A new microalgal negative carbon technology for landfill leachate treatment: Simultaneous removal of nitrogen and phosphorus.

Replete with ammonia nitrogen and organic pollutants, landfill leachate typically undergoes treatment employing expensive and carbon-intensive integrated techniques. We propose a novel microalgae technology for efficient, low-carbon simultaneous treatment of carbon, nitrogen, and phosphorus in landfill leachate (LL). The microbial composition comprises a mixed microalgae culture with Chlorella accounting for 82.58%. After seven days, the process with an N/P ratio of approximately 14:1 removed 98.81% of NH4+-N, 88.62 % of TN, and 99.55% of TP. Notably, the concentrations of NH4+-N and TP met the discharge standards, while the removal rate of NH4+-N was nearly three times higher than previously reported in relevant studies. The microalgae achieved a removal efficiency of 64.27% for Total Organic Carbon (TOC) and 99.26% for Inorganic Carbon (IC) under mixotrophic cultivation, yielding a biomass of 1.18 g/L. The treatment process employed in this study results in a carbon emissions equivalent of -8.25 kgCO2/kgNremoved, representing a reduction of 33.56 kgCO2 compared to the 2AO + MBR process. In addition, shake flask experiments were conducted to evaluate the biodegradability of leachate after microalgae treatment. After microalgae treatment, the TOCB (Biodegradable Total Organic Carbon)/TOC ratio decreased from 56.54% to 27.71%, with no significant improvement in biodegradability. It establishes a fundamental foundation for further applied research in microalgae treatment of leachate.

Enhancement of biomass productivity and biochemical composition of alkaliphilic microalgae by mixotrophic cultivation using cheese whey for biofuel production.

The growth of microalgae under alkaline conditions ensures an ample supply of CO2 from the atmosphere, with a low risk of crashing due to contamination and predators. The present study investigated the mixotrophic cultivation of two alkaliphilic microalgae (Tetradesmus obliquus and Cyanothece sp.) using cheese whey as an organic carbon source. The variation in cheese whey concentration (0.5-4.5% (v/v)), culture pH (7-11), and NaNO3 concentrations (0-2 gL-1) was evaluated using central composite design in response to biomass productivity and the contents of lipids, total proteins, and soluble carbohydrates. Both investigated microalgae effectively utilized cheese whey as an organic carbon source. The optimum conditions for simultaneously maximizing biomass and lipid productivity in T. obliquus were 3.5% (v/v) whey, pH 10.0, and 0.5 g L-1 NaNO3. Under these conditions, the biomass, lipid, soluble carbohydrate, and protein productivities were 48.69, 20.64, 7.02, and 10.97 mg L-1 day-1, respectively. Meanwhile, Cyanothece produced 52.78, 11.42, 4.31, and 7.89 mg L-1 day-1 of biomass, lipid, carbohydrate, and protein, respectively, at 4.5% (v/v) whey, pH 9.0, and 1.0 g L-1 NaNO3. The lipids produced under these conditions were rich in saturated fatty acids (FAs) and monounsaturated FAs, with no polyunsaturated FAs in both microalgae. Moreover, several biodiesel characteristics were estimated, and results fell within the ranges specified by international standards. These findings indicate that the mixotrophic cultivation of alkaliphilic microalgae could open new avenues for promoting microalgae productivity through low-cost biofuel production.

Assessment of hydrocarbon degradation capacity and kinetic modeling of Chlorella vulgaris and Scenedesmus quadricauda for crude oil phycoremediation under mixotrophic conditions.

Crude oil spills imperil aquatic ecosystems globally, prompting innovative solutions such as microalgae-based bioremediation. This study explores the potential of Chlorella vulgaris and Scenedesmus quadricauda, for crude oil spill phycoremediation under mixotrophic conditions and varying crude oil concentrations (0.5-2%). C. vulgaris demonstrated notable resilience, thriving up to 1% crude oil exposure, while S. quadricauda adapted to lower concentrations. Optimal growth for both was observed at 0.5% exposure. Chlorophyll a content in C. vulgaris increases at 0.5% exposure but declines above 1%, while a decline was noticeable in chlorophyll b in treatment groups above 1%. Carotenoid levels varied, displaying the highest levels at higher concentrations above 1.5%. Similarly, S. quadricauda showed increased chlorophyll a content at 0.5% exposure, with stable carotenoid levels and a decline in chlorophyll b content at higher concentrations. GC/MS analyses indicated C. vulgaris efficiently degraded aliphatic compounds like decane and tridecane, surpassing S. quadricauda in degrading both aliphatic and aromatic hydrocarbons. Growth kinetics was best represented by the modified Gompertz and logistic models. These findings highlight the species-specific adaptability and optimal concentration for microalgae to degrade crude oil effectively, advancing phycoremediation processes and strategies critical for environmental restoration.

This study marks the first exploration of both Chlorella vulgaris and the previously unexplored Scenedesmus quadricauda for crude oil phycoremediation potential under mixotrophic conditions. Additionally, it pioneers the modeling and study of algae growth kinetics in response to crude oil exposure. Notably, this research demonstrated the adaptability and efficiency of C. vulgaris in degrading crude oil components under mixotrophic conditions up to a level of 1%, while S. quadricauda showed similar capabilities at a concentration of 0.5%.

Microalgae-mediated biofixation as an innovative technology for flue gases towards carbon neutrality: A comprehensive review.

Microalgae-mediated industrial flue gas biofixation has been widely discussed as a clean alternative for greenhouse gas mitigation. Through photosynthetic processes, microalgae can fix carbon dioxide (CO2) and other compounds and can also be exploited to obtain high value-added products in a circular economy. One of the major limitations of this bioprocess is the high concentrations of CO2, sulfur oxides (SOx), and nitrogen oxides (NOx) in flue gases, according to the origin of the fuel, that can inhibit photosynthesis and reduce the process efficiency. To overcome these limitations, researchers have recently developed new technologies and enhanced process configurations, thereby increased productivity and CO2 removal rates. Overall, CO2 biofixation rates from flue gases by microalgae ranged from 72 mg L-1 d -1 to over 435 mg L-1 d-1, which were directly influenced by different factors, mainly the microalgae species and photobioreactor. Additionally, mixotrophic culture have shown potential in improving microalgae productivity. Progress in developing new reactor configurations, with pilot-scale implementations was observed, resulting in an increase in patents related to the subject and in the implementation of companies using combustion gases in microalgae culture. Advancements in microalgae-based green technologies for environmental impact mitigation have led to more efficient biotechnological processes and opened large-scale possibilities.

Mixotrophic aerobic denitrification facilitated by denitrifying bacterial-fungal communities assisted with iron in micro-polluted water: Performance, metabolic activity, functional genes abundance, and community co-occurrence.

Low-dosage nitrate pollutants can contribute to eutrophication in surface water bodies, such as lakes and reservoirs. This study employed assembled denitrifying bacterial-fungal communities as bio-denitrifiers, in combination with zero-valent iron (ZVI), to treat micro-polluted water. Immobilized bacterial-fungal mixed communities (IBFMC) reactors demonstrated their ability to reduce nitrate and organic carbon by over 43.2 % and 53.7 %, respectively. Compared to IBFMC reactors, IBFMC combined with ZVI (IBFMC@ZVI) reactors exhibited enhanced removal efficiencies for nitrate and organic carbon, reaching the highest of 31.55 % and 17.66 %, respectively. The presence of ZVI in the IBFMC@ZVI reactors stimulated various aspects of microbial activity, including the metabolic processes, electron transfer system activities, abundance of functional genes and enzymes, and diversity and richness of microbial communities. The contents of adenosine triphosphate and electron transfer system activities enhanced more than 5.6 and 1.43 folds in the IBFMC@ZVI reactors compared with IBFMC reactors. Furthermore, significant improvement of crucial genes and enzyme denitrification chains was observed in the IBFMC@ZVI reactors. Iron played a central role in enhancing microbial diversity and activity, and promoting the supply, and transfer of inorganic electron donors. This study presents an innovative approach for applying denitrifying bacterial-fungal communities combined with iron enhancing efficient denitrification in micro-polluted water.

Treatment of high ammonia anaerobically digested molasses wastewater using aerobic granular sludge reactor.

This study addressed the treatment of high ammonia, low biodegradable chemical oxygen demand (bCOD) anaerobically digested molasses wastewater, utilizing an aerobic granular sludge (AGS) reactor. The AGS achieved 99 % ammonia removal regardless of the bCOD supplementation. By adding low ammonia (<60 mg/L), high bCOD raw molasses wastewater (before anaerobic digestion) as a carbon source, enhanced nitrogen removal, increasing from 10 % to 97 %, and improved sludge settleability via bio-induced calcite precipitation were observed. Functional genes prediction suggested two potential denitrification pathways, including heterotrophic denitrification by Paracoccus and Thauera, and autotrophic denitrification, specifically sulfide-oxidizing autotrophic denitrification by Thiobacillus. An increase in the relative abundance of microorganisms involved in heterotrophic denitrification was observed with the addition of high bCOD raw molasses wastewater. Consequently, incorporating raw molasses wastewater into the AGS presents a sustainable approach to achieve mixotrophic denitrification, maintain stable granular sludge and ensure stable treatment performance when treating anaerobically digested molasses wastewater.

Partial nitrification and simultaneous denitrification in sequential anaerobic and aerobic reactors: performance and microbial community dynamics.

ABSTRACTThe removal of organic matter and nitrogen from domestic sewage was evaluated using a system composed of two sequential reactors: an anaerobic reactor (ANR) with suspended sludge and an aerobic (AER) reactor with suspended and adhered sludge to polyurethane foams. Nitrogen removal consisted of AER operating at low dissolved oxygen (DO) concentrations; this favoured the simultaneous nitrification and denitrification (SND) process. The concentration of COD and N were 440 mgO2.L-1 and 37 mgTN.L-1, respectively. The operation was divided into three phases (P), lasting 51, 53, and 46 days, respectively. The initial DO concentrations applied in the AER were: 3.0 (PI) and 1.5 mg.L-1 (PII and PIII). In PIII, the AER effluent was recirculated to the ANR at a ratio of 0.25. Kinetic assays were performed to determine the nitrification and denitrification rates of the biomasses (ANR and AER in PIII). Changes in the microbial community were evaluated throughout phases PI to PIII by massive sequencing. In PIII, the best results obtained for chemical oxygen demand (COD) and total nitrogen (TN-N) removal efficiencies, were close to 94% and 65%, respectively. Under these conditions, system effluent concentrations below 30 mg COD.L-1 and 15 mg TN-N.L-1 were verified. The nitritation and nitration rates were 10.5 and 6.5 mg N.g VSS-1.h-1, while the denitrification via nitrite and nitrate were 6.8 and 5.8 mg N.g VSS-1.h-1, respectively. A mixotrophic community was prevalent, with Rhodococcus, Nitrosomonas, Pseudomnas, and Porphyromonas being dominant or co-dominant in most of the samples, confirming the SND process in the AER sludge.

Mixotrophic denitrification of waste activated sludge fermentation liquid as an alternative carbon source for nitrogen removal: Reducing N2O emissions and costs.

Heterotrophic-sulfur autotrophic denitrification (HAD) has been proposed to be a prospective nitrogen removal process. In this work, the potential of fermentation liquid (FL) from waste-activated sludge (WAS) as the electron donor for denitrification in the HAD system was explored and compared with other conventional carbon sources. Results showed that when FL was used as a carbon source, over 99% of NO3--N was removed and its removal rate exceeded 14.00 mg N/g MLSS/h, which was significantly higher than that of methanol and propionic acid. The produced sulfate was below the limit value and the emission of N2O was low (1.38% of the NO3--N). Microbial community analysis showed that autotrophic denitrifiers were predominated in the HAD system, in which Thiobacillus (16.4%) was the dominant genus. The economic analysis showed the cost of the FL was 0.062 €/m3, which was 30% lower than that in the group dosed with methanol. Our results demonstrated the FL was a promising carbon source for the HAD system, which could reduce carbon emission and cost, and offer a creative approach for waste-activated sludge resource reuse.

Significantly Enhanced Nitrate and Phosphorus Removal by Pyrite/Sawdust Composite-Driven Mixotrophic Denitrification with Boosted Electron Transfer: Comprehensive Evaluation of Water-Gas-Biofilm Phases during a Long-Term Study.

Further reducing total nitrogen (TN) and total phosphorus (TP) in the secondary effluent needs to be realized effectively and in an eco-friendly manner. Herein, four pyrite/sawdust composite-based biofilters were established to treat simulated secondary effluent for 304 days. The results demonstrated that effluent TN and TP concentrations from biofilters under the optimal hydraulic retention time (HRT) of 3.5 h were stable at <2.0 and 0.1 mg/L, respectively, and no significant differences were observed between inoculated sludge sources. The pyrite/sawdust composite-based biofilters had low N2O, CH4, and CO2 emissions, and the effluent's DOM was mainly composed of five fluorescence components. Moreover, mixotrophic denitrifiers (Thiothrix) and sulfate-reducing bacteria (Desulfosporosinus) contributing to microbial nitrogen and sulfur cycles were enriched in the biofilm. Co-occurrence network analysis deciphered that Chlorobaculum and Desulfobacterales were key genera, which formed an obvious sulfur cycle process that strengthened the denitrification capacity. The higher abundances of genes encoding extracellular electron transport (EET) chains/mediators revealed that pyrite not only functioned as an electron conduit to stimulate direct interspecies electron transfer by flagella but also facilitated EET-associated enzymes for denitrification. This study comprehensively evaluates the water-gas-biofilm phases of pyrite/sawdust composite-based biofilters during a long-term study, providing an in-depth understanding of boosted electron transfer in pyrite-based mixotrophic denitrification systems.

Bioreactor Expansion Affects Microbial Succession of Mixotrophic Acidophiles and Bioremediation of Cadmium-Contaminated Soils.

Microbial scale-up cultivation is the first step to bioremediating cadmium (Cd)-contaminated soils at the industrial scale. However, the changes in the microbial community as the bioreactor volume expands and their associations with soil Cd removal remain unclear. Herein, a six-stage scale-up cultivation process of mixotrophic acidophiles was conducted, scaling from 0.1 L to 10 m3, to remediate Cd-contaminated soils. The findings showed that bioreactor expansion led to a delay in sulfur and glucose oxidations, resulting in a reduced decline in solution pH and cell density. There were minimal differences observed in bacterial alpha-diversity and community structure as the bioreactor volume increased, except for the 10 m3 scale. However, bioreactor expansion decreased fungal alpha-diversity, changed the community structure, and simplified fungal community compositions. At the family level, Acidithiobacillaceae and Debaryomycetaceae dominated the bacterial and fungal communities throughout the scale-up process, respectively. Correlation analysis indicated that the indirect effect of mixotrophic acidophiles played a significant role in soil Cd removal. Bacterial community shifts, driven by changes in bioreactor volume, decreased the pH value through sulfur oxidation, thereby indirectly enhancing Cd removal efficiency. This study will contribute to the potential industrial application of mixotrophic acidophiles in bioremediating Cd-contaminated soils.

Mixotrophic culture enhances fucoxanthin production in the haptophyte Pavlova gyrans.

Fucoxanthin is a versatile substance in the food and pharmaceutical industries owing to its excellent antioxidant and anti-obesity properties. Several microalgae, including the haptophyte Pavlova spp., can produce fucoxanthin and are potential industrial fucoxanthin producers, as they lack rigid cell walls, which facilitates fucoxanthin extraction. However, the commercial application of Pavlova spp. is limited owing to insufficient biomass production. In this study, we aimed to develop a mixotrophic cultivation method to increase biomass and fucoxanthin production in Pavlova gyrans OPMS 30543X. The effects of culturing OPMS 30543X with different organic carbon sources, glycerol concentrations, mixed-nutrient conditions, and light intensities on the consumption of organic carbon sources, biomass production, and fucoxanthin accumulation were analyzed. Several organic carbon sources, such as glycerol, glucose, sucrose, and acetate, were examined, revealing that glycerol was well-consumed by the microalgae. Biomass and fucoxanthin production by OPMS 30543X increased in the presence of 10 mM glycerol compared to that observed without glycerol. Metabolomic analysis revealed higher levels of the metabolites related to the glycolytic, Calvin-Benson-Bassham, and tricarboxylic acid cycles under mixotrophic conditions than under autotrophic conditions. Cultures grown under mixotrophic conditions with a light intensity of 100 µmol photons m-2 s-1 produced more fucoxanthin than autotrophic cultures. Notably, the amount of fucoxanthin produced (18.9 mg/L) was the highest reported thus far for Pavlova species. In conclusion, the use of mixotrophic culture is a promising strategy for increasing fucoxanthin production in Pavlova species. KEY POINTS: • Glycerol enhances biomass and fucoxanthin production in Pavlova gyrans • Metabolite levels increase under mixotrophic conditions • Mixotrophic conditions and medium-light intensity are appropriate for P. gyrans.

Effect of pyrite particle size on the denitrification performance of autotrophic or split-mixotrophic bioreactors supported by pyrite/polycaprolactone.

In this study, a pyrite-based autotrophic denitrification (PAD) system, a polycaprolactone (PCL)-supported heterotrophic denitrification (PHD) system, and a pyrite+PCL-based split-mixotrophic denitrification (PPMD) system were constructed. The pyrite particle size was controlled in 1-3, 3-5, or 5-8 mm in both the PAD and PPMD systems to investigate the effect of pyrite particle size on the denitrification performance of autotrophic or split-mixotrophic bioreactors. It was found that the PAD system achieved the best denitrification efficiency with an average removal rate of 98.98% in the treatment of 1- to 3-mm particle size, whereas it was only 19.24% in the treatment of 5- to 8-mm particle size. At different phases of the whole experiment, the nitrate removal rates of both the PHD and PPMD systems remained stable at a high level (>94%). Compared with the PAD or PHD system, the PPMD system reduced the concentrations of sulfate and chemical oxygen demand in the final effluent efficiently. The interconnection network diagram explained the intrinsic metabolic pathways of nitrogen, sulfur, and carbon in the three denitrification systems at different phases. In addition, the microbial community analysis showed that the PPMD system was beneficial for the enrichment of Firmicutes. Finally, the impact mechanism of pyrite particle size on the performance of the PPMD system was proposed. PRACTITIONER POINTS: The reduction of pyrite particle size was beneficial for improving the efficiency of the PAD process. The change in particle size had an effect on NO2 --N accumulation in the PAD system. The accumulation of NH4 +-N in the PPMD system increased with the decrease in particle size. The reduction of pyrite particle size increased the production of SO4 2- in the PAD and PPMD systems. The correlations among the effluent indicators of the PAD and PPMD systems could be well explained.

Enhancing nitrogen removal in the partial denitrification/anammox processes for SO4- - Rich wastewater treatment: Insights into autotrophic and mixotrophic strategies.

The investigation of partial denitrification/anammox (PD/anammox) processes was conducted under autotrophic (N-S cycle) and mixotrophic (N-S-C cycle) conditions over 180 days. Key findings revealed the remarkable capability of SO42--dependent systems to produce NO2- effectively, supporting anaerobic NH4+ oxidation. Additionally, SO42- served as an additional electron acceptor in sulfate reduction ammonium oxidation (SRAO). Increasing influent SO42- concentrations notably improved ammonia utilization rates (AUR) and NH4+ and total nitrogen (TN) utilization efficiencies, peaking at 57% for SBR1 and nearly 100% for SBR2. Stoichiometric analysis showed a 7.5-fold increase in AUR (SRAO and anammox) in SBR1 following SO42- supplementation. However, the analysis for SBR2 indicated a shift towards SRAO and mixotrophic denitrification, with anammox disappearing entirely by the end of the study. Comparative assessments between SBR1 and SBR2 emphasized the impact of organic compounds (CH3COONa) on transformations within the N-S-C cycle. SBR1 performance primarily involved anammox, SRAO and other SO42- utilization pathways, with minimal S-dependent autotrophic denitrification (SDAD) involvement. In contrast, SBR2 performance encompassed SRAO, mixotrophic denitrification, and other pathways for SO42- production. The SRAO process involved two dominant genera, such as Candidatus Brocadia and PHOS-HE36.

A Response Surface Methodology Study for Chlorella vulgaris Mixotrophic Culture Optimization.

Glycerol is a carbon source that produces good biomass under mixotrophic conditions. Enhancing the composition of culture media in algae biomass production improves growth rates, biomass yield, nutrient utilization efficiency, and overall cost-effectiveness. Among the key nutrients in the medium, nitrogen plays a pivotal role. Urea can be effectively used as a nitrogen source and is considered a low-cost form of nitrogen compared to other sources. Urea metabolism releases some CO2 in photosynthesis, and magnesium plays a major role in urea uptake. Magnesium is another key nutrient that is key in photosynthesis and other metabolic reactions. To maximize glycerol consumption in the mixotrophic system and to obtain high biomass and lipid productions, the variations in MgSO4·7H2O and urea concentrations were evaluated in the growth medium of the microalgae. A response surface methodology (RSM) using a central composite design (CCD) was designed to maximize glycerol consumption at the initial cellular growth rates (up to four days). The magnesium and urea supply varied from 0.3 to 1.7 g L-1. Response surface methodology was utilized to analyze the results, and the highest glycerol consumption rate, 770.2 mg L-1 d-1, was observed when C. vulgaris was grown at 1.7 g L-1 urea, 1.0 g L-1 MgSO4·7H2O. Using the optimal urea and magnesium concentrations with acetate, glucose, and glycerol as carbon sources, the same lipid content (10% average) was achieved on day 4 of mixotrophic C. vulgaris culture. Overall, the results show that mixotrophic growth of C. vulgaris using urea with an optimum magnesium concentration yields large amounts of fatty acids and that the carbon source greatly influences the profile of the fatty acids.

A robust multinutrient kinetic model for enhanced lutein and biomass yields in mixotrophic microalgae cultivation: A step towards successful large-scale productions.

Mixotrophic cultivation holds great promise to significantly enhance the productivities of biomass and valuable metabolites from microalgae. In this study, a new kinetic model is developed, explicitly describing the effect of the most influential environmental factors on both biomass growth and the production of the high-value product lutein. This extensive study of multinutrient kinetics for Tetradesmus obliquus in a mixotrophic regime covers various nutritional conditions. Crucial nutrients governing the model include nitrate, phosphate, and glucose. Using seven state variables and 13 unknown parameters, the model's accuracy was ensured through a well-designed two-factor, four-level experimental setup, providing ample data for reliable calibration and validation. Results accurately predict dynamic concentration profiles for all validation experiments, revealing broad applicability. Optimizing nitrogen availability led to significant increases in biomass (up to fourfold) and lutein production (up to 12-fold), with observed maximum biomass concentration of 6.80 g L-1 and lutein reaching 25.58 mg L-1. Noticeably, the model exhibits a maximum specific growth rate of 4.03 day-1, surpassing reported values for photoautotrophic and heterotrophic conditions, suggesting synergistic effects. Valuable guidance is provided for applying the method to various microalgal species and results are large-scale production-ready. Future work will exploit these results to develop real-time photobioreactor operation strategies.

Exposure to the mixotrophic dinoflagellate Lepidodinium sp. and its cues increase toxin production of Pseudo-nitzschia multiseries.

The present study examined the defense responses of toxigenic Pseudo-nitzschia species (P. multiseries) to a mixotrophic dinoflagellate, Lepidodinium sp., and its associated cues. We evaluated their responses to different predation risks, including direct physical contact and indirect interactions facilitated by cues from Lepidodinium sp. during active feeding on heterospecific prey (Rhodonomas salina), limited feeding on conspecific prey (P. multiseries) and non-feeding (autotrophic growth in f/2 medium) states. This study is the first investigation of these trophic interactions. Our results demonstrated a significant increase in cellular domoic acid (cDA) in P. multiseries when exposed to Lepidodinium sp. and its associated cues, which was 1.38 to 2.42 times higher than the non-induced group. Notably, this increase was observed regardless of Lepidodinium sp. feeding on this toxic diatom and nutritional modes. However, the most significant increase occurred when they directly interacted. These findings suggest that P. multiseries evaluates predation risk and increases cDA production as a defensive strategy against potential grazing threats. No morphological changes were observed in P. multiseries in response to Lepidodinium sp. or its cues. P. multiseries cultured in flasks of Group L+P-P showed a decrease in growth, but Group L-P and Group L+R-P did not exhibit any decrease. These results suggest a lack of consistent trade-offs between the defense response and growth, thus an increase in cDA production may be a sustainable and efficient defense strategy for P. multiseries. Furthermore, our findings indicate that P. multiseries had no significant impact on the fitness (cell size, growth and/or grazing) of Lepidodinium sp. and R. salina, which suggests no evident toxic or allelopathic impacts on these two phytoplankton species. This study enhances our understanding of the trophic interactions between toxic diatoms and mixotrophic dinoflagellates and helps elucidate the dynamics of Harmful Algal Blooms, toxin transmission, and their impact on ecosystem health.

The role of the combined nitrogen-sulfur-carbon cycles for efficient performance of anammox-based systems.

The combined anammox/mixotrophic denitrification process was conducted in two granular sequencing batch reactors (SBRs) during a 200-day operation. Both reactors were fed with synthetic medium, but SBR2 was enriched with additional sulfate (SO42-) which influenced sulfate reduction ammonium oxidation (SRAO) and heterotrophic reduction of SO42- by sulfate reducing bacteria. It was hypothesized that the addition of SO42- could positively impact the removal rates of N-S-C compounds. A low C/N ratio (0.4-1.6) was maintained to prevent inhibition of anaerobic ammonium oxidizing bacteria (AnAOB), and alternating chemical oxygen demand (COD) on/off conditions were used to regenerate AnAOB during COD-off phases and heterotrophic denitrifiers during COD-on phases. Stoichiometric analysis showed that introducing SO42- in SBR2 enhanced the ammonium utilization rate, which was approximately 10 % higher compared to SBR1 in the final stage of the experiment (25.8 vs. 22.8 mg N/(g VSS·h)). The total nitrogen removal efficiencies ranged from 62 % to 99 % in both reactors, with SBR2 consistently exhibiting approximately 4 % higher efficiency than SBR1. In SBR2, the maximum overall SO42- utilization efficiency reached 27 % under COD-off conditions, while overall COD utilization was almost complete under COD-on conditions. A strong correlation (R2 = 0.98) was observed between SO42- production and COD utilization. The key players responsible for N and S transformations in response to SO42- addition were Candidatus Brocadia and Chloroflexi - Anaerolineae. This study highlights the potential to enhance the overall efficiency of N-S-C removal by implementing an integrated anammox/mixotrophic denitrification process. The combination of cycles emerges as a sustainable approach for treating wastewater rich in N-S-C compounds.

Deciphering the influencing mechanism of hydraulic retention time on purification performance of a mixotrophic system from the perspective of reaction kinetics.

At present, eutrophication is increasingly serious, so it is necessary to effectively reduce nitrogen and phosphorus in water bodies. In this study, a pyrite/polycaprolactone-based mixotrophic denitrification (PPMD) system using pyrite and polycaprolactone (PCL) as electron donors was developed and compared with pyrite-based autotrophic denitrification (PAD) system and PCL-based heterotrophic denitrification (PHD) system through continuous flow experiment. The removal efficiency of NO3--N (NRE) and PO43--P (PRE) and the contribution proportion of PAD in the PPMD system were significantly increased by prolonging hydraulic retention time (HRT, from 1 to 48 h). When HRT was equal to 24 h, the PPMD system conformed to the zero-order kinetic model, so NRE and PRE were mainly limited by the PAD process. When HRT was equal to 48 h, the PPMD system met the first-order kinetic model with NRE and PRE reaching 98.9 ± 1.1% and 91.8 ± 4.5%, respectively. When HRT = 48 h, the NRE and PRE by PAD system were 82.7 ± 9.1% and 88.5 ± 4.7%, respectively, but the effluent SO42- concentration was as high as 152.1 ± 13.7 mg/L (the influent SO42- concentration was 49.2 ± 3.3 mg/L); the NRE by PHD system was 98.5 ± 1.7%, but the PO43--P could not be removed ideally. The concentrations of NO3--N, total nitrogen, PO43--P, and SO42- in the PPMD system also showed distinct changes along the reactor column. In addition, the microbial diversity analysis showed that prolonging HRT (from 24 to 48 h) increased the abundance of autotrophic denitrifying microorganisms in the PPMD system, ultimately increasing the contribution proportion of PAD.

Third generation biobutanol production by Clostridium beijerinckii in a medium containing mixotrophically cultivated Dunaliella salina biomass.

This study aims the third generation biobutanol production in P2 medium supplemented D. salina biomass mixotrophically cultivated with marble waste (MW). The wastes derived from the marble industry contain approximately 90% of carbon-rich compounds. Microalgal growth in mixotrophic conditions was optimized in the 0.4-2 g/L of MW concentration range. The highest microalgal concentration was obtained as 0.481 g/L in the presence of 1 g/L MW. Furthermore, some important parameters for the production of biobutanol, such as microalgal cultivation conditions, initial mixotrophic microalgal biomass loading (50-300 g/L), and fermentation time (24-96 h) were optimized. The highest biobutanol, total ABE, biobutanol yield and productivity were determined as 11.88 g/L, 13.89 g/L, 0.331 g/g and 0.165 g/L/h at the end of 72 h in P2 medium including 60 g/L glucose and 200 g/L microalgal biomass cultivated in 1 g/L MW, respectively. The results show that D. salina is a suitable raw material for supporting Clostridium beijerinckii DSMZ 6422 cells on biobutanol production. To the best of our knowledge, this is the first study on the use of MW which is a promising feedstock on the mixotrophic cultivation of D. salina for biobutanol production.

Insight into the potential mechanism of bicarbonate assimilation promoted by mixotrophic in CO2 absorption and microalgae conversion system.

CO2 absorption-microalgae conversion (CAMC) system is a promising carbon capture and utilization technology. However, the use of HCO3- as a carbon source often led to a slower growth rate of microalgae, which also limited the application of CAMC system. In this study, the assimilation efficiency of HCO3- in CAMC system was improved through mixotrophic, and the potential mechanism was investigated. The HCO3- assimilation efficiency and biomass under mixotrophic were 34.79% and 31.76% higher than that of control. Mixotrophic increased chlorophyll and phycocyanin content, which were beneficial to capture more light energy. The content of ATP and NADPH reached 566.86 µmol/gprot and 672.86 nmol/mgprot, which increased by 31.83% and 27.67% compared to autotrophic. The activity of carbonic anhydrase and Rubisco increased by 18.52% and 22.08%, respectively. Transcriptome showed that genes related to photosynthetic and respiratory electron transport were up-regulated. The synergy of photophosphorylation and oxidative phosphorylation greatly improved energy metabolism efficiency, thus accelerating the assimilation of HCO3-. These results revealed a potential mechanism of promoting the HCO3- assimilation under mixotrophic, it also provided a guidance for using CAMC system to serve carbon neutrality.