Methanogenic population dynamics during semi‐continuous biogas fermentation and acidification by overloading

Aims: This study was to investigate the methanogenic community in a biogas reactor from start‐up to acidification conditions. Furthermore, reliability and accuracy of the applied quantitative real‐time PCR method (Q‐PCR) was briefly evaluated. Methods and Results: A mesophilic (37°C), maize silage fed, continuously stirred tank reactor was surveyed. It was operated semi‐continuously with increasing daily organic loading rates (OLRs) to reach acidification. Gas production and organic acid composition were measured. Methanogenic community structure was determined by 16S rDNA‐based Q‐PCR to estimate the abundance of key methanogenic micro‐organisms. 16S rDNA of hydrogenotrophic Methanobacteriales was most abundant at OLRs of ≥3·7 g dry organic matter (DOM) l^?1 day^?1. By contrast, that of aceticlastic Methanosaetaceae predominated at lower OLRs but disappeared at OLRs of ≥4·1 g DOM l^?1 day^?1. At the same OLR, the propionate concentration increased dramatically indicating the acidification of the digester. Application of internal standards to examine Q‐PCR’s accuracy revealed that the detected amount of 16S rDNA may vary within one log cycle. Conclusions: These results suggest that the absence of Methanosaetaceae might be taken as biological indicator for process’ instability. Inhibitory effects on Q‐PCR analyses could not be determined based on the spiking experiments. Significance and Impact of the Study: In this study, reactors’ microbiology was observed over time using Q‐PCR. Insights into the abundance of different methanogens might be used to improve the performance of biogas reactors.     

Improving biogas production from protein-rich distillery wastewater by decreasing ammonia inhibition

A large quantity of protein-rich distillery wastewater is produced during the process of bio-ethanol production from kitchen waste. It is difficult, however, to treat protein-rich distillery wastewater by anaerobic digestion due to ammonia inhibition. In this study, a novel method was investigated to reduce ammonia inhibition during thermophilic anaerobic digestion through the recirculation of water-washed biogas into the headspace (R1 system) or liquid phase (R2 system) of the reactors. The results show that the method greatly improved biogas production from distillery wastewater. R2 system achieved stable biogas production at a higher organic loading rate (OLR) of 4.0 g VTS/L/d than R1 system at 3.0 g VTS/L/d. At the same OLR, we observed a higher biogas production rate but lower accumulation of NH₄⁺ and volatile fatty acids in the reactor, and higher ammonia absorption rate in the water tank of R2 system than R1 system. The better performance of R2 system could be attributed to the more efficient removal of ammonia from liquid phase. In addition, adjusting the C/N ratio of distillery wastewater from 9.0 to 11.4 significantly enhanced the maximum OLR from 3.0 to 7.0 g VTS/L/d in R1 system.     

In situ ammonia production as a sanitation agent during anaerobic digestion at mesophilic temperature

Aim: To measure the sanitizing effect of mesophilic (37°C) anaerobic digestion in high ammonia concentrations produced in situ. Methods and Results: Indicator organisms and salmonella were transferred to small‐scale anaerobic batch cultures and D‐values were calculated. Batch cultures were started with material from two biogas processes operating at high (46 mmol l^?1) and low (1·6 mmol l^?1) ammonia concentration. D‐values were shortened from c. 3 days to <1 day for the bacteria. MS2 had the same D‐value (1·3 days) independent of ammonia concentration whereas ΦX174 and 28B were faster inactivated in the control (1·1 and 7·9 days) than in the high ammonia (8·9 and 39 days) batch cultures. Conclusion: Running biogas processes at high levels of ammonia shortens the time to meet EU regulation concerning reduction of salmonella and enterococci (5 log). Unless a minimum retention time of 2 days, post‐treatment digestion is needed to achieve sufficient sanitation in continuous biogas processes. Significance and Impact of the Study: Running mesophilic biogas processes at high ammonia level produces residue with a high fertilizer value. With some stipulations concerning management parameters, such processes provide a method of bacterial sanitation without preceding pasteurization of the incoming organic waste.     

Molecular microbial ecology of stable versus failing rice straw anaerobic digesters

Waste rice straw (RS) is generated in massive quantities around the world and is often burned, creating greenhouse gas and air quality problems. Anaerobic digestion (AD) may be a better option for RS management, but RS is presumed to be comparatively refractory under anaerobic conditions without pre-treatment or co-substrates. However, this presumption assumes frequent reactor feeding regimes but less frequent feeding may be better for RS due to slow hydrolysis rates. Here, we assess how feeding frequency (FF) and organic loading rate (OLR) impacts microbial communities and biogas production in RS AD reactors. Using 16S rDNA amplicon sequencing and bioinformatics, microbial communities from five bench-scale bioreactors were characterized. At low OLR (1.0 g VS l⁻¹ day⁻¹), infrequently fed units (once every 21 days) had higher specific biogas yields than more frequent feeding (five in 7 days), although microbial community diversities were statistically similar (P > 0.05; ANOVA with Tukey comparison). In contrast, an increase in OLR to 2.0 g VS l⁻¹ day⁻¹ significantly changed Archaeal and fermenting Eubacterial sub-communities and the least frequency fed reactors failed. ‘Stable’ reactors were dominated by Methanobacterium, Methanosarcina and diverse Bacteroidetes, whereas ‘failed’ reactors saw shifts towards Clostridia and Christensenellaceae among fermenters and reduced methanogen abundances. Overall, OLR impacted RS AD microbial communities more than FF. However, combining infrequent feeding and lower OLRs may be better for RS AD because of higher specific yields.     

Bioaugmentation of Syntrophic Acetate-Oxidizing Culture in Biogas Reactors Exposed to Increasing Levels of Ammonia

The importance of syntrophic acetate oxidation for process stability in methanogenic systems operating at high ammonia concentrations has previously been emphasized. In this study we investigated bioaugmentation of syntrophic acetate-oxidizing (SAO) cultures as a possible method for decreasing the adaptation period of biogas reactors operating at gradually increased ammonia concentrations (1.5 to 11 g NH₄⁺-N/liter). Whole stillage and cattle manure were codigested semicontinuously for about 460 days in four mesophilic anaerobic laboratory-scale reactors, and a fixed volume of SAO culture was added daily to two of the reactors. Reactor performance was evaluated in terms of biogas productivity, methane content, pH, alkalinity, and volatile fatty acid (VFA) content. The decomposition pathway of acetate was analyzed by isotopic tracer experiments, and population dynamics were monitored by quantitative PCR analyses. A shift in dominance from aceticlastic methanogenesis to SAO occurred simultaneously in all reactors, indicating no influence by bioaugmentation on the prevailing pathway. Higher abundances of Clostridium ultunense and Tepidanaerobacter acetatoxydans were associated with bioaugmentation, but no influence on Syntrophaceticus schinkii or the methanogenic population was distinguished. Overloading or accumulation of VFA did not cause notable dynamic effects on the population. Instead, the ammonia concentration had a substantial impact on the abundance level of the microorganisms surveyed. The addition of SAO culture did not affect process performance or stability against ammonia inhibition, and all four reactors deteriorated at high ammonia concentrations. Consequently, these findings further demonstrate the strong influence of ammonia on the methane-producing consortia and on the representative methanization pathway in mesophilic biogas reactors.     

Effects of ferrous iron on the performance and microbial community in aerobic granular sludge in relation to nutrient removal

Lab‐scale experiments were conducted to investigate the effects of ferrous iron on nutrient removal performance and variations in the microbial community inside aerobic granular sludge for 408 days. Two reactors were simultaneously operated, one without added ferrous iron (SBR1), and one with 10 mg Fe²⁺ L^?1 of added ferrous iron (SBR2). A total of 1 mg Fe²⁺ L^?1 of added ferrous iron was applied to SBR1 starting from the 191st day to observe the resulting variations in the nutrient removal performance and the microbial community. The results show that ammonia‐oxidizing bacteria (AOB) could not oxidize ammonia due to a lack of iron compounds, but they could survive in the aerobic granular sludge. Limited ferrous iron addition encouraged nitrification. Enhanced biological phosphorus removal (EBPR) from both reactors could not be maintained regardless of the amount of ferrous iron that was applied. EBPR was established in both reactors when the concentration of mixed liquor suspended solid (MLSS) and the percentage of Accumulibacteria increased. A total of 10 mg Fe²⁺ L^?1 of added ferrous iron had a relatively adverse effect on the growth of AOB species compared to 1 mg Fe²⁺ L^?1 of added ferrous iron, but it encouraged the growth of Nitrospira sp. and Accumulibacteria, which requires further study. It could be said that the compact and stable structure of aerobic granular sludge preserved AOB and NOB from Fe‐deficient conditions, and wash‐out during the disintegration period. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:716–725, 2017     

Corynebacterium glutamicum,a natural overproducer of succinic acid?

Corynebacterium glutamicum is well known as an important industrial amino acid producer. For a few years, its ability to produce organic acids, under micro‐aerobic or anaerobic conditions was demonstrated. This study is focused on the identification of the culture parameters influencing the organic acids production and, in particular, the succinate production, by this bacterium. Corynebacterium glutamicum 2262, used throughout this study, was a wild‐type strain, which was not genetically designed for the production of succinate. The oxygenation level and the residual glucose concentration appeared as two critical parameters for the organic acids production. The maximal succinate concentration (4.9 g L^?1) corresponded to the lower k_La value of 5 h^?1. Above 5 h^?1, a transient accumulation of the succinate was observed. Interestingly, the stop in the succinate production was concomitant with a lower threshold glucose concentration of 9 g L^?1. Taking into account this threshold, a fed‐batch culture was performed to optimize the succinate production with C. glutamicum 2262. The results showed that this wild‐type strain was able to produce 93.6 g L^?1 of succinate, which is one of the highest concentration reported in the literature.     

Toxicity of ammonia and nitrite to yellow catfish (Pelteobagrus fulvidraco)

The 96‐h LC₅₀ (median lethal concentration, LC₅₀) tests were conducted using four different sizes of yellow catfish Pelteobagrus fulvidraco to provide primary information on the sensitivity of this species to elevated ammonia and/or nitrite, and to determine if the sensitivity is mediated by size under the same conditions. The results showed that 96‐h LC₅₀ of fish weighing 0.034 ± 0.002, 0.296 ± 0.049, 3.52 ± 0.95 and 32.96 ± 5.75 g to total ammonia nitrogen‐N was 24.96, 35.85, 47.44 and 68.79 mg L^?1, respectively; un‐ionized ammonia nitrogen‐N was 0.34, 0.49, 0.65 and 0.94 mg L^?1 in test conditions of pH 7.42 and 23°C; and that nitrite nitrogen‐N was 69.06, 97.23, 133.61 and 196.05 mg L^?1 in test conditions of pH 7.58 and 23°C, respectively. The NOEL (No Observable Effect Level) of fish (body weight from 0.03 to 30 g) to ammonia and nitrite was 2.25–6.22 mg L^?1 total ammonia nitrogen‐N, 0.03–0.10 mg L^?1 un‐ionized ammonia nitrogen‐N in test conditions of pH 7.42 and 23°C, and 6.27–17.68 mg L^?1 nitrite nitrogen‐N in test conditions of pH 7.58 and 23°C, respectively. These results indicate that the susceptibility of this fish to total ammonia or nitrite was reduced with increasing size, and that a dose‐dependent relationship might exist between them. The 96‐h LC₅₀ and NOEL of different sizes of fish to total ammonia, un‐ionized ammonia and nitrite would be important to know for water quality standards in yellow catfish aquaculture.     

Functionally redundant but dissimilar microbial communities within biogas reactors treating maize silage in co-fermentation with sugar beet silage

Numerous observations indicate a high flexibility of microbial communities in different biogas reactors during anaerobic digestion. Here, we describe the functional redundancy and structural changes of involved microbial communities in four lab-scale continuously stirred tank reactors (CSTRs, 39°C, 12 L volume) supplied with different mixtures of maize silage (MS) and sugar beet silage (SBS) over 80 days. Continuously stirred tank reactors were fed with mixtures of MS and SBS in volatile solid ratios of 1:0 (Continuous Fermenter (CF) 1), 6:1 (CF2), 3:1 (CF3), 1:3 (CF4) with equal organic loading rates (OLR 1.25 kgVS m⁻³ d⁻¹) and showed similar biogas production rates in all reactors. The compositions of bacterial and archaeal communities were analysed by 454 amplicon sequencing approach based on 16S rRNA genes. Both bacterial and archaeal communities shifted with increasing amounts of SBS. Especially pronounced were changes in the archaeal composition towards Methanosarcina with increasing proportion of SBS, while Methanosaeta declined simultaneously. Compositional shifts within the microbial communities did not influence the respective biogas production rates indicating that these communities adapted to environmental conditions induced by different feedstock mixtures. The diverse microbial communities optimized their metabolism in a way that ensured efficient biogas production.     

Enrichment and description of novel bacteria performing syntrophic propionate oxidation at high ammonia level

Inefficient syntrophic propionate degradation causes severe operating disturbances and reduces biogas productivity in many high-ammonia anaerobic digesters, but propionate-degrading microorganisms in these systems remain unknown. Here, we identified candidate ammonia-tolerant syntrophic propionate-oxidising bacteria using propionate enrichment at high ammonia levels (0.7–0.8 g NH₃ L⁻¹) in continuously-fed reactors. We reconstructed 30 high-quality metagenome-assembled genomes (MAGs) from the propionate-fed reactors, which revealed two novel species from the families Peptococcaceae and Desulfobulbaceae as syntrophic propionate-oxidising candidates. Both MAGs possess genomic potential for the propionate oxidation and electron transfer required for syntrophic energy conservation and, similar to ammonia-tolerant acetate degrading syntrophs, both MAGs contain genes predicted to link to ammonia and pH tolerance. Based on relative abundance, a Peptococcaceae sp. appeared to be the main propionate degrader and has been given the provisional name “Candidatus Syntrophopropionicum ammoniitolerans”. This bacterium was also found in high-ammonia biogas digesters, using quantitative PCR. Acetate was degraded by syntrophic acetate-oxidising bacteria and the hydrogenotrophic methanogenic community consisted of Methanoculleus bourgensis and a yet to be characterised Methanoculleus sp. This work provides knowledge of cooperating syntrophic species in high-ammonia systems and reveals that ammonia-tolerant syntrophic propionate-degrading populations share common features, but diverge genomically and taxonomically from known species.     

Effects of ammonia on growth,digestion and antioxidant capacity in juvenile yellow catfish Pelteobagrus fulvidraco (Richardson, 1846)

Effects of ammonia stress on food ingestion, growth, digestion and antioxidant capacity were investigated in juvenile yellow catfish Pelteobagrus fulvidraco (Richardson) with initial body weights of 20.24 ± 0.18 g. The fish were reared in triplicate in 15 experimental tanks at a rate of 30 fish per tank for 56 days. Water was maintained at a dissolved oxygen (DO) level of over 6.2 mg L^?1, pH 7.2–7.6, and temperature of 29.0 ± 1.5°C under a natural 12L: 12D photoperiod. Survival, food ingestion (FI), specific growth rate (SGR), food conversion efficiency (FCE), apparent digestibility coefficient (ADC), total antioxidant capacity (T‐AOC), levels of glutathione (GSH) and malonaldehyde (MDA), and activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH‐PX) of the juveniles were determined in total ammonia nitrogen (NH₃‐N+NH₄‐N) levels of 0 (control group), 3.36, 6.72, 13.44 and 26.88 mg L^?1. The results show that the survival, FI, SGR, FCE, and ADC decreased significantly with an increase in total ammonia nitrogen (p < .05), and a significantly negative correlation between SGR and total ammonia nitrogen levels (p < .05). T‐AOC, SOD, CAT, GSH‐PX and GSH in the blood, liver and gills were found to decline significantly with an increase in the total ammonia nitrogen level (p < .05), while the MDA in the blood, liver and gills was elevated significantly with the increase in total ammonia nitrogen (p < .05). The results indicate a threshold in the induction of the T‐AOC and activities of antioxidant enzymes in yellow catfish tissues with a total ammonia nitrogen increase. In the present study the total ammonia nitrogen threshold thus changed from 6.72 mg L^?1 in the juvenile yellow catfish.     

Start-up and operation strategies on the liquefied food waste anaerobic digestion and a full-scale case application

Batch anaerobic digestion was employed to investigate the efficient start-up strategies for the liquefied food waste, and sequencing batch digestion was also performed to determine maximum influent organic loading rate (OLR) for efficient and stable operation. The results indicated that the start-up could be well improved using appropriate wastewater organic load and food-to-microorganism ratios (F/M). When digestion was initialized at low chemical oxygen demand (COD) concentration of 20.0 gCOD L^?1, the start-up would go well using lower F/M ratio of 0.5–0.7. The OLR 7.0 gCOD L^?1 day^?1 was recommended for operating the ASBR digestion, in which the COD conversion of 96.7 ± 0.53 % and biomethane yield of 3.5 ± 0.2 L gCOD^?1 were achieved, respectively. The instability would occur when OLR was higher than 7.0 gCOD L^?1 day^?1, and this instability was not recoverable. Lipid was suggested to be removed before anaerobic digestion. The anaerobic digestion process in engineering project ran well, and good performance was achieved when the start-up and operational strategies from laboratory study were applied. For case application, stable digestion performance was achieved in a digester (850 m³ volume) with biogas production of 1.0–3.8 m³ m^?3 day^?1.     

Effect of chitosan on UASB treating POME during a transition from mesophilic to thermophilic conditions

The effects of chitosan addition on treatment of palm oil mill effluent were investigated using two lab-scale upflow anaerobic sludge bed (UASB) reactors: (1) with chitosan addition at the dosage of 2 mg chitosan per g volatile suspended solids on the first day of the operation (R1), (2) without chitosan addition (the control, R2). The reactors were inoculated with mesophilic anaerobic sludge which was acclimatized to a thermophilic condition with a stepwise temperature increase of 5 °C from 37 to 57 °C. The OLR ranged from 2.23 to 9.47 kg COD m⁻³ day⁻¹. The difference in biogas production rate increased from non-significant to 18% different. The effluent volatile suspended solids of R1 was 65 mg l⁻¹ lower than that of R2 on Day 123. 16S rRNA targeted denaturing gradient gel electrophoresis (DGGE) fingerprints of microbial community indicated that some methanogens in the genus Methanosaeta can be detected in R1 but not in R2.     

Metabolism in 1,3‐propanediol fed‐batch fermentation by a D‐lactate deficient mutant of Klebsiella pneumoniae

Klebsiella pneumoniae HR526, a new isolated 1,3‐propanediol (1,3‐PD) producer, exhibited great productivity. However, the accumulation of lactate in the late‐exponential phase remained an obstacle of 1,3‐PD industrial scale production. Hereby, mutants lacking D ‐lactate pathway were constructed by knocking out the ldhA gene encoding fermentative D ‐lactate dehydrogenase (LDH) of HR526. The mutant K. pneumoniae LDH526 with the lowest LDH activity was studied in aerobic fed‐batch fermentation. In experiments using pure glycerol as feedstock, the 1,3‐PD concentrations, conversion, and productivity increased from 95.39 g L^?1, 0.48 and 1.98 g L^?1 h^?1 to 102. 06 g L^?1, 0.52 mol mol^?1 and 2.13 g L^?1 h^?1, respectively. The diol (1,3‐PD and 2,3‐butanediol) conversion increased from 0.55 mol mol^?1 to a maximum of 0.65 mol mol^?1. Lactate would not accumulate until 1,3‐PD exceeded 84 g L^?1, and the final lactate concentration decreased dramatically from more than 40 g L^?1 to <3 g L^?1. Enzymic measurements showed LDH activity decreased by 89–98% during fed‐batch fermentation, and other related enzyme activities were not affected. NADH/NAD⁺ enhanced more than 50% in the late‐exponential phase as the D ‐lactate pathway was cut off, which might be the main reason for the change of final metabolites concentrations. The ability to utilize crude glycerol from biodiesel process and great genetic stability demonstrated that K. pnemoniae LDH526 was valuable for 1,3‐PD industrial production. Biotechnol. Bioeng. 2009; 104: 965–972. © 2009 Wiley Periodicals, Inc.     

Laboratory scale examination of the effects of overloading on the anaerobic digestion by glycerol

The anaerobic digestion of pure glycerol, which produces a baseline acetic acid to propionic acid ratio of 0.2, was studied in laboratory scale reactors (3 l working volume) at mesophilic temperature (37 °C) with 3000 mg chemical oxygen demand (COD) l⁻¹d⁻¹. During the experiment tVFA and C2-C6 VFA analysis and daily biogas yield measurement were carried out. Following 10 days of a 15% d⁻¹ increase in the organic loading rate (OLR) of 3.0-10.5 g COD l⁻¹d⁻¹, the concentration of propionic acid increased to 6200-8000 mg l⁻¹. Then the inoculum was divided into three parts feeding with 100% glycerol, 50% glycerol + 50% acetic acid, and 50% glycerol + 50% thick stillage, (presented in % of 2.60 g COD l⁻¹d⁻¹ OLR), respectively. The application of co-substrates reduced the recovery period by 5 days compared to feeding with pure glycerol. When the reactors were loaded with glycerol again (10% OLR raise per day) the previously applied co-substrates had a positive effect on the VFA composition and the biogas yield as well.     

Polysialic acid production using Escherichia coli K1 in a disposable bag reactor

Polysialic acid (polySia), consisting of α‐(2,8)‐linked N‐acetylneuraminic acid monomers plays a crucial role in many biological processes. This study presents a novel process for the production of endogenous polySia using Escherichia coli K1 in a disposable bag reactor with wave‐induced mixing. Disposable bag reactors provide easy and fast production in terms of regulatory requirements as GMP, flexibility, and can easily be adjusted to larger production capacities not only by scale up but also by parallelization. Due to the poor oxygen transfer rate compared to a stirred tank reactor, pure oxygen was added during the cultivation to avoid oxygen limitation. During the exponential growth phase the growth rate was 0.61 h^?1. Investigation of stress‐related product release from the cell surface showed no significant differences between the disposable bag reactor with wave‐induced mixing and the stirred tank reactor. After batch cultivation a cell dry weight of 6.8 g L^?1 and a polySia concentration of 245 mg L^?1 were reached. The total protein concentration in the supernatant was 132 mg L^?1. After efficient and time‐saving downstream processing characterization of the final product showed a protein content of below 0.04 mg_protein/g_polySia and a maximal chain length of ～90 degree of polymerization.     

Ammonia may play an important role in the succession of cyanobacterial blooms and the distribution of common algal species in shallow freshwater lakes

With the human intensification of agricultural and industrial activities, large amount of reduced nitrogen enter into the biosphere, which consequently results in the development of global eutrophication and cyanobacterial blooms. However, no research had reported the effect of ammonia toxicity on the algal succession. In this study, we investigated the ammonia toxicity to 19 algal species or strains to test the hypothesis that ammonia may regulate the succession of cyanobacterial blooms and the distribution of common algal species in freshwater lakes. The bloom‐forming cyanobacterium Microcystis aeruginosa PCC 7806 suffered from ammonia toxicity at high pH value and light intensity conditions. Low NH₄Cl concentration (0.06 mmol L^?1) resulted in the decrease of operational PSII quantum yield by 50% compared with the control exposed to 1000 μmol photons m^?2 s^?1 for 1 h at pH 9.0 ± 0.2, which can be reached in freshwater lakes. Furthermore, the tolerant abilities to NH₃ toxicity of 18 freshwater algal species or strains were as follows: hypertrophication species > eutrophication species > mesotrophication species > oligotrophication species. The different sensitivities of NH₃ toxicity in this study could well explain the distributing rule of common algal species in the freshwater lakes of different trophic states. Meanwhile, the cyanobacterial bloom (e.g. M. aeruginosa) always happened at the low concentration of ammonia in summer, and disappeared with the decrease of ammonia. This may be attributed to the toxic effect of ammonia to M. aeruginosa in spring (the average and maximum ammonia concentration were 0.08 and 0.72 mmol L^?1 in 33 Chinese lakes), and the low level of NH₃‐N in summer and fall in the lakes might be used as preferred nitrogen nutrition by M. aeruginosa, rather than with toxicity. Therefore, ammonia could be a key factor to determine the distribution of common algal species and cyanobacterial bloom in the freshwater systems.     

Heterotrophic production of Chlorella sp. TISTR 8990—biomass growth and composition under various production conditions

The green microalga Chlorella sp. TISTR 8990 was grown heterotrophically in the dark using various concentrations of a basal glucose medium with a carbon‐to‐nitrogen mass ratio of 29:1. The final biomass concentration and the rate of growth were highest in the fivefold concentrated basal glucose medium (25 g L^?1 glucose, 2.5 g L^?1 KNO₃) in batch operations. Improving oxygen transfer in the culture by increasing the agitation rate and decreasing the culture volume in 500‐mL shake flasks improved growth and glucose utilization. A maximum biomass concentration of nearly 12 g L^?1 was obtained within 4 days at 300 rpm, 30°C, with a glucose utilization of nearly 76% in batch culture. The total fatty acid (TFA) content of the biomass and the TFA productivity were 102 mg g^?1 and 305 mg L^?1 day^?1, respectively. A repeated fed‐batch culture with four cycles of feeding with the fivefold concentrated medium in a 3‐L bioreactor was evaluated for biomass production. The total culture period was 11 days. A maximum biomass concentration of nearly 26 g L^?1 was obtained with a TFA productivity of 223 mg L^?1 day^?1. The final biomass contained (w/w) 13.5% lipids, 20.8% protein and 17.2% starch. Of the fatty acids produced, 52% (w/w) were saturated, 41% were monounsaturated and 7% were polyunsaturated (PUFA). A low content of PUFA in TFA feedstock is required for producing high quality biodiesel. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1589–1600, 2017     

Constructed wetlands with free water surface for treatment of aquaculture effluents

The aim of the study was to determine the reduction of the overall environmental load (in terms of organic and nutrient load) in effluents of a flow‐through trout farm. Effluents of a flow‐through system for rainbow trout (Oncorhynchus mykiss) production passed through constructed wetlands with free water surface. Removal of nutrients was determined in three wetlands of 350 m² each at hydraulic residence times (HRTs) of 3.5, 5.5 and 11 h. The areal load of total suspended solids (TSS), chemical oxygen demand (COD), total phosphorus (TP), and total nitrogen (TN) varied in terms of HRTs from 12.3–36.8 g m^?2 day^?1, 21.7–65.2 g m^?2 day^?1, 0.23–0.70 g m^?2 day^?1, and 1.46–4.37 g m^?2 day^?1. Values for reduction of suspended solids, COD, TP, and TN were 67–72%, 30–31%, 41–53% ,and 19–30%, respectively. Significantly lower nutrient concentrations in the effluent among the wetlands were only found for nitrogen parameters: TN and ammonia concentrations were lower in the wetlands with a HRT of 5.5 h (0.89 mg L^?1, 0.11 mg L^?1) and 11 h (0.81 mg L^?1, 0.11 mg L^?1) compared with the one with 3.5 h (0.96 mg L^?1, 0.16 mg L^?1).