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The main purpose of this research was to clarify the influence of the addition of iron (Fe) alone (0-100 mg/L) or 50 mg/L of Fe with 2 mg/L each of cobalt (Co), copper (Cu) and nickel (Ni) on the methanogenic activity of a mesophilic two-stage UASB system treating ethanol wastewater at a fixed chemical oxygen demand (COD) loading rate of 16 kg/m

To achieve advanced nitrogen removal from actual municipal sewage, a novel multistage anoxic/aerobic process with sludge regeneration zone (R-MAO) was developed. The reactor was used to treat actual domestic sewage and the nitrogen removal capacity of the sludge regeneration zone (R zone) was investigated during the long-term operation. The best performance was obtained at the R zone's Oxidation-Reduction Potential (ORP) of -50±30 mV and hydraulic residence times (HRT) of 1.2 hr. The average effluent COD, TN, NH

Soluble microbial products (SMPs), dissolved organic matter excreted by activated sludge, can interact with antibiotics in wastewater and natural water bodies. Interactions between SMPs and antibiotics can influence antibiotic migration, transformation, and toxicity but the mechanisms involved in such interactions are not fully understood. In this study, integrated spectroscopy approaches were used to investigate the mechanisms involved in interactions between SMPs and a representative antibiotic, trimethoprim (TMP), which has a low biodegradation rate and has been detected in wastewater. The results of liquid chromatography-organic carbon detection-organic nitrogen detection indicated that the SMPs used in the study contained 15% biopolymers and 28% humic-like substances (based on the total dissolved organic carbon concentration) so would have contained sites that could interact with TMP. A linear relationship of fluorescent intensities of tryptophan protein-like substances in SMP was observed (R

In this study, a modified continuous-flow nitrifying reactor was successfully operated for rapid cultivation of micro-granules and achieving robust nitritation. Results showed that sludge granulation with mean size of ca. 100 µm was achieved within three weeks by gradually increasing settling velocity-based selection pressure from 0.48 to 0.9 m/hr. Though Nitrospira like nitrite-oxidizing bacteria (NOB) were enriched in the micro-granules with a ratio between ammonia-oxidizing bacteria (AOB) and NOB of 5.7%/6.5% on day 21, fast nitritation was achieved within one-week by gradually increasing of influent ammonium concentration (from 50 to 200 mg/L). Maintaining ammonium in-excess was the key for repressing NOB in the micro-granules. Interestingly, when the influent ammonium concentration switched back to 50 mg/L still with the residual ammonium of 15-25 mg/L, the nitrite accumulation efficiency increased from 90% to 98%. Experimental results suggested that the NOB repression was intensified by both oxygen and nitrite unavailability in the inner layers of micro-granules. Unexpectedly, continuous operation with ammonium in excess resulted in overproduction of extracellular polysaccharides and overgrowth of some bacteria (e.g., Nitrosomonas, Arenimonas, and Flavobacterium), which deteriorated the micro-granule stability and drove the micro-granules aggregation into larger ones with irregular morphology. However, efficient nitritation was stably maintained with extremely high ammonium oxidation potential (50 mg/g VSS/hr) and nearly complete washout of NOB was obtained. This suggested that smooth and spherical granule was not a prerequisite for achieving NOB wash-out and maintaining effective nitritation in the granular reactor. Overall, the micro-granules exhibited a great practical potential for high-rate nitritation.

The growing interest in the anaerobic ammonium oxidizing (AMX) process in treating high nitrogen containing wastewaters and a comprehensive study into the granulation mechanism of these bacteria under diverse environmental conditions over the years have been unequal. To this effect, the distinctive differences in saline adapted AMX (S_AMX) and non-saline adapted AMX (NS_AMX) granules are presented in this study. It was observed that substrate utilisation profiles, granule formation mechanism, and pace towards granulation differed marginally for the two adaptation conditions. The different microbial dominant aggregation types aided in splitting the 471 days operated lab-scale SBRs into three distinct phases. In both reactors, phase III (granules dominant phase) showed the highest average nitrogen removal efficiency of 87.9% ± 4.8% and 85.6% ± 3.6% for the S_AMX and NS_AMX processes, respectively. The extracellular polymeric substances (EPS) quantity and major composition determined its role either as a binding agent in granulation or a survival mechanism in saline adaptation. It was also observed that granules of the S_AMX reactor were mostly loosely and less condensed aggregates of smaller sub-units and flocs while those of the NS_AMX reactor were compact agglomerates. The ionic gradient in saline enrichment led to an increased activity of the Na

The relatively poor settling characteristics of particles produced in moving bed biofilm reactor (MBBR) outline the importance of developing a fundamental understanding of the characterization and settleability of MBBR-produced solids. The influence of carrier geometric properties and different levels of biofilm thickness on biofilm characteristics, solids production, particle size distribution (PSD), and particle settling velocity distribution (PSVD) is evaluated in this study. The analytical ViCAs method is applied to the MBBR effluent to assess the distribution of particle settling velocities. This method is combined with microscopy imaging to relate particle size distribution to settling velocity. Three conventionally loaded MBBR systems are studied at a similar loading rate of 6.0 g/(m

Successful scale-up of Bacillus subtilis culture for poly-β-hydroxybutyrate (PHB) production was performed in 5-l stirred-tank reactor using batch, fed-batch, and two-stage culture strategies. The kinetics of biomass production, substrate consumption, and PHB production were established in the stirred tank bioreactor in all the studies. A mathematical model was developed to investigate the role of limiting substrate on overall culture metabolism. A fed-batch strategy was predicted on the basis of computer simulations, for maximum PHB production. This was performed by extrapolation of batch model for predicting the feeding rate and suitable time of feeding. Substrate inhibition was studied and the substrate inhibition terms were incorporated in the model. The maximum cell biomass concentration in batch culture (24 h) and fed-batch culture (30 h) was 1.79 ± 0.03 g/l on dry cell weight (DCW) basis and 1.66 ± 0.050 g/l on DCW basis and the corresponding PHB content was 68.71% and 85.54% of DCW, respectively. Glucose was found to be the major limiting nutrient during the bioreactor culture. A two-stage culture, where cells were first grown in stage I in LBG media containing excess carbon and thereafter in stage II in OM media, showed biomass production of 1.95 ± 0.045 g/l at 4 h and PHB production of 93.33% of DCW at 16 h. A 9% increase in growth and 25% increase in PHB yield were obtained using two-stage culture with computer-simulated feeding strategy in the 5 l reactor. Oxygen limitation was overcome in modified two-stage culture to obtain a PHB production of 98% at 30 h. KEY POINTS: • Polyhydroxybutyrate production was studied in a 5-l stirred-tank bioreactor using HPLC • Mathematical model-assisted fed-batch strategy was implemented in bioreactor • Two-stage fed-batch cultivation was implemented and PHB production was 93% of dry weight in Gram-positive bacteria.

In this work, the effect of activated carbon particles on the production of xylonic acid from xylose by Gluconobacter oxydans in a stirred tank bioreactor was investigated. The enhancement of the oxygen transfer coefficient by activated carbon particles was experimentally evaluated under different solids volume fractions, agitation and aeration rates conditions. The experimental conditions optimized by response surface methodology (agitation speed 800 rpm, aeration rate 7 L min

Extracellular production of target proteins simplifies downstream processing due to obsolete cell disruption. However, optimal combinations of a heterologous protein, suitable signal peptide and secretion host can currently not be predicted, resulting in large strain libraries that need to be tested. On the experimental side, this challenge can be tackled by miniaturization, parallelization and automation, which provide high-throughput screening data. These data need to be condensed into a candidate ranking for decision making to focus bioprocess development on the most promising candidates. We screened for Bacillus subtilis signal peptides mediating Sec secretion of two polyethylene terephthalate degrading enzymes (PETases), leaf-branch compost cutinase (LCC) and polyester hydrolase (PE-H) mutants, by Corynebacterium glutamicum. We developed a fully automated screening process and constructed an accompanying Bayesian statistical modeling framework, which we applied in screenings for highest activity in 4-nitrophenyl palmitate degradation. In contrast to classical evaluation methods, batch effects and biological errors are taken into account and their uncertainty is quantified. Within only two rounds of screening, the most suitable signal peptide was identified for each PETase. Results from LCC secretion in microliter-scale cultivation were shown to be scalable to laboratory-scale bioreactors. This work demonstrates an experiment-modeling loop that can accelerate early-stage screening in a way that experimental capacities are focused to the most promising strain candidates. Combined with high-throughput cloning, this paves the way for using large strain libraries of several hundreds of strains in a Design-Build-Test-Learn approach.

The field of nanomedicine-catalyzed tumor therapy has achieved a lot of progress; however, overcoming the limitations of the tumor microenvironment (TME) to achieve the desired therapeutic effect remains a major challenge. In this study, a nanocomposite hydrogel (GH@LDO) platform combining the nanozyme CoMnFe-layered double oxides (CoMnFe-LDO) and natural enzyme glucose oxidase (GOX) was engineered to remodel the TME to enhance tumor catalytic therapy. The CoMnFe-LDO is a nanozyme that can convert endogenous H

Biohydrogen production using renewable sources has been regarded as one of the most sustainable ways to develop low-cost and green production technology. In order to achieve this objective, herein biohydrogen production has been conducted using the combination of untreated secondary sewage sludge (Sss), algal biomass hydrolyzate (Abh), graphene oxide (GO) and bacterial consortia that forms a granular system. Thus, naturally formed granular system produced cumulative H

Respirogram technology has been widely applied for aerobic process, however, the response of respirogram to anoxic denitrification is still unclear. To reveal such response may help to design a new method for the evaluation of the performance of denitrification. The size distribution of flocs measured at different denitrification moments demonstrated a clear expansion of flocs triggered by denitrification, during which higher specific endogenous and quasi-endogenous respiration rates (SOUR

This article describes fabrication of a customizable bioreactor, which comprises a perfusion system and coverslip-based tissue culture chamber that allow centimeter-scale vascularized or otherwise canalized tissue constructs to be maintained in weeks long static and/or perfusion culture at an exceptionally low cost, with intermittent live imaging and media sampling capabilities. The perfusion system includes a reusable polydimethylsiloxane (PDMS) lid generated from a three-dimensional (3D)-printed poly-lactic acid (PLA) mold and several lengths of perfusion tubing. The coverslip tissue culture chamber includes PDMS components built with 3D-printed PLA molds, as well as 3D-printed PLA frames and glass coverslips that house perfusable hydrogel constructs. As proof of concept, we fabricated a vascularized hydrogel construct, which was subjected to static and perfusion tissue culture, as well as flow studies using fluorescent beads and widefield fluorescent microscopy. This system can be readily reproduced, promoting the advancement of tissue engineering and regenerative medicine research.

As an ecofriendly photocatalytic antifouling technology for membrane bioreactors (MBRs), photocatalytic optical fibers (POFs) can decrease the replacement cost of modified membranes and prevent the proliferation of photosynthetic bacteria caused by direct light illumination. Here, POFs were applied in situ in an anaerobic ammonium oxidation (anammox) MBR for membrane biofouling control. Compared with the control MBR without POFs treatment, the average fouling cycle of the POFs-loaded MBR was extended by 137%, and the energy consumption caused by membrane fouling was saved by 18%. In the antibiofouling process

Human epidermal growth factor (hEGF) has multiple biological functions, such as promoting cell proliferation, differentiation, and migration. In addition, it is a very expensive polypeptide with attractive market prospects. However, the production of hEGF needs for high cost to manufacture polypeptide demands reinvestigations of process conditions so as to enhance economic benefits. Improving the expression of soluble hEGF is the fundamental method to reduce the cost. In this study, a non-extracellular engineered strain of expressed hEGF was constructed, using plasmid pET-22b(+) in Escherichia coli. Preliminary fermentation and high cell density cultivation were carried out in shake flasks and in a 5 L bioreactor, respectively. A high yield of 98 ± 10 mg/L of soluble hEGF and a dry cell weight (DCW) of 6.98 ± 0.3 g/L were achieved in shake flasks. Then, fermentation conditions were optimized for large-scale production, while taking into consideration the expensive equipment required for cooling and conforming to industrial standards. A yield of 285 ± 10 mg/L of soluble hEGF, a final cell density of 57.4 ± 2 g/L DCW (OD

Anaerobic digestion (AD) is the most widely used technology for organic matter treatment. However, multiple types of research have reported on improving the process because different operation inhibition factors and limitations affect the performance of AD process. Owing to the increasing use of iron-nanoparticles (Fe-NP) on AD, this review addresses the knowledge gaps and summarizes the finding from academic articles based on (i) the AD upgrading operations: limitations and upgrade techniques, (ii) Fe-NPs mechanisms on AD, (iii) Fe-NP effect on microbial communities associated to AD systems, and (iv) perspectives. The selected topics give the Fe-NP positive effects on the AD methane-production process in terms of gas production, effluent quality, and process optimization. The main results of this work indicate that (i) Fe-NP addition can be adapted among different feedstocks and complement other pretreatments, (ii) Fe-NP physicochemical characteristics enhance biogas production via direct interspecies electron transfer (DIET) mechanisms, and Fe-ion release due to their structure and their conductivity capability, and (iii) syntrophic bacteria and acetoclastic methanogens have been reported as the communities that better uptake Fe-NPs on their metabolisms. Finally, our research perspectives and gaps will be discussed to contribute to our knowledge of using Fe-NPs on AD systems.

Bacteria–algae consortia in the light bring the benefit of O2 production and CO2 reduction for wastewater treatment, while the bottleneck for application is how it behaves in the dark. In this study, inoculum ratio and sludge retention time (SRT) affected nutrient removal rather than chemical oxygen demand (COD) removal. Dark conditions (with a sludge/Chlorella inoculum ratio of 1:2 at a SRT of 15 d) achieved comparable performance to those of light conditions, due to bacteria contribution and mechanical aeration. Compared with light conditions, the ratio of Chla/Chlb decreased and Caro/(Chla + Chlb) increased to response oxidative stress. In the dark, algae were associated with Nitrosomonas and Dechloromonas. Flavobacterium disassociated with Chlorella in the dark but associated with Chlorella in the light. Moreover, nitritation genes (amo and Hao) and denitrifying gene (narH) were up-regulated, while P metabolism genes (PPX and PPK) were down-regulated. It is proposed to enrich Nitrosomonas in the night and denitrify polyphosphate accumulating organisms (DPAO) in the daytime to establish short-cut nitrification and denitrifying phosphorus removal in practical applications.

Significant progress in understanding the key enzymes or species of anammox has been made; however, the nitrogen removal mechanism in complex coupling systems centered on anammox remains limited. In this study, by the combination of metagenomics-metatranscriptomics analyses, the nitrogen removal in the anammox-centered coupling system that entails partial denitrification (PD) and hydrolytic acidification (HA, A-PDHA) was elucidated to be the nitrogen transformation driven by the electron generation-transport-consumption process. The results showed that a total nitrogen (TN) removal efficiency of98%, with a TN effluence of1 mg/L and a TN removal contribution via anammox of98%, was achieved after 59 days under famine operation and alkaline conditions during the start-up process. Further investigation confirmed that famine operation promoted the activity of genes responsible for electron generation in anammox, and increased the abundance or expression of genes related to electron consumption. Alkaline conditions enhanced the electron generation for PD by upregulating the activity of glyceraldehyde 3-phosphate dehydrogenase and strengthened electron transfer by increasing the gene encoding quinone pool. Altogether, these variations in the electron flow led to efficient nitrogen removal. These results improve our understanding of the nitrogen removal mechanism and application of the anammox-centered coupling systems in treating nitrogen wastewater.

Role of biochar in promoting methanogenesis during anaerobic processes was investigated in this research. Biochar produced from Himalayan pine needles was used as medium for conductive material mediated interspecies electron transfer (CM-IET) amongst the electron producing microorganisms and electron consuming methanogenic archaea. Three anaerobic continuous stirrer tank reactors (CSTRs) with 0, 5 and 10 g/L pine needle biochar (PNB) were operated at steady state organic loading rate (OLR) of 2.0–2.5 kgCOD/(m3.d). R0 (0 g/L PNB), representing indirect interspecies electron transfer (IIET), failed at an OLR of 2.0 kgCOD/(m3.d) due to the highest volatile fatty acid (VFA) concentration of 6,300 mg/L among the three CSTRs. On the other hand, at an OLR of 2.5 kgCOD/(m3.d), R2 (10 g/L PNB) showed the most superior performance with chemical oxygen demand (COD) removal of 55% and volatile fatty acid (VFA) concentration of 3,500 mg/L, while R1 (5 g/L PNB) recorded COD removal of 45% and VFA concentration of 4,400 mg/L. In comparison, fixed biofilm reactor (FBR) with 80 g/L of PNB as support material operated satisfactorily at OLR of 13.8 kgCOD/(m3.d) with 70% COD removal and VFA concentration of 1,400 mg/L. These investigations confirmed the beneficial role of biochar in anaerobic processes by promoting CM-IET amongst VFA degrading bacteria and methane producing archaea.

Cells grown in chemically defined medium are sensitive to shear force, potentially resulting in decreased cell growth. We optimized the perfusion process for HEK293 cell-based recombinant adenovirus-vectored zoster vaccine (Ad-HER) production with chemically defined medium.We first studied the pseudo-continuous strategies in shake flasks as a mimic of the bioreactor equipped with perfusion systems. Using design of experiment (DoE) in shake flasks, we obtained the regression models between Ad-HER titer/virus input-output ratio and three production process parameters: time of infection (TOI), multiplicity of infection (MOI), and virus production pH (pH). We then confirmed the effect of Pluronic F68 (PF-68) at 3.0 g/L on HEK293 cell growth and Ad-HER production in shake flasks and a 2 L benchtop bioreactor.The optimized process was scale-up to a 2 L benchtop bioreactor with the PATFP perfusion system, which yielded cell density of 7.4 × 10This optimization strategy could be used to develop a robust process with stable cell culture performance and adenovirus titer. Increasing PF-68 concentration in chemically defined medium could protect cells from shear stress generated by perfusion system.

Microbial chain elongation is a newly developed carboxylate platform-based bioprocess, which often encounters high salinity stress due to saline feedstock and pH adjustment. In this study, we systematically investigated the effects of salt types (Na

In order to investigate the cause of accumulation of the inorganic suspended solid (ISS) in biochemical tank for wastewater treatment plant (WWTP) in recent years, the influent quality of one WWTP in Chongqing was monitored in one year, and the removal efficiency of ISS during the pretreatment process was studied. Results showed that the low removal efficiency of ISS (

Over the last decades, the success of advanced cell therapies and the increasing production volumes of vaccines, proteins, or viral vectors have raised the need of robust cell-based manufacturing processes for ensuring product quality and satisfying good manufacturing practice requirements. The cultivation process of cells needs to be highly controlled for improved productivity, reduced variability, and optimized bioprocesses. Cell cultures can be easily monitored using different technologies, which could deliver direct or indirect assessment of the cells' viability. Among these techniques, nuclear magnetic resonance (NMR) spectroscopy is a powerful technology that permits the evaluation and the identification of key endogenous metabolites. NMR can provide information on the cell metabolic pathways, on the bioprocesses, and is also capable to quickly test for impurities. In this study, NMR was successfully used as a technology for monitoring cell viability and expansion in different supports for cell growth (including bioreactors), to predict the bioprocess output and for the early identification of key metabolites linked to cell starvation. This investigation will allow the timely control of culture conditions and favor the optimization of the bioprocesses.

Dosing powdered activated carbon (PAC) has been proven to be an economical and effective method to mitigate membrane fouling. However, the effects of pretreated PAC with different redox properties on membrane fouling still need to be further investigated. Here, the impact of commercial PAC, oxidized-PAC, and reduced-PAC on membrane fouling was investigated in membrane bioreactors (MBRs). Surprisingly, the filtration cycles were extended from 12-36 h to 132-156 h only by dosing reduced-PAC and commercial PAC with a finial dosage of 3 g/L, which were provided with reductive properties. However, few improvements of filtration cycle (less than 50 h) were achieved by dosing oxidized-PAC in the same dosage, which had the same adsorption performance as reduced-PAC and commercial PAC. The biomass and foulant concentration suggested that the enhanced anti-fouling performances by PAC with reductive properties were mainly attributed to the reduction of extracellular polymer substances (EPS) and soluble microbial products (SMP) content in the bulk solutions after 14 days of continuous operation. The model foulant degradation tests and the confocal laser scanning microscope (CLSM) images of activated sludge further demonstrated that PAC with reductive properties directly affected the microbial activities by controlling the EPS and SMP concentrations in the bulk solution, thereby suppressing membrane fouling. Such a finding provides new insights into anti-fouling mechanisms that the redox properties of PAC played a decisive role in membrane fouling mitigation, and also provides a strategy to prolong the anti-fouling effects by restoring the reductive properties of PAC. KEY POINTS: • The anti-fouling mechanisms of PAC with reductive property were investigated. • Reductive property was the main reason for fouling control instead of adsorption. • PAC with reductive property hindered the sludge activity to produce fewer foulants.

In microbiology, cultivation is a central approach for uncovering novel physiology, ecology, and evolution of microorganisms, but conventional methods have left many microorganisms found in nature uncultured. To overcome the limitations of traditional methods and culture indigenous microorganisms, we applied a two-stage approach: enrichment/activation of indigenous organisms by using a continuous-flow down-flow hanging sponge bioreactor and subsequent selective batch cultivation. Here, we provide a protocol for this bioreactor-mediated technique using activation of deep marine sediment microorganisms and downstream isolation of a syntrophic co-culture containing an archaeon closely related to the eukaryote ancestor (Candidatus Promethearchaeum syntrophicum strain MK-D1) as an example. Both stages can easily be tailored to target other environments and organisms by modifying the inoculum, feed solution/gases, attachment material and/or cultivation media. We anaerobically incubate polyurethane sponges inoculated with deep-sea methane seep sediment in a reactor at 10 °C and feed anaerobic artificial seawater medium and methane. Once phylogenetically diverse and metabolically active microorganisms are adapted to synthetic conditions in the reactor, we transition to growing community samples in glass tubes with the above medium, simple substrates and selective compounds (e.g., antibiotics). To accommodate for the slow growth anticipated for target organisms, primary cultures can be incubated for ≥6-12 months and analyzed for community composition even when no cell turbidity is observed. One casamino acid- and antibiotic-amended culture prepared in this way led to the enrichment of uncultured archaea. Through successive transfer in vitro combined with molecular growth monitoring, we successfully obtained the target archaeon with its partner methanogen as a pure syntrophic co-culture.

Biobutanol produced in acetone-butanol-ethanol fermentation at batch mode cannot compete with chemically derived butanol because of the low reactor productivity. Continuous fermentation can dramatically enhance productivity and lower capital and operating costs but are rarely used in industrial fermentation because of increased risks in culture degeneration, cell washout, and contamination. In this study, cells of the asporogenous Clostridium acetobutylicum ATCC55025 were immobilized in a single-pass fibrous-bed bioreactor (FBB) for continuous production of butanol from glucose and butyrate at various dilution rates. Butyric acid in the feed medium helped maintaining cells in the solventogenic phase for stable continuous butanol production. At the dilution rate of 1.88 h , butanol was produced at 9.55 g/L with a yield of 0.24 g/g and productivity of 16.8 g/L∙h, which was the highest ever achieved for biobutanol fermentation and an 80-fold improvement over the conventional ABE fermentation. The extremely high productivity was attributed to the high density of viable cells (~100 g/L at >70% viability) immobilized in the fibrous matrix, which also enabled the cells to better tolerate butanol and butyric acid. The FBB was stable for continuous operation for an extended period of over one month.

Veterinary antibiotics in swine wastewater has drawn great public attention. The removal processes of sulfamethizole (SMZ), enrofloxacin (ENR) and chlortetracycline (CTC) were investigated in the high-rate anaerobic process. The continuous experiments demonstrated that in 3 L working volume and with the organic loading rate 5 kg/(m3·d) rised to 20 kg/(m3·d), the average removal efficiencies of the high-rate anaerobic bioreactor for SMZ, ENR and CTC were 0, 54 and 100%, respectively. By using fixed-bed adsorption models, the saturation time of SMZ, ENR and CTC were 4 hydraulic retention time (HRT) (24 h), 8 HRT (48 h) and 372 HRT (2,232 h). In the batch experiments, the adsorption and biodegradation characteristics of anaerobic granular sludge were determined. In the high-rate anaerobic bioreactor, SMZ removal process mainly relied on the adsorption but it was very weak; ENR removal process was based on the adsorption and biodegradation; CTC removal process was based to a large extent on the adsorption because of the big capacity of AnGS. These results were helpful to create a rational basis for designing more suitable treatment systems as feasible barriers to the release of antibiotics into the environment.

As an alternative for commercial enzyme, crude enzyme of fungal mash could promote food waste (FW) hydrolysis, but its specific effects coupled pH adjusting on the production of volatile fatty acids (VFAs) remains unknown. The crude enzyme produced from an Aspergillus awamori, named complex-amylase (CA), was added to short-term anaerobic system of FW fermentation. Results showed that adding CA significantly improved the solubility and degradability of biodegradable and non-biodegradable organics in FW, where the SCOD concentration with adding CA increased by 116.9% relative to the control but a marginal enhancement on VFAs yield. In contrast, adding CA combined with adjusting pH 8 markedly increased the VFAs production to 32.0 g COD/L, almost 10 times as much as the control. Besides, pH adjusting altered the metabolic pathway from lactate-type to butyrate-type. Adding CA coupled pH adjusting significant increase the component of butyrate compared with pH adjusting alone. Moreover, microbial community analysis indicated that adding CA reinforced proportion of the butyrate-producing bacteria (e.g., Dialister) under basic conditions, thus enhancing the butyrate metabolic pathways. This study demonstrated that fungal mash pretreatment coupled pH conditioning could be an economical way to enhance VFAs yield for FW valorization during anaerobic fermentation.

The demand for cheap, healthy, and sustainable alternative protein sources has turned research interest into microbial proteins. Mycoproteins prevail due to their quite balanced amino acid profile, low carbon footprint and high sustainability potential. The goal of this research was to investigate the capability of Pleurotus ostreatus to metabolize the main sugars of agro-industrial side streams, such as aspen wood chips hydrolysate, to produce high-value protein with low cost. Our results indicate that P. ostreatus LGAM 1123 could be cultivated both in a C-6 (glucose)- and C-5(xylose)-sugar-containing medium for mycoprotein production. A mixture of glucose and xylose was found to be ideal for biomass production with high protein content and rich amino acid profile. P. ostreatus LGAM 1123 cultivation in a 4 L stirred-tank bioreactor using aspen hydrolysate was achieved with 25.0 ± 3.4 g L−1 biomass production, 1.8 ± 0.4 d−1 specific growth rate and a protein yield of 54.5 ± 0.5% (g/100 g sugars). PCA analysis of the amino acids revealed a strong correlation between the amino acid composition of the protein produced and the ratios of glucose and xylose in the culture medium. The production of high-nutrient mycoprotein by submerged fermentation of the edible fungus P. ostreatus using agro-industrial hydrolysates is a promising bioprocess in the food and feed industry.

Fe

The rate and extent of microbial polyhydroxyalkanoates (PHAs) production rely on the availability of substrates, growth of microbial biomass, and intracellular accumulation of polymer under nitrogen-limited conditions. The dynamics of PHAs production captured through various structured or unstructured models can be extended to design an optimal feeding strategy for process intensification. Large variability in process assumptions, choices of kinetics, and model complexity is expected depending on substrate(s), microbial metabolism, and discretization of the process under consideration. This communication attempts to review the estimation of stoichiometric yield coefficients, metabolic modelling, and choices of unstructured kinetics in microbial PHA production. Implementational irregularities in parameter estimation and quality check in modelling exercises have also been reviewed. It is observed that the scope of the majority of the "modelling" studies is confined to the estimation of stoichiometric parameters with limited utility. In dynamic models, microbial growth is often described using either Monod or logistic variants, while PHAs production adopts a Luedeking-Piret expression with or without substrate inhibition. Though model selection, regression with experimental data, parameter estimation, and model validation are integral parts of the exercise, very few provide sufficient coverage on all those aspects. Application of the model to control or optimize the bioprocess has rarely been attempted.

Mesenchymal stromal cells (MSC) are a promising platform for regenerative medicine applications because of their multilineage differentiation abilities and ease of collection, isolation, and growth ex vivo. To meet the demand for clinical applications, large-scale manufacturing will be required using three-dimensional culture platforms in vessels such as stirred suspension bioreactors. As MSCs are an adherent cell type, microcarriers are added to the culture to increase the available surface area for attachment and growth. Although extensive research has been performed on efficiently culturing MSCs using microcarriers, challenges persist in downstream processing, including harvesting, filtration, and volume reduction, which all play a critical role in the translation of cell therapies to the clinic. The objective of this review is to assess the current state of downstream technologies available for microcarrier-based MSC cultures. This includes a review of current research within the three stages: harvesting, filtration, and volume reduction. Using this information, a downstream process for MSCs is proposed, which can be applied to a wide range of applications.

In the context of regenerative endodontics research with the development of biomaterials, this work aimed to develop and test a prototype biomimetic bioreactor of a human tooth. The bioreactor was designed to reproduce a shaped dental canal connected with a cavity reproducing the periapical region and irrigated through two fluidic channels intended to reproduce the apical residual vascular supply. A test biomaterial composed of polylactic acid/polycaprolactone-tannic acid (PLA/PCL-TA) was produced by electrospinning/electrospraying and calibrated to be inserted in a dental canal. This biomaterial was first used to evaluate its imbibition capacity and the oximetry inside the bioreactor. Then, Dental Pulp Stem Cells (DPSCs) were cultured on PLA/PCL-TA cones for 1-3 weeks in the bioreactor; afterward cell adhesion, proliferation, and migration were histologically assessed. Complete imbibition biomaterial was obtained in 10 min and oximetry was stable over time. In the bioreactor, DPSCs were able to adhere, proliferate and migrate onto the surface and inside the biomaterial. In conclusion, this bioreactor was used successfully to test a biomaterial intended to support pulp regeneration and constitutes a new in vitro experimental model closer to clinical reality.

The article aimed to investigate performance and evaluation of combining chemical coagulation pretreatment unit with a pilot-scale fourth generation downflow hanging sponge (DHS-4G) for slaughterhouse wastewater (SWW) treatment. This combination of compact treatment technique was tested as a novel system for the treatment of SWW. A DHS-4G reactor with a capacity of about 100 liters was installed after a 1 m

Directly integrating anammox into sewage treatment is attractive, but anammox bacteria (AnAOB) enrichment is complex due to vicious competition from heterotrophic bacteria (HB). A novel strategy of optimal organics management using a preanaerobic stage and subsequent limited-oxygen conditions (0.32 ± 0.15 mg-O

Microbial consortia constitute a promising tool for achieving high-value chemical bio-production. However, customizing the consortium ratio remains challenging. Herein, an asymmetry distribution-based synthetic consortium (ADSC) was developed to switch cell phenotypes using shikimate synthesis for proof of concept. First, the cell pole-organizing protein PopZ was screened as a mediator of asymmetric protein distribution in Escherichia coli. The ADSC was then constructed to incorporate PopZ-mediated asymmetry distribution and a TetR-based transcription repression switch to achieve the dynamical control of microbial population ratio. Finally, the ADSC was used to decouple cell growth from shikimate synthesis by effectively coordinating the ratio of growing cells and production cells at the consortium level, thereby increasing shikimate titer to 30.1 g/L in the 7.5-L bioreactor with a minimal medium. This titer was further improved to 82.5 g/L when using rich medium fermentation. Our results illustrate a novel approach to control consortium structure through ADSC-mediated regulation, highlighting its potential as an efficient strategy for controlling metabolic state in microbes.

Nitrous oxide (N

To explore the effect of low temperature on the anaerobic digestion of pig manure, the anaerobic digestion experiment was carried out under the conditions of inoculum concentration of 30% and TS of 8%. Five low-temperature gradients of 4, 8, 12, 16 and 20 °C were set to study the activities of gas production, pH, solluted chemical oxygen demand (SCOD), volatile fatty acids (VFAs), coenzymes F420 and archaea community composition in the digestion process. The results were demonstrated: as the temperature decreased, the more unstable the gas production became, the less gas production produced, and the later the gas peak occurred. There were no significant peaks at either 4 °C or 8 °C, and the SCOD was unstable over time. From 12 °C, the SCOD increased over time, and the higher the temperature, the faster the growth trend. The pH was always greater than 7.6. 8, 12, 16, 20 °C had different degrees of VFAs accumulation at the late digestion stage. The higher the temperature, the greater the amount of volatile acid accumulation. When the VFAs of each reactor reached the maximum, the proportion of acetic acid also reached the highest. The digestion system of the five treatment groups was dominated by hydrogen-nutrient methanogenic pathway. The results could provide a further reference for the mechanism of anaerobic digestion of pig manure at low temperatures.

A new generation of bioreactors with integrated six degrees of freedom (6 DOF) aims to mimic more accurately the natural intervertebral disc (IVD) load. We developed and validated in a biological and mechanical study a specimen holder and corresponding ex vivo IVD organ model according to the bioreactor requirements for multiaxial loading and a long-term IVD culture. IVD height changes and cell viability were compared between the 6 DOF model and the standard 1 DOF model throughout the 3 weeks of cyclic compressive loading in the uniaxial bioreactor. Furthermore, the 6 DOF model and holder were loaded for 9 days in the multiaxial bioreactor under development using the same conditions, and the IVDs were evaluated for cell viability. The interface of the IVD model and specimen holder, enhanced with fixation screws onto the bone, was tested in compression, torsion, lateral bending, and tension. Additionally, critical motions such as tension and bending were assessed for a combination of side screws and top screws or side screws and adhesive. The 6 DOF model loaded in the uniaxial bioreactor maintained similar cell viability in the IVD regions as the 1 DOF model. The viability was high after 2 weeks throughout the whole IVD and reduced by more than 30% in the inner annulus fibrous after 3 weeks. Similarly, the IVDs remained highly viabile when cultured in the multiaxial bioreactor. In both models, IVD height changes after loading were in the range of typical physiological conditions. When differently directed motions were applied, the holder-IVD interface remained stable under hyper-physiological loading levels using a side screw approach in compression and torsion and the combination of side and top screws in tension and bending. We thus conclude that the developed holding system is mechanically reliable and biologically compatible for application in a new generation of multiaxial bioreactors., ACS Biomaterials Science & Engineering, 8 (9), ISSN:2373-9878

Hydrogen gas (H

Groundwater and surface water bodies may have contaminants from urban, industrial, or agricultural wastewater, including emerging contaminants (ECs) or micropollutants (MPs). Frequently, they are not efficiently removed by microbial action due to their minimal concentration in water and the low microbiota affinity for complex compounds. This work developed a process allowing the adsorption of contaminants and their simultaneous biodegradation using horizontal tubular fixed-bed biofilm reactors (HTR). Each HTR has two zones: an equalizer-aerator of the incoming liquid flow and a fixed bed zone. This zone was packed with a mixed support material consisting of granular bio-activated carbon (Bio-GAC) and porous material that increases the bed permeability, thus decreasing the pressure drop. Five microbial communities were acclimated and immobilized in granular activated carbon (GAC) to obtain different specialized Bio-GAC particles able to remove the micropollutants ibuprofen, naproxen, atrazine, carbaryl, and diazinon. The Bio-GAC particles were transferred to HTRs continuously run in microaerophilia at several MPs loading rates. Under these conditions, the removal efficiencies of MPs, except atrazine and carbaryl, were around 100.

P-nitrophenol (PNP) is highly toxic and difficult to degrade, causing great harm to the ecological environment and human health. A two-stage bench-scale membrane biofilm reactor (MBfR) was constructed to treat wastewater containing high concentration of PNP and the generated nitrogen without external organic carbon sources. The two reactors were supplied with oxygen and methane, respectively. O2-MBfR was used for the degradation of PNP and the improvement of wastewater biodegradability. CH4-MBfR was used for the total nitrogen (TN) removal treatment from O2-MBfR effluent. In this experiment, the performance of the two-stage MBfR process was evaluated and optimized by adjusting operational parameters (aeration pressure, HRT, and pH). Under the optimal operation parameters, the removal efficiencies of PNP (100 mg/L) and TN attained 89.70% and 69.24%, respectively, and the removal loads were 0.930 g·m−2·d−1 and 241.42 mg·m−2·d−1, respectively. The reactor was able to accommodate the concentrations of PNP up to 200–400 mg/L, and the reactor reached maximum efficiency throughout the process when the concentration of PNP in the wastewater was 250 mg/L. The removal rates of PNP and TN reached 95.0% and 69.48%, respectively, and the removal loads were 2.37 g·m−2·d−1 and 96.22 mg·m−2·d−1, respectively. This research provides a better solution for multi-MBfR to treat toxic industrial wastewater containing phenol, nitrophenol, and further TN removal, which would not release any air pollutants into the atmosphere.

Conventional bioprocess models for wastewater treatment are based on aggregated bulk biomass concentrations and do not incorporate microbial physiological diversity. Such a broad aggregation of microbial functional groups can fail to predict ecosystem dynamics when high levels of physiological diversity exist within trophic guilds. For instance, functional diversity among nitrite-oxidizing bacteria (NOB) can obfuscate engineering strategies for their out-selection in activated sludge (AS), which is desirable to promote energy-efficient nitrogen removal. Here, we hypothesized that different NOB populations within AS can have different physiological traits that drive process performance, which we tested by estimating biokinetic growth parameters using a combination of highly replicated respirometry, genome-resolved metagenomics, and process modeling. A lab-scale AS reactor subjected to a selective pressure for over 90 days experienced resilience of NOB activity. We recovered three coexisting

Efficient and large-scale expansion of mesenchymal stem/stromal cells (MSCs) has always been a formidable challenge to researchers in cell-based therapies and regenerative medicine. To reconcile major drawbacks of 2D planar culturing system, we innovatively developed an automated closed industrial scale cell production (ACISCP) platform based on GMP-grade microcarrier for culture of umbilical cord-mesenchymal stem/stromal cells (UCMSCs), in accordance with the criteria of stem cell bank. ACISCP system is a fully closed system, which employs different models of vivaSPIN bioreactors (CytoNiche Biotech, China) for scale-up cell culture and vivaPREP (CytoNiche Biotech, China) for automated cell harvesting and cell dosage preparation. To realize industrial scale expansion of UCMSCs, a three-stage expansion was conducted with 1 L, 5 and 15 L vivaSPIN bioreactors. Using 3D TableTrixsup®/supand ACISCP system, we inoculated 1.5 × 10sup7/supof UCMSCs into 1 L vivaSPIN bioreactor and finally scaled to two 15 L bioreactor. A final yield of 2.09 × 10sup10/supcells with an overall expansion factor of 1975 within 13 days. The cells were harvested, concentrated, washed and prepared automatically with vivaPREP. The entire process was realized with ACISCP platform and was totally enclosed. Critical quality attributes (CQA) assessments and release tests of MSCs, including sterility, safety, purity, viability, identity, stability and potency were performed accordingly. The quality of cells harvested from 3D culture on the ACISCP and conventional 2D planar culture counterpart has no significant difference. This study provides a bioprocess engineering platform, harnessing GMP-grade 3D TableTrixsup®/supmicrocarriers and ACISCP to achieve industrial-scale manufacturing of clinical-grade hMSCs.

The legislation for environment protection requires strict controls of the wastewater releasing in water bodies. The wastewater treatment plants (WWTP) have been used for organic matter degradation; however, the residual total phosphorus (TP) removal has not been efficient. TP and nitrogen present in wastewater are associated to eutrophication of water bodies and algae growth. Therefore, this study discusses the efficiency of phosphorus removal by a slow filter (SF), complementary to a WWTP and the microbial community involved. The results showed that the use of SF, with or without macrophytes, is not suitable to remove TP. Spatial variation in microbial communities distributed in three distinct zones was identified in the SF. Proteobacteria, Bacteroidetes, Chloroflexi and Firmicutes covered the hydrolytic and fermentative bacteria. The acetogenesis, nitrification, and denitrification, as well as the removal of phosphorus from the effluent, were performed by representatives affiliated to different groups. Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria among these, Dokdonella sp., Frateuria sp., Comamonas sp., Diaphorobacter sp., Nitrosospira sp., Ferruginibacter sp., Flavobacterium sp., and the uncultured OD1 were the most abundant bacteria in the SF. The low efficiency for TP removing from SF effluents can be explained by the low abundance of phosphorus accumulating organisms (PAOs), with the association of the low concentration of biodegradable organic matter in the inlet effluent. Therefore, the alternative to using SF as a complement to WWTPs, as recommended by some Brazilian environmental agencies, did not prove to be viable and new approaches must be evaluated. KEY POINTS: • The phosphorus removal was performed by a slow filter system in a WWTP but obtained a low efficiency. • Microbial spatial variation was distributed into distinct zones from slow filter. • Low abundance of PAOs was observed due to the low availability of organic matter.

There is increasing interest in thiosulfate-driven denitrification for low C/N wastewater treatment, but the denitrification performance varies with the thiosulfate oxidation pathways. Models have been developed to predict the products of denitrification, but few consider thiosulfate reduction to elemental sulfur (S

Producción Científica, Biogas-based biopolymer production represents an alternative biogas valorization route with potential to cut down plastic pollution and greenhouse gas emissions. This study investigated for the first time the continuous bioconversion of methane, contained in biogas, into poly(3-hydroxybutyrate) (PHB) by a mixed methanotrophic culture using an innovative high mass-transfer Taylor flow bioreactor. Following a hydrodynamic flow regime mapping, the influence of the gas residence time and the internal gas recirculation on CH4 abatement was assessed under non nutrient limiting conditions. Under optimal operational conditions (gas residence time of 60 min and internal gas recycling ratio of 17), the bioreactor was able to support a CH4 removal efficiency of 63.3%, a robust CH4 elimination capacity (17.2 g-CH4 m-3h-1) and a stable biomass concentration (1.0 g L-1). The simultaneous CH4 abatement and PHB synthesis was investigated under 24-h:24-h nitrogen feast/famine continuous operation. The cyclic nitrogen starvation and the Taylor flow imposed in the bioreactor resulted in a relatively constant biomass concentration of 0.6 g L-1 with PHB contents ranging from 11 to 32% w w-1 (on a dry weight basis), entailing an average PHB productivity of 5.9 g-PHB m-3 d-1 with an associated PHB yield of 19.8 mg-PHB g-CH4-1. Finally, the molecular analysis of the microbial population structure indicated that type II methanotrophs outcompeted non-PHB accumulating type I methanotrophs, with a heterotrophic-methanotrophic consortium enriched in Methylocystis, Hyphomicrobium, Rubinisphaeraceae SH PL14 and Pseudonocardia., Ministerio de Ciencia, Innovación y Universidades (project RTI2018-0-096441-B-I00), Erasmus+ International Master of Science in Environmental Technology and Engineering (project 2017-1957/001-001-EMJMD), Junta de Castilla y Leon - Fondo Europeo de Desarrollo Regional (grants CLU 2017-09 and UIC 315)

The present study assessed the performance and fouling of adding granular activated carbon (GAC) and sponge (BioCube), as two different media, to a membrane sequencing batch reactor (MSBR) system in wastewater treatment containing Acid Red 18 (AR 18). Anaerobic phase, aerobic phase, and hydraulic retention times (HRTs) of 24 h, 12 h, and 72 h were considered for 500 mg/L AR 18 removal at a sludge retention time (SRT) of 20 days by separately adding up to 35% BioCube volume and 8 g/L GAC to the reactors. Based on the kinetic study, 63 mg/L (87% removal) and 115 mg/L (77% removal) remaining dye were reported in the GAC and BioCube membrane sequencing batch reactors (GAC-MSBR and BioCube-MSBR), respectively. A gradual oxidation-reduction potential decline toward -416 mV confirmed better dye removal in GAC-MSBR than BioCube-MSBR, observing a sudden drop to -354 mV. The morphology can explain better biological treatment in GAC-MSBR in addition to the adsorption process. Soluble microbial products (SMPs) of 126.92 mg/L and 395.18 mg/L were obtained for GAC-MSBR and BioCube-MSBR, respectively. Chemical oxygen demand (COD) and SMP indicated that the GAC-MSBR water quality is better than that of the other reactor.

Landfill leachate, due to its recalcitrant nature and toxicity, poses a serious environmental threat, which requires the implementation of effective treatment processes. In this work, a full-scale treatment system consisting of two Sequencing Batch Reactors (SBRs) was used for the processing of landfill leachate of intermediate to mature age (BOD/COD ratio of 0.16). Biosystem operation resulted in BOD

Conventional activated sludge (AS) systems are widely used to treat domestic sewage worldwide. However, the removal of nitrogen in the AS system is limited, and its concentration in the effluent exceeds the recommended values in the discharge standards. In this study, a pilot experiment was conducted to improve nitrogen removal during municipal sewage treatment by operating AS and anaerobic-aerobic (AO) systems under low dissolved oxygen (DO) conditions of less than 0.5 mg L