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Multiple‐batch production is the most widely practiced method of raising channel catfish. Producers are increasingly adopting intensified production practices in multiple‐batch systems by increasing stocking density and aeration rates as a means to improve cost efficiencies. Proven stocking recommendations are required for the efficient implementation of recent developments in multiple‐batch production. Twelve 0.4‐ha ponds were understocked with 17,484, 20,612, and 26,124 fingerlings/ha (mean weight = 40 g/fish) over equal weights of carryover fish (0.46 kg/fish @ 4,589 kg/ha). Fish were fed once daily to apparent satiation with a 28% protein floating feed and aerated with a single 7.4‐kW electric paddlewheel aerator. Density‐dependent significant differences were absent for gross, net, daily net yields, marketable yields (≥0.54 kg), growth (g/day), and survival. Sub‐marketable yield (<0.54 kg) and feeding rate increased significantly with increased understocking density. Economic analysis revealed increased breakeven prices and diminished net returns with increased stocking density when sub‐marketable fish were not considered as revenue. These differences in production costs and profits among the three treatments became minimal when sub‐marketable fish were included as revenue. All three density treatments attained positive annual net cash flows. This study validates channel catfish understocking densities of 17,000–26,000 fish/ha to improve cost efficiency in intensively aerated, multiple‐batch production systems.

With an increasing demand for efficacious, safe, and affordable vaccines for human and animal use, process intensification in cell culture-based viral vaccine production demands advanced process strategies to overcome the limitations of conventional batch cultivations. However, the use of fed-batch, perfusion, or continuous modes to drive processes at high cell density (HCD) and overextended operating times has so far been little explored in large-scale viral vaccine manufacturing. Also, possible reductions in cell-specific virus yields for HCD cultivations have been reported frequently. Taking into account that vaccine production is one of the most heavily regulated industries in the pharmaceutical sector with tough margins to meet, it is understandable that process intensification is being considered by both academia and industry as a next step toward more efficient viral vaccine production processes only recently. Compared to conventional batch processes, fed-batch and perfusion strategies could result in ten to a hundred times higher product yields. Both cultivation strategies can be implemented to achieve cell concentrations exceeding 10⁷ cells/mL or even 10⁸ cells/mL, while keeping low levels of metabolites that potentially inhibit cell growth and virus replication. The trend towards HCD processes is supported by development of GMP-compliant cultivation platforms, i.e., acoustic settlers, hollow fiber bioreactors, and hollow fiber-based perfusion systems including tangential flow filtration (TFF) or alternating tangential flow (ATF) technologies. In this review, these process modes are discussed in detail and compared with conventional batch processes based on productivity indicators such as space-time yield, cell concentration, and product titers. In addition, options for the production of viral vaccines in continuous multi-stage bioreactors such as two- and three-stage systems are addressed. While such systems have shown similar virus titers compared to batch cultivations, keeping high yields for extended production times is still a challenge. Overall, we demonstrate that process intensification of cell culture-based viral vaccine production can be realized by the consequent application of fed-batch, perfusion, and continuous systems with a significant increase in productivity. The potential for even further improvements is high, considering recent developments in establishment of new (designer) cell lines, better characterization of host cell metabolism, advances in media design, and the use of mathematical models as a tool for process optimization and control.

The increasing demand for biopharmaceuticals produced in mammalian cells has driven the industry to enhance productivity of bioprocesses through different strategies. This is why fed-batch and perfusion cultures are considered more attractive choices than batch processes. In this context, the availability of reliable online measuring systems for cell density and metabolic activity estimation will help the application of these processes. The present work focuses on the comparison of two different monitoring tools for indirect estimation of biomass concentration in a HEK293 cell cultures producing IFN-γ: on one side, the oxygen uptake rate (O.U.R.) determination, by means of application of the dynamic method measurement which is already a widely used tool and, on the other side, a new robust online monitoring tool based on the alkali buffer addition used to maintain the pH set point. Both strategies allow a proper monitoring of cell growth and metabolic activity, with precise identification of the balanced cell growth and the most important action in the process, as is the media feeding. The application of these monitoring systems in fed-batch processes allows extending the growth of HEK293 cells, which in turn results in higher final cell concentrations compared with Batch strategy (7 · 106 cells mL−1), achieving 14 · 106 cells mL−1 for the fed-batch based on O.U.R. and 19 · 106 cells mL−1 for the fed-batch based on the alkali addition. Product titter is also increased in respect of the batch strategy (3.70 mg L−1), resulting in 8.27 mg L−1 when fed-batch was based on O.U.R. and 11.49 mg L−1 when it was based on the alkali buffer strategy. Results prove that fed-batch strategy based on the alkali buffer addition is a robust online monitoring method that has shown its great potential to optimize the feeding strategy in HEK293 fed-batch cultures.

The effects of biological factors including dissolved oxygen (DO), pH, carbon/nitrogen (C/N) and hydraulic retention times (HRT) on the performance of simultaneous nitrification and denitrification (SND) in a moving bed sequencing batch reactor (MBSBR) were investigated. A low DO was found to be advantageous to the SND in that nitrification was not inhibited, while pH and C/N ratio were shown to have positive effects on SND, and HRT needed to be controlled in a suitable range. A desirable SND efficiency was obtained at a DO of 2.5 mg L−1, pH of approximately 8.0, C/N ratio of 10 and HRT of 10 h in the MBSBR. High-throughput sequencing analysis showed that different operating conditions impacted microbial communities, resulting in different nitrogen removal mechanisms. Autotrophic and heterotrophic nitrification together contributed to the good nitrification performance, while denitrification was conducted by combined anoxic and aerobic processes. Furthermore, the results of principal component analyses (PCA) and the abundance of the predominant nitrification and denitrification genera both showed that DO and HRT might be regarded as the dominant variable factors influencing community structure analysis during SND, while the linear discriminant analysis (LDA) effect size (LEfSe) algorithm showed differences in abundance among the biofilm microbial communities with different DO. Overall, the results of this study improve our understanding of the bacterial community structure with different operating conditions in MBSBRs.

Municipal wastewater treatment plants (WWTPs) have been identified as a key source of microplastics (MPs) release into rivers and oceans. Varied extents of MPs pollution have been observed at different WWTPs with limited information in Thailand. This research aimed to study the occurrence of MPs in municipal WWTPs in Thailand by measuring MPs of three WWTPs which employ different treatment process. The WWTPs were selected to represent MP pollution from urbanized and suburbanized areas with different treatment set-ups, i.e., sequence batch reactor (WWTP-A), oxidation ditch (WWTP-B), and conventional activated sludge (WWTP-C). Water and sludge sampling was performed at the inlet and outlet of primary and secondary treatment units. The results indicate that the average MPs removal efficiency of the WWTPs was ca. 84%, with the aeration tank as the main removal unit by transferring MPs from the water phase to sludge. Primary treatment comprising of screening and grit chambers could not remove MPs effectively. Most of the MPs observed in the WWTPs were fibers (32–57%), mostly made of polyester, polyethylene, polyacrylate, and polypropylene. From these results, it is suggested to implement tertiary treatment options to improve MPs removal efficiency in WWTPs, and to apply post-treatment to the WWTPs’ raw sludge to prevent the MPs’ release into the environment when the sludge is applied to agricultural land.

The simulations and predictions obtained from mathematical models of bioprocesses conducted by microorganisms are not overvalued. Mechanistic models are bringing a better process understanding and the possibility of simulating unmeasurable variables. The Dynamic Energy Budget (DEB) model is an energy balance that can be formulated for any living organism and can be classified as a structured model. In this study, the DEB model was used to describe E. coli growth in a batch reactor in carbon and nitrogen substrate limitation conditions. The DEB model provides a possibility to follow the changes in the microbes’ cells including their elemental composition and content of some important cell ingredients in different growth phases in substrate limitation conditions which makes it more informative compared to Monod’s model. The model can be used as an optimal choice between Monod-like models and flux-based approaches. Key points • The DEB model can be used to catch changes in elemental composition of E. coli • Bacteria batch culture growth phases can be explained by the DEB model • The DEB model is more informative compared to Monod’s based models

The relationship of microbial community and cometabolic consumption of 2-chlorophenol (2-CP) in a nitrifying sequencing batch reactor (SBR) was studied. The assessment of the population dynamics of the nitrifying sludge during the cometabolic 2-CP consumption with increasing ammonium (NH 4 + ) concentrations in the SBR showed the presence of 39 different species of which 10 were always present in all cycles. Fifty-five percent of the species found were grouped as Proteobacteria (45% as β-proteobacteria and 10% as γ-proteobacteria class), 30% as Acidobacteria, and 15% as Deinococcus-Thermus phyla. NH 4 + and cometabolic 2-CP consumption could be related to the presence and permanence of ammonium-oxidizing bacteria (AOB) species and heterotrophic bacteria, while the complete nitrification to the presence of nitrite-oxidizing bacteria (NOB) species. A correlation analysis showed that the complete and stable nitrifying performance (NH 4 + consumption efficiencies (ENH 4 + -N) > 99% and nitrate production yields (YNO 3 − -N) between 0.93 and 0.99), as well as the increase in specific rates (ammonium (qNH 4 + -N) and 2-CP (q2-CP-C) consumption and nitrate production (qNO 3 − -N)), was associated with the homogeneity of the bacterial community ( J index = 0.99). The increase in the proportion of individuals of AOB species such as Nitrosomonas oligotropha and Nitrosomonas marina was associated with the increase in qNH 4 + -N ( r  ≥ 0.69) and q2-CP-C ( r  ≥ 0.64) and, therefore, with the 2-CP cometabolic consumption in the SBR. Finally, the increase in the proportion of individuals of heterotrophic species such as Dokdonella ginsengisoli , Deinococcus peraridilitoris , Truepera radiovictrix , and Stenotrophobacter terrae was associated with the increase in q2-CP-C ( r  ≥ 0.59). Key points • Thirty-nine bacterial species were identified in the nitrifying sludge population of the SBR. • β-Proteobacteria and Acidobacteria were the prevalent (85%) bacterial groups. • AOB and heterotrophic bacteria participate in NH 4 + and cometabolic 2-CP consumption. Graphical abstract

Purpose Fish waste was studied as a raw material for the simultaneous production of protein hydrolysates, collagen and fish oil. Enzymatic hydrolysis was selected for recovering these by-products with high value-added. Methods Alcalase 2.4 L was used to hydrolyze fish waste in a batch reactor under controlled conditions (180 min, 50 °C and pH 8). The influence of hydrolysis degree on by-products recovery was analyzed for different enzyme and substrate concentrations. Results Results suggested that the enzyme/substrate ratio was the main factor controlling the hydrolysis rate. Linear relationships were found between the degree of hydrolysis and the amount of each of the obtained by-products. From these relationships, the amounts of by-products with high added value can be predicted by only knowing the degree of hydrolysis reached. In optimal conditions (DH = 25%), 430 g of protein hydrolysate, 10 g of collagen and 350 g of oil could be obtained from 1000 g of fish waste. The use of fish waste as raw material for by-product fabrication resulted in a 79% reduction of waste disposed to landfill. Conclusion Therefore, this study shows the enzymatic hydrolysis of fish waste as a feasible solution to obtain high value-added products and an alternative to landfilling disposal. Graphic Abstract

Wastewater treatment plants (WWTPs) are one of the most important ways of releasing microplastics (MPs) into the environment. In this study, the size, number, color, and shape of MPs during the wastewater treatment process were investigated in six WWTPs with different processes, which include WWTPs A and B (activated sludge process, ASP), WWTP C and D (aerated lagoon, AL), WWTP E (sequencing batch reactor process, SBR), and WWTP F (stabilization pond, SP). The MP particles were detected by the polarized light microscopy. In all six WWTPs, the clear color was observed as the dominant color in the effluent. Among the forms of MPs, fibers had the highest removal efficiency in WWTPs A (97.3%), B (99.2%), C (95.5%), and D (94.3%). In both WWTPs E and F, the highest removal rate of MP shapes was related to films (96.1%) and granules (86.1%), respectively. MPs with size 25–125 μm had the highest amount (0.39 to 4.08 MP/L) in the effluent of WWTPs compared to larger sizes. With respect to the type of the wastewater treatment process, the number of MPs during the treatment process decreased from 3.75–25.31 to 0.51–6.28MP/L. Based on the results of this study, ASP with a removal rate of 91.87% had the highest efficiency compared to other processes. However, daily 4.95 × 104 to 1.49 × 108 MP enter the environment via the effluent of these WWTPs. The study recommends reducing the use of MPs and plasticizers in widely used products as much as possible and replacing them with nature-friendly materials.

Polyphosphate accumulating organisms (PAOs) are responsible for enhanced biological phosphate removal (EBPR) from wastewater, where they grow embedded in a matrix of extracellular polymeric substances (EPS). EPSs comprise a mixture of biopolymers like polysaccharides or (glyco)proteins. Despite previous studies, little is known about the dynamics of EPS in mixed cultures, and their production by PAOs and potential consumption by flanking microbes. EPSs are biodegradable and have been suggested to be a substrate for other organisms in the community. Studying EPS turnover can help elucidate their biosynthesis and biodegradation cycles. We analyzed the turnover of proteins and polysaccharides in the EPS of an enrichment culture of PAOs relative to the turnover of internal proteins. An anaerobic-aerobic sequencing batch reactor (SBR) simulating EBPR conditions was operated to enrich for PAOs. After achieving a stable culture, carbon source was switched to uniformly 13C-labeled acetate. Samples were collected at the end of each aerobic phase. EPSs were extracted by alkaline treatment. 13C enrichment in proteins and sugars (after hydrolysis of polysaccharides) in the extracted EPS were measured by mass spectrometry. The average turnover rate of sugars and proteins (0.167 and 0.192 d−1 respectively) was higher than the expected value based on the solid removal rate (0.132 d−1), and no significant difference was observed between intracellular and extracellular proteins. This indicates that EPS from the PAO enriched community is not selectively degraded by flanking populations under stable EBPR process conditions. Instead, we observed general decay of biomass, which corresponds to a value of 0.048 d−1.Key Points• Proteins showed a higher turnover rate than carbohydrates.• Turnover of EPS was similar to the turnover of intracellular proteins.• EPS is not preferentially consumed by flanking populations.