A review on co-metabolic degradation of organic micropollutants during anaerobic digestion: Linkages between functional groups and digestion stages.

The emerging presence of organic micropollutants (OMPs) in water bodies produced by human activities is a source of growing concern due to their environmental and health issues. Biodegradation is a widely employed treatment method for OMPs in wastewater owing to its high efficiency and low operational cost. Compared to aerobic degradation, anaerobic degradation has numerous advantages, including energy efficiency and superior performance for certain recalcitrant compounds. Nonetheless, the low influent concentrations of OMPs in wastewater treatment plants (WWTPs) and their toxicity make it difficult to support the growth of microorganisms. Therefore, co-metabolism is a promising mechanism for OMP biodegradation in which co-substrates are added as carbon and energy sources and stimulate increased metabolic activity. Functional microorganisms and enzymes exhibit significant variations at each stage of anaerobic digestion affecting the environment for the degradation of OMPs with different structural properties, as these factors substantially influence OMPs' biodegradability and transformation pathways. However, there is a paucity of literature reviews that explicate the correlations between OMPs' chemical structure and specific metabolic conditions. This study provides a comprehensive review of the co-metabolic processes which are favored by each stage of anaerobic digestion and attempts to link various functional groups to their favorable degradation pathways. Furthermore, potential co-metabolic processes and strategies that can enhance co-digestion are also identified, providing directions for future research.

Thermocaproicibacter melissae gen. nov., sp. nov., a thermophilic chain-elongating bacterium, producing n-caproate from polymeric carbohydrates.

Strain MDTJ8T is a chain-elongating thermophilic bacterium isolated from a thermophilic acidogenic anaerobic digestor treating human waste while producing the high commodity chemical n-caproate. The strain grows and produces formate, acetate, n-butyrate, n-caproate and lactate from mono-, di- and polymeric saccharides at 37-60 °C (optimum, 50-55 °C) and at pH 5.0-7.0 (optimum, pH 6.5). The organism is an obligate anaerobe, is motile and its cells form rods (0.3-0.5×1.0-3.0 µm) that stain Gram-positive and occur primarily as chains. Phylogenetic analysis of both the 16S rRNA gene and full genome sequence shows that strain MDTJ8T belongs to a group that consists of mesophylic chain-elongating bacteria within the family Oscillospiraceae, being nearest to Caproicibacter fermentans EA1T (94.8 %) and Caproiciproducens galactitolivorans BS-1T (93.7 %). Its genome (1.96 Mbp) with a G+C content of 49.6 mol% is remarkably smaller than those of other chain-elongating bacteria of the family Oscillospiraceae. Pairwise average nucleotide identity and DNA-DNA hybridization values between strain MDJT8T and its mesophilic family members are less than 70 and 35 %, respectively, while pairwise average amino acid identity values are less than 68 %. In addition, strain MDJT8T uses far less carbohydrate and non-carbohydrate substrates compared to its nearest family members. The predominant cellular fatty acids of strain MDTJ8T are C14 : 0, C14 : 0 DMA (dimethyl acetal) and C16 : 0, while its polar lipid profile shows three unidentified glycophospholipids, 11 glycolipids, 13 phospholipids and six unidentified lipids. No respiratory quinones and polyamines are detected. Based on its phylogenetic, genotypic, morphological, physiological, biochemical and chemotaxonomic characteristics, strain MDTJ8T represents a novel species and novel genus of the family Oscillospiraceae and Thermocaproicibacter melissae gen. nov., sp. nov. is proposed as its name. The type strain is MDTJ8T (=DSM 114174T=LMG 32615T=NCCB 100883T).

Corrigendum to 'Amyloid adhesin production in activated sludge is enhanced in lab-scale sequencing batch reactors: Feeding regime impacts microbial community and amyloid distribution' [Water Research X 17 (2022) 100162].

[This corrects the article DOI: 10.1016/j.wroa.2022.100162.].

Quantification of hydrolysis activity in a biological wastewater treatment context.

This paper reviews currently available methods for hydrolysis activity monitoring of the most commonly encountered enzyme categories in biological wastewater treatment. While highlighting the relevant methods for protein, lipid, carbohydrate, organic phosphate, and ester hydrolysis, the discussion of their pros and cons is predominantly aimed at revealing the relevance of the to-be-hydrolyzed substrates that are used in the methods. These "substrates" should mimic the proteins, lipids, or other polymers that are present in the wastewater and are in the reviewed methods (i) real substrates (i.e., naturally present in the wastewater), (ii) chromogenic substrates, or (iii) fluorogenic substrates. We conclude that exploiting relevant substrates such as casein or starch, containing fluorophores, has the highest potential for meaningful high throughput hydrolysis quantification and that lipase activity monitoring is still cumbersome. Monitoring the hydrolysis activity in biological wastewater treatment systems is an underdeveloped area. With this review, which aims at providing a condensed and practice-oriented overview, we hope to facilitate the start or continuation of such monitoring. This monitoring will only grow in importance, given the transition from wastewater treatment plants towards water resource recovery facilities. KEY POINTS: • Colorimetric-based methods are vulnerable to sludge matrix interference. • Bonds in p-nitrophenol-based methods are not representative for the targeted substrates. • Direct methods with relevant/real substrates are preferred. • Fluorophore-containing (real) substrates enable high throughput screening.

Isolation and characterization of a thermophilic chain elongating bacterium that produces the high commodity chemical n-caproate from polymeric carbohydrates.

A thermophilic chain elongating bacterium, strain MDTJ8, was isolated from a thermophilic acidogenic anaerobic digestor producing n-caproate from human waste, growing optimally at 50-55 °C and pH 6.5. 16S rRNA gene analysis suggests that MDTJ8 represents a new species/genus within a group currently composed of mesophilic chain elongators of the Oscillospiraceae family. Genome analysis showed that strain MDTJ8 contains homologues of genes encoding for chain elongation and energy conservation but also indicated n-caproate production from carbohydrates including polymeric substances. This was confirmed by culturing experiments in which MDTJ8 converted, at pH 6.5 and 55 °C, mono-, di- and polymeric carbohydrates (starch and hemicellulose) to n-caproate reaching concentrations up to 283 mg/L and accounting for up to 10 % of the measured fermentation products. MDTJ8 is the first axenic organism that thermophilically performs chain elongation, opening doors to understand and intensify thermophilic bioprocesses targeting anaerobic digestion towards the production of the value-added chemical n-caproate.

Amyloid adhesin production in activated sludge is enhanced in lab-scale sequencing batch reactors: Feeding regime impacts microbial community and amyloid distribution.

Amyloid adhesins are ß-sheet-rich extracellular proteins thought to contribute to bioflocculation. They are present in activated sludge to varying extent. However, it remains unclear which operational conditions promote their production. To this end, the abundance and distribution of amyloids and their potential producers were monitored in two lab-scale reactors operated in sequencing batch mode with an unaerated and aerated reaction phase. Various feeding regimes ranging from feast-famine to nearly continuous feeding were applied. Thioflavin T staining revealed more amyloids in the lab-scale reactors during all operational stages compared to the full-scale industrial and municipal inocula. Furthermore, the feeding regime impacted the distribution of produced amyloids from dense clusters during feast-famine conditions towards a dispersed distribution during nearly continuous feeding. This dispersed presence did not negatively impact the bioflocculation (towards average floc size and shear sensitivity). 16S rRNA sequencing detected several known EPS and amyloid producers. More continuous and, hence, partially aerobic feeding promoted the relative abundance of denitrifiers. Sequential Thioflavin T staining and fluorescence in situ hybridization identified Zoogloea and Ca. Competibacter as potential amyloid producers under the applied conditions. This experiment confirms that amyloid producers need to be triggered for production and that the feeding regime impacts the microbial community composition, which in turn influences the amyloid production and distribution.

Integrated membrane bioreactors modelling: A review on new comprehensive modelling framework.

Integrated Membrane Bioreactor (MBR) models, combination of biological and physical models, have been representing powerful tools for the accomplishment of high environmental sustainability. This paper, produced by the International Water Association (IWA) Task Group on Membrane Modelling and Control, reviews the state-of-the-art, identifying gaps for future researches, and proposes a new integrated MBR modelling framework. In particular, the framework aims to guide researchers and managers in pursuing good performances of MBRs in terms of effluent quality, operating costs (such as membrane fouling, energy consumption due to aeration) and mitigation of greenhouse gas emissions.

A high-throughput assay to quantify protein hydrolysis in aerobic and anaerobic wastewater treatment processes.

Proteins, an important fraction of the organic matter in wastewater, typically enter a treatment facility as high molecular weight components. These components are degraded by extracellular protein hydrolytic enzymes, denoted as proteases. Adequate protein hydrolysis monitoring is crucial, since protein hydrolysis is often a rate-limiting step in wastewater treatment. However, current monitoring tools lack a high sample throughput and reliable quantification. Here, we present an improved assay for high-throughput protein hydrolysis rate measurements in wastewater treatment applications. A BODIPY FL casein model substrate was implemented in a microplate format for continuous fluorescent quantification. Case studies on a conventional and a high-rate aerobic municipal wastewater treatment plant and a lab-scale, two-stage, anaerobic reactor provided proof-of-concept. The assay presented in this study can help to obtain monitoring-based process insights, which will in turn allow improving biological performance of wastewater treatment installations in the future. KEY POINTS: • Protein hydrolysis is a crucial step in biological wastewater treatment. • Quantification of the protein hydrolysis rate enables in-depth process knowledge. • BODIPY FL casein is a suitable model substrate for a protein hydrolysis assay. • High sample throughput was obtained with fluorescent hydrolysis quantification. Graphical abstract.

Improving anaerobic digestion via direct interspecies electron transfer requires development of suitable characterization methods.

Recent anaerobic digestion studies commonly attribute performance improvements (e.g. increased methane production, enhanced process stability, reduced startup times) to direct interspecies electron transfer (DIET), even though only indirect evidence of DIET is available and DIET alone does not explain enhanced performance in many cases. This review evaluates methods believed to confirm the occurrence of DIET in anaerobic systems. 16S rRNA gene sequencing and meta-omics approaches are necessary to further DIET knowledge but are limited in their ability to confirm the occurrence of DIET. In situ use of cyclic voltammetry should be explored further, as well as microscopy and image analysis procedures to quantify stained cytochromes. Furthermore, linking interspecies distance, interspecies mixing, and cellular activity to a DIET-based electron transfer model is promising but needs further validation for anaerobic digestion systems. In short, a combination of methods is necessary to confirm the occurrence and expand our knowledge of DIET.

Ultrasound assisted chrome tanning: Towards a clean leather production technology.

Nowadays, there is a growing demand for a cleaner, but still effective alternative for production processes like in the leather industry. Ultrasound (US) assisted processing of leather might be promising in this sense. In the present paper, the use of US in the conventional chrome tanning process has been studied at different pH, temperature, tanning time, chrome dose and US exposure time by exposing the skin before tanning and during tanning operation. Both prior exposure of the skin to US and US during tanning improves the chrome uptake and reduces the shrinkage significantly. Prior exposure of the skin to US increase the chrome uptake by 13.8% or reduces the chrome dose from 8% to 5% (% based on skin weight) and shorten the process time by half while US during tanning increases the chrome uptake by 28.5% or reduces the chrome dose from 8% to 4% (half) and the tanning time to one third compared to the control without US. Concomitantly, the resulting leather quality (measured as skin shrinkage) improved from 5.2% to 3.2% shrinkage in the skin exposed to US prior tanning and to 1.3% in the skin exposed to US during the tanning experiment. This study confirms that US chrome tanning is an effective and eco-friendly tanning process which can produce a better quality leather product in a shorter process time with a lower chromium dose.

Impact of Tannery Effluent on the Self-purification Capacity and Biodiversity Level of a River.

The present study investigates the impact of tannery effluents on the self-purification capacity and the local macroinvertebrate community of one natural stream. As the concentration of chromium and sulfide increased from up- to downstream sites, the reduction of suspended solids, 5-days biological oxygen demand (BOD5), chemical oxygen demand and nitrification capacity decreased by 61 %, 21 %, 30 % and 74 %, respectively. Similarly, the share of Ephemeroptera, Plecoptera, and Trichoptera on the macroinvertebrate community decreased from 24 % to 0 %. Also the diversity (Simpson's) index and the correlation between the physicochemical parameters, BOD5 reduction, the macroinvertebrate abundance and the chromium concentration underpin the importance of the contamination by tannery effluents for the degradation of the stream habitat quality. In conclusion, although the physicochemical parameters indicate that the self-purification of the river can be maintained for a certain stream section, the biodiversity of the river is severely compromised.

Addition of polyaluminiumchloride (PACl) to waste activated sludge to mitigate the negative effects of its sticky phase in dewatering-drying operations.

This paper presents a new application of polyaluminiumchloride (PACl) as a conditioner for waste activated sludge prior its dewatering and drying. It is demonstrated at lab scale with a shear test-based protocol that a dose ranging from 50 to 150 g PACl/kg MLSS (mixed liquor suspended solids) mitigates the stickiness of partially dried sludge with a dry solids content between 25 and 60 %DS (dry solids). E.g., at a solids dryness of 46% DS the shear stress required to have the pre-consolidated sludge slip over a steel surface is reduced with 35%. The salient feature of PACl is further supported by torque data from a full scale decanter centrifuge used to dewater waste sludge. The maximal torque developed by the screw conveyor inside the decanter centrifuge is substantially reduced with 20% in the case the sludge feed is conditioned with PACl. The beneficial effect of waste sludge conditioning with PACl is proposed to be the result of the bound water associated with the aluminium polymers in PACl solutions which act as a type of lubrication for the intrinsically sticky sludge solids during the course of drying. It can be anticipated that PACl addition to waste sludge will become a technically feasible and very effective method to avoid worldwide fouling problems in direct sludge dryers, and to reduce torque issues in indirect sludge dryers as well as in sludge decanter centrifuges.

Identifying sustainable rehabilitation strategies for urban wastewater systems: a retrospective and interdisciplinary approach. Case study of Coronel Oviedo, Paraguay.

Many wastewater managers in developing countries struggle with the daily operation of urban wastewater systems. Technically well-designed wastewater collection and treatments are often degraded and/or not properly functioning. In this paper, a realistic rehabilitation strategy is developed for the urban wastewater system of Coronel Oviedo (Paraguay), in which the actual performance is unsatisfactory, as revealed by a detailed technical assessment, including water quantity and quality monitoring data. Understanding the history, starting from the initial planning and design process, allows to explain the current failing status of the urban wastewater system of Coronel Oviedo. The key information for the specific local rehabilitation strategy was extracted from an interdisciplinary assessment of shortcomings of urban wastewater systems in Paraguay which were revealed by a survey of all existing wastewater systems. Opting for a stepwise rehabilitation strategy allows the wastewater manager to gradually improve the performance of the wastewater system. Reusing the wastewater in agriculture and recovering the energy of methane gas are possible advantageous options for attracting external financial resources. Finally, the crucial role that the wastewater manager must play for sustainable wastewater management to become effective in practice is discussed, and recommendations are provided on how decision makers, researchers and consultants can contribute by anticipating the challenging circumstances inherent to developing countries.

Liver X receptors regulate cholesterol homeostasis in oligodendrocytes.

Cholesterol synthesis and transport in oligodendrocytes are essential for optimal myelination and remyelination in pathological conditions such as multiple sclerosis. However, little is known about cholesterol homeostasis in the myelin-forming oligodendrocytes. Liver X receptors (LXRs) are nuclear oxysterol receptors that regulate genes involved in cholesterol homeostasis and may therefore play an important role in de- and remyelination. We investigated whether LXRs regulate cholesterol homeostasis in oligodendrocytes. mRNA expression of genes encoding LXR-α and LXR-ß and their target genes (ABCA1, ABCG1, ABCG4, apoE, and LDLR) was detected in oligodendrocytes derived from both neonatal and adult rats using quantitative real-time PCR. The expression of LXR-ß and several target genes was increased during oligodendrocyte differentiation. We further demonstrated that treatment of primary neonatal rat oligodendrocytes with the synthetic LXR agonist T0901317 induced the expression of several established LXR target genes, including ABCA1, ABCG1, apoE, and LDLR. Treatment of oligodendrocytes with T0901317 resulted in an enhanced cholesterol efflux in the presence of apolipoprotein A-I or high-density lipoprotein particles. These data show that LXRs are involved in regulating cholesterol homeostasis in oligodendrocytes.

Simvastatin interferes with process outgrowth and branching of oligodendrocytes.

Statins have attracted interest as a treatment option for multiple sclerosis (MS) because of their pleiotropic antiinflammatory and immunomodulatory effects. However, contradictory results have been described when they are applied to oligodendrocytes (OLGs), the cell type predominantly affected in MS. In this study we focus on the in vitro effect of statins on process outgrowth in OLN-93 cells, a well-characterized OLG-derived cell line, and primary cultures of neonatal rat OLGs. Application of the lipophilic simvastatin, as low as 0.1-1 µM, disturbs process formation of both cell types, leading to less ramified cells. We show that both protein isoprenylation and cholesterol synthesis are required for the normal differentiation of OLGs. It is further demonstrated that the expression of 2',3'-cyclic-nucleotide-3' phosphodiesterase (CNP) and tubulin is lowered, concomitant with a reduction of membrane-bound CNP as well as tubulin. Therefore, we propose that lack of isoprenylation of CNP could help to explain the altered morphological and biochemical differentiation state of treated OLGs. Moreover, expression of specific myelin markers, such as myelin basic protein, myelin-associated glycoprotein, and myelin oligodendrocyte glycoprotein, was compromised after treatment. We conclude that simvastatin treatment has detrimental effects on OLG process outgrowth, the prior step in (re)myelination, thereby mortgaging long-term healing of MS lesions.

Analysis of mitochondrial pH and ion concentrations.

Detailed practical information is provided with emphasis on mapping cytosolic and mitochondrial pH, mitochondrial Na(+), and briefly also aspects related to mitochondrial Ca(2+) measurements in living cells, as grown on (un)coated glass coverslips. This chapter lists (laser scanning confocal) microscope instrumentation and setup requirements for proper imaging conditions, cell holders, and an easy-to-use incubator stage. For the daily routine of preparing buffer and calibration solutions, extensive annotated protocols are provided. In addition, detailed measurement and image analysis protocols are given to routinely obtain optimum results with confidence, while avoiding a number of typical pitfalls.

Morphological changes do not reflect biochemical and functional differentiation in OLN-93 oligodendroglial cells.

OLN-93 cells, a cell line established from spontaneously transformed rat brain glial cultures, are used as a model for oligodendrocytes. These cells are known to undergo morphological changes upon serum deprivation. The objective of the present study is to investigate a possible correlation between these morphological changes and (1) the loss or gain of oligodendrocyte markers and (2) the electrophysiological properties of these cells. Using RT-PCR and immunocytochemistry, we demonstrate that the OLN-93 cell line expresses a broad range of markers (NG2, CNP, MAG, MOG) both when cultured in medium containing 10% or 0.5% fetal calf serum. Whole-cell patch-clamp recordings demonstrate that, regardless of the culture conditions, OLN-93 cells mainly express delayed-rectifying K+ currents, a characteristic of immature oligodendrocytes. These currents are in part mediated by the shaker family of voltage-gated potassium channels. Kv1.1 and Kv1.3-expression are present at the mRNA and at the protein levels, and functional evidence for Kv1.3 mediated currents was obtained by using the selective blocker margatoxin. Under low serum conditions, OLN-93 cells exhibit differentiation-like morphological changes. However, we provide evidence that these morphological modifications do not necessarily correlate with biochemical or functional changes. Based on these data, we conclude that the OLN-93 cell line can be situated at a developmental stage between a late pre-oligodendrocyte and a late immature oligodendrocyte, regardless of serum concentration.

Assessment of activated sludge stability in lab-scale experiments.

The majority of activated sludge research is conducted in a laboratory environment with, most often, the start-up sludge being taken from a large-scale wastewater treatment plant. Inoculating this sludge in a lab-scale set-up induces a transient period, which, evidently, has a direct impact on the experimental results during this period of acclimatization. In the currently published literature, the acclimatization period is either neglected or fixed to two or three times the sludge age, without any guarantee that stable conditions are indeed reached. To develop a strategy that assesses the stability of activated sludge, three experiments were performed during which the activated sludge was extensively monitored through a series of physical, microscopic and biochemical analyses. It is demonstrated that it is possible to objectively quantify activated sludge stability through the monitoring of the total averaged filament length per image, the sludge volume index and the maximum specific oxygen uptake rate. Hereto, a moving window approach is adopted: within a 7 days interval the mean slope and the gap between the maximum and minimum value has to be smaller than a pre-specified threshold value. Once stability is reached, the true impact of test conditions can be studied without interference of adaptation phenomena.

Characterisation and optimisation of individual wastewater treatment systems.

Onsite individual wastewater treatment systems can provide a financially attractive alternative to a sewer connection in locations far from the existing sewer network. These systems are, however, relatively new, and practical experiences, especially long-term field studies, are lacking. Therefore, a thorough study of two compact biofilm-based, aerobic onsite systems, both of five population equivalents, was started in 2001. The assessment of the treatment performance of these systems, as well as the maintenance requirements and the characterisation of the feed are of great importance for the better understanding of the systems in order to optimise their design and performance. This paper presents an evaluation and discussion of the start-up and a starvation period of the two studied systems, followed by a characterisation of the incoming wastewater using activated sludge respirometry experiments in the context of the assessment and improvement of the denitrification process. Individual wastewater treatment systems are characterised by a rather long start-up period of 70-120 days. An important characteristic during the start-up is the nitrite peak, which indicates the initiation of the nitrification process. The respirometric experiments reveal that the failing denitrification is probably caused by an insufficient amount of readily biodegradable COD in the influent.

Efficient tracking of the dominant eigenspace of a normalized kernel matrix.

Various machine learning problems rely on kernel-based methods. The power of these methods resides in the ability to solve highly nonlinear problems by reformulating them in a linear context. The dominant eigenspace of a (normalized) kernel matrix is often required. Unfortunately, the computational requirements of the existing kernel methods are such that the applicability is restricted to relatively small data sets. This letter therefore focuses on a kernel-based method for large data sets. More specifically, a numerically stable tracking algorithm for the dominant eigenspace of a normalized kernel matrix is proposed, which proceeds by an updating (the addition of a new data point) followed by a downdating (the exclusion of an old data point) of the kernel matrix. Testing the algorithm on some representative case studies reveals that a very good approximation of the dominant eigenspace is obtained, while only a minimal amount of operations and memory space per iteration step is required.