral biogas upgrading and pollutants removal from piggery wastewater by different microalgae-based technologies.

Microalgae-based technologies are promising strategies for efficient wastewater treatment and biogas upgrading. In this study, three types of microalga-fungi/bacteria symbiotic systems stimulated with the strigolactone analog (GR24) were used to simultaneously remove nutrients from treated piggery wastewater and CO2 from biogas. The effects of initial concentrations of chemical oxygen demand (COD) and GR24 on nutrient removal and biogas upgrading were investigated. When the initial COD concentration was 1200 mg/L, the Chlorella vulgaris-Ganoderma lucidum-endophytic bacteria co-cultivation systems achieved the best photosynthetic performance and microalgae growth. Moreover, under the appropriate COD concentration (1200 mg/L), the highest nutrient/CO2 removal efficiencies were obtained. In addition, 10-9 M GR24 significantly accelerated nutrient/CO2 removal efficiencies. These findings provide a theoretical basis for scale-up experiments using microalgae-based technologies.

Yak rumen fluid inoculum increases biogas production from sheep manure substrate.

Hydrolytic bacteria are essential for the degradation of lignocellulose to produce biogas and organic fertilizers. In this study, sheep manure was used as substrate, and sheep manure slurry, yak rumen fluid and slurry from a biogas reactor (SBR) were used as inocula in single-stage anaerobic digestion. The SBR and rumen fluid inocula increased biogas production by 23% and 43%, respectively, when compared to solely sheep manure in the single-stage anaerobic digestion. The two-stage anaerobic digestion, with yak rumen fluid as inoculum in the hydrolytic reactor, increased the biogas production by 59, 86, and 58% compared with the control. Microbial analysis of the effluent revealed that yak rumen fluid contained hydrolytic bacteria such as Proteiniphilum, Jeotgalibaca, Fermentimonas, and Atopostipes to enhance the degradation of sheep manure and increase biogas production. It was concluded that yak rumen fluid, rich in hydrolytic bacteria, increases the degradability of sheep manure and improves production of volatile fatt acids and biogas.

Confirming the presence of selected antibiotics and steroids in Norwegian biogas digestate.

Farms utilizing sewage sludge and manure in their agronomic plant production are recognized as potential hotspots for environmental release of antibiotics and the resulting promotion of antibiotic resistance. As part of the circular economy, the use of biogas digestates for soil fertilizing is steadily increasing, but their potential contribution to the spreading of pharmaceutical residues is largely unknown. Digestates can be produced from a variety of biowaste resources, including sewage sludge, manure, food waste, and fish ensilage. We developed a method for the detection of 17 antibiotics and 2 steroid hormones and applied the method to detect pharmaceutical residues in digestates from most municipal biogas plants in Norway, covering a variety of feedstocks. The detection frequency and measured levels were overall low for most compounds, except a few incidents which cause concern. Specifically, relatively high levels of amoxicillin, penicillin G, ciprofloxacin, and prednisolone were detected in different digestates. Further, ipronidazole was detected in four digestates, although no commercial pharmaceutical products containing ipronidazole are currently registered in Norway. A simplified risk assessment showed a high risk for soil microorganisms and indicates the tendency for antibiotic-resistant bacteria for penicillin G and amoxicillin. For prednisolone and ipronidazole; however, no toxicity data is available for reliable risk assessments.