Danish Wastewater Treatment Plants Research Articles

Toward smart wastewater treatment plants: a novel data-driven sludge blanket model based on stochastic differential equations

ABSTRACT A novel data-driven model for forecasting the sludge blanket height in secondary clarifiers is presented. The model is trained on sensor measurements of the sludge blanket height and used as inputs such as (1) the clarifier feed flow rate, (2) feed suspended solids concentration, and (3) the clarifier recycle flow rate. The model’s prediction accuracy is evaluated based on data from two Danish wastewater treatment plants by means of root-mean-square errors (RMSEs), and results are compared against a persistence model. We demonstrate that the developed model is superior to the persistence forecast at both plants during high blanket dynamics. In the best scenario, the model improves the RMSE by 0.1/0.4 m at prediction horizons of 2.5/10 h, assuming known inputs. The model performance is subsequently considered with forecasted inputs using two different forecast scenarios. We discuss differences in the two plants’ performance and requirements to achieve good model performance. The model is well-suited for a model predictive control strategy, whose purpose ultimately is to improve clarifier control, increasing hydraulic capacity and reducing overflow suspended solids.

Exploring data quality and seasonal variations of N2O in wastewater treatment: a modeling perspective

Abstract In this work, operational data collected from four Danish wastewater treatment plants (WWTP) are assessed for quality issues and analyzed to investigate the feasibility of data-driven modeling for control purposes. All plants have permanent N2O sensors installed in the biological reactors, and N2O data are collected on the same terms as other operational data. We present and deploy a six-dimensional data quality assessment to the operational data evaluating (1) relevance, (2) accuracy, (3) completeness, (4) consistency, (5) comparability, and (6) accessibility. To increase the accuracy and completeness of the stored data, it is suggested that future initiatives are taken toward the collection and storing of metadata in WWTPs. Furthermore, seasonal variations and time-varying relationships between N2O, nitrogenous variables, and oxygen are investigated and compared across various case plants and process designs. Results show that the quality of the operational data varies substantially between plants. The investigation of time-varying interrelation between N2O and nitrogenous variables showed no clear pattern within or across different case plants. Furthermore, it is recommended that future research should consider adapting models so that more influence is linked to reliable measurements, contrary to assuming that all variables are of equal quality.

Genetic potential for exopolysaccharide synthesis in activated sludge bacteria uncovered by genome-resolved metagenomics

A good floc formation of activated sludge (AS) is crucial for solid-liquid separation and production of clean effluent during wastewater treatment. Floc formation is partly controlled by self-produced extracellular polymeric substances (EPS) such as exopolysaccharides, proteins, and nucleic acids. Little is known about the composition, structure, and function of EPS in AS and which bacteria produce them. To address this knowledge gap for the exopolysaccharides, we took advantage of 1083 high-quality metagenome-assembled genomes (MAGs) obtained from 23 Danish wastewater treatment plants. We investigated the genomic potential for exopolysaccharide biosynthesis in bacterial species typical in AS systems based on genome mining and gene synteny analyses. Putative gene clusters associated with the biosynthesis of alginate, cellulose, curdlan, diutan, hyaluronic acids, Pel, poly-β-1,6-N-acetyl-d-glucosamine (PNAG), Psl, S88 capsular polysaccharide, salecan, succinoglycan, and xanthan were identified and linked to individual MAGs, providing a comprehensive overview of the genome-resolved potential for these exopolysaccharides in AS bacteria. The approach and results provide a starting point for a more comprehensive understanding of EPS composition in wastewater treatment systems, which may facilitate a more refined regulation of the activated sludge process for improved stability.

Diversity and Ecophysiology of the Genus OLB8 and Other Abundant Uncultured Saprospiraceae Genera in Global Wastewater Treatment Systems.

The Saprospiraceae family within the phylum Bacteroidota is commonly present and highly abundant in wastewater treatment plants (WWTPs) worldwide, but little is known about its role. In this study, we used MiDAS 4 global survey with samples from 30 countries to analyze the abundance and distribution of members of Saprospiraceae. Phylogenomics were used to delineate five new genera from a set of 31 high-quality metagenome-assembled genomes from Danish WWTPs. Newly designed probes for fluorescence in situ hybridization (FISH) revealed rod-shaped morphologies for all genera analyzed, including OLB8, present mostly inside the activated sludge flocs. The genomes revealed potential metabolic capabilities for the degradation of polysaccharides, proteins, and other complex molecules; partial denitrification; and storage of intracellular polymers (glycogen, polyphosphate, and polyhydroxyalkanoates). FISH in combination with Raman microspectroscopy confirmed the presence of intracellular glycogen in Candidatus Brachybacter, Candidatus Parvibacillus calidus (both from the former genus OLB8), and Candidatus Opimibacter, and the presence of polyhydroxyalkanoates in Candidatus Defluviibacterium haderslevense and Candidatus Vicinibacter. These results provide the first overview of the most abundant novel Saprospiraceae genera present in WWTPs across the world and their potential involvement in nutrient removal and the degradation of macromolecules.

Low Global Diversity of Candidatus Microthrix, a Troublesome Filamentous Organism in Full-Scale WWTPs.

Candidatus Microthrix is one of the most common bulking filamentous microorganisms found in activated sludge wastewater treatment plants (WWTPs) across the globe. One species, Ca. M. parvicella, is frequently observed, but global genus diversity, as well as important aspects of its ecology and physiology, are still unknown. Here, we use the MiDAS ecosystem-specific 16S rRNA gene database in combination with amplicon sequencing of Danish and global WWTPs to investigate Ca. Microthrix spp. diversity, distribution, and factors affecting their global presence. Only two species were abundant across the world confirming low diversity of the genus: the dominant Ca. M. parvicella and an unknown species typically present along with Ca. M. parvicella, although usually in lower abundances. Both species were mostly found in Europe at low-to-moderate temperatures and their growth was favored in municipal WWTPs with advanced process designs. As no isolate is available for the novel species, we propose the name “Candidatus Microthrix subdominans.” Ten high-quality metagenome-assembled genomes recovered from Danish WWTPs, including 6 representing the novel Ca. M. subdominans, demonstrated high genetic similarity between the two species with a likely preference for lipids, a putative capability to reduce nitrate and nitrite, and the potential to store lipids and poly-P. Ca. M. subdominans had a potentially more versatile metabolism including additional sugar transporters, higher oxygen tolerance, and the potential to use carbon monoxide as energy source. Newly designed fluorescence in situ hybridization probes revealed similar filamentous morphology for both species. Raman microspectroscopy was used to quantify the in situ levels of intracellular poly-P. Despite the observed similarities in their physiology (both by genomes and in situ), the two species showed different seasonal dynamics in Danish WWTPs through a 13-years survey, possibly indicating occupation of slightly different niches. The genomic information provides the basis for future research into in situ gene expression and regulation, while the new FISH probes provide a useful tool for further characterization in situ. This study is an important step toward understanding the ecology of Ca. Microthrix in WWTPs, which may eventually lead to optimization of control strategies for its growth in this ecosystem.

High Diversity and Functional Potential of Undescribed "Acidobacteriota" in Danish Wastewater Treatment Plants.

Microbial communities in water resource recovery facilities encompass a large diversity of poorly characterized lineages that could have undescribed process-critical functions. Recently, it was shown that taxa belonging to “Acidobacteriota” are abundant in Danish full-scale activated sludge wastewater treatment plants (WWTP), and here we investigated their diversity, distribution, and functional potential. “Acidobacteriota” taxa were identified using a comprehensive full-length 16S rRNA gene reference dataset and amplicon sequencing surveys across 37 WWTPs. Members of this phylum were diverse, belonging to 14 families, eight of which are completely uncharacterized and lack type strains. Several lineages were abundant, with relative abundances of up to 5% of the microbial community. Genome annotation and metabolic reconstruction of 50 high-quality “Acidobacteriota” metagenome-assembled genomes (MAGs) from 19 WWTPs showed high metabolic diversity and potential involvement in nitrogen and phosphorus removal and iron reduction. Fluorescence in situ hybridization (FISH) using newly-designed probes revealed cells with diverse morphologies, predominantly located inside activated sludge flocs. FISH in combination with Raman microspectroscopy revealed ecophysiological traits in probe-defined cells from the families Holophagaceae, Thermoanaerobaculaceae, and Vicinamibacteraceae, and families with the placeholder name of midas_f_502, midas_f_973, and midas_f_1548. Members of these lineages had the potential to be polyphosphate-accumulating organisms (PAOs) as intracellular storage was observed for the key compounds polyphosphate and glycogen.

Characterizing the growing microorganisms at species level in 46 anaerobic digesters at Danish wastewater treatment plants: A six-year survey on microbial community structure and key drivers

Anaerobic digestion (AD) is a key technology at many wastewater treatment plants (WWTPs) for converting primary and surplus activated sludge to methane-rich biogas. However, the limited number of surveys and the lack of comprehensive datasets have hindered a deeper understanding of the characteristics and associations between key variables and the microbial community composition. Here, we present a six-year survey of 46 anaerobic digesters, located at 22 WWTPs in Denmark, which is the first and largest known study of the microbial ecology of AD at WWTPs at a regional scale. For three types of AD (mesophilic, mesophilic with thermal hydrolysis pretreatment, and thermophilic), we present the typical value range of 12 key parameters including operational variables and performance parameters. High-resolution bacterial and archaeal community analyses were carried out at species level using amplicon sequencing of >1,000 samples and the new ecosystem-specific MiDAS 3 reference database. We detected 42 phyla, 1,600 genera, and 3,584 species in the bacterial community, where 70% of the genera and 93% of the species represented environmental taxa that were only classified based on MiDAS 3 de novo placeholder taxonomy. More than 40% of the bacterial species were found not to grow in the mesophilic and thermophilic digesters and were only present due to immigration with the feed sludge. Ammonium concentration was the main driver shaping the bacterial community while temperature and pH were main drivers for the archaea in the three types of ADs. Sub-setting for the growing microbes improved significantly the correlation analyses and revealed the main drivers for the presence of specific species. Within mesophilic digesters, feed sludge composition and other key parameters (organic loading rate, biogas yield, and ammonium concentration) correlated with specific growing species. This survey provides a comprehensive insight into community structure at species level, providing a foundation for future studies of the ecological significance/characteristics and function of the many novel or poorly described taxa.

Candidatus Amarolinea and Candidatus Microthrix Are Mainly Responsible for Filamentous Bulking in Danish Municipal Wastewater Treatment Plants.

Filamentous bulking is a common serious operational problem leading to deteriorated sludge settling that has long been observed in activated sludge biological wastewater treatment systems. A number of bacterial genera found therein possess filamentous morphology, where some have been shown to be implicated in bulking episodes (e.g., Ca. Microthrix), the impact of many others is still not clear. In this study we performed a survey of 17 Danish municipal wastewater treatment plants (WWTPs) with nutrient removal using 16S rRNA amplicon sequencing over a period of 13 years, where all known filamentous bacteria from 30 genera were analyzed. The filamentous community constituted on average 13 ± 6%, and up to 43% of total read abundance with the same genera common to all plants. Ca. Microthrix and several genera belonging to phylum Chloroflexi were among the most abundant filamentous bacteria. The effect of filamentous bacteria on sludge settling properties was analyzed using measurements of the diluted sludge volume index (DSVI). Strong positive correlations with DSVI were observed only for Ca. Microthrix and Ca. Amarolinea, the latter being a novel, recently characterized genus belonging to the phylum Chloroflexi. The bulking potential of other filamentous bacteria was not significant despite their presence in many plants. Low phylogenetic diversity was observed for both Ca. Microthrix and Ca. Amarolinea, making physiological characterization of individual species and potential development of control strategies more feasible. In this study we show that, despite the high diversity of filamentous phylotypes in Danish WWTPs, only few of them were responsible for severe bulking episodes.

MiDAS 3: An ecosystem-specific reference database, taxonomy and knowledge platform for activated sludge and anaerobic digesters reveals species-level microbiome composition of activated sludge

The function of the microbiomes in wastewater treatment systems and anaerobic digesters is dictated by the physiological activity of their members and complex interactions between them. Since functional traits are often conserved at low taxonomic ranks (genus, species, strain), high resolution taxonomic classification is crucial to understand the role of microbes in any ecosystem. Here we present MiDAS 3, a comprehensive 16S rRNA gene reference database based on full-length 16S rRNA gene amplicon sequence variants (FL-ASVs) derived from activated sludge and anaerobic digester systems in Denmark. The new database proposes unique provisional names for all unclassified microorganisms down to species level, providing a new and much-needed tool for microbiome research. The MiDAS 3 database was used to analyze the microbiome in 20 Danish wastewater treatment plants with nutrient removal, sampled over 13 years. The 50 most abundant species belonged to 42 genera, including 14 genera with provisional ‘midas’ name. Of those, 20 have no known function in the system, which highlights the need for more efforts towards elucidating the role of important members of wastewater treatment ecosystems. The new MiDAS 3 database also forms the backbone of the MiDAS Field Guide – an online resource linking the identity of microorganisms in wastewater treatment systems to available data related to their functional importance. The new field guide contains a complete list of genera (>1800) and species (>4200) found in activated sludge and anaerobic digesters in Denmark, but is also relevant to wastewater systems across the world. The identity of the microbes is linked to functional information, where available, and the website provides the possibility to BLAST new sequences against the MiDAS 3 database. The MiDAS Field Guide is a collaborative platform acting as an online knowledge repository, facilitating understanding of wastewater treatment ecosystem function.

Bacteria from the Genus Arcobacter Are Abundant in Effluent from Wastewater Treatment Plants.

Pathogenic bacteria in wastewater are generally considered to be efficiently removed in biological wastewater treatment plants. This understanding is almost solely based on culture-based control measures, and here we show, by applying culture-independent methods, that the removal of species in the genus Arcobacter was less effective than for many other abundant genera in the influent wastewater. Arcobacter was one of the most abundant genera in influent wastewater at 14 municipal wastewater treatment plants and was also abundant in the "clean" effluent from all the plants, reaching up to 30% of all bacteria as analyzed by 16S rRNA gene amplicon sequencing. Metagenomic analyses, culturing, genome sequencing of Arcobacter isolates, and visualization by fluorescent in situ hybridization (FISH) confirmed the presence of the human-pathogenic Arcobacter cryaerophilus and A. butzleri in both influent and effluent. The main reason for the high relative abundance in the effluent was probably that Arcobacter cells, compared to those of other abundant genera in the influent, did not flocculate and attach well to the activated sludge flocs, leaving a relatively large fraction dispersed in the water phase. The study shows there is an urgent need for new standardized culture-independent measurements of pathogens in effluent wastewaters, e.g., amplicon sequencing, and an investigation of the problem on a global scale to quantify the risk for humans and livestock.IMPORTANCE The genus Arcobacter was unexpectedly abundant in the effluent from 14 Danish wastewater treatment plants treating municipal wastewater, and the species included the human-pathogenic A. cryaerophilus and A. butzleri Recent studies have shown that Arcobacter is common in wastewater worldwide, so the study indicates that discharge of members of the genus Arcobacter may be a global problem, and further studies are needed to quantify the risk and potentially minimize the discharge. The study also shows that culture-based analyses are insufficient for proper effluent quality control, and new standardized culture-independent measurements of effluent quality encompassing most pathogens should be considered.

Novel syntrophic bacteria in full-scale anaerobic digesters revealed by genome-centric metatranscriptomics

Short-chain fatty acid (SCFA) degradation is an important process in methanogenic ecosystems, and is usually catalyzed by SCFA-oxidizing bacteria in syntrophy with methanogens. Current knowledge of this functional guild is mainly based on isolates or enrichment cultures, but these may not reflect the true diversity and in situ activities of the syntrophs predominating in full-scale systems. Here we obtained 182 medium to high quality metagenome-assembled genomes (MAGs) from the microbiome of two full-scale anaerobic digesters. The transcriptomic response of individual MAG was studied after stimulation with low concentrations of acetate, propionate, or butyrate, separately. The most pronounced response to butyrate was observed for two MAGs of the recently described genus Candidatus Phosphitivorax (phylum Desulfobacterota), expressing a butyrate beta-oxidation pathway. For propionate, the largest response was observed for an MAG of a novel genus in the family Pelotomaculaceae, transcribing a methylmalonyl-CoA pathway. All three species were common in anaerobic digesters at Danish wastewater treatment plants as shown by amplicon analysis, and this is the first time their syntrophic features involved in SCFA oxidation were revealed with transcriptomic evidence. Further, they also possessed unique genomic features undescribed in well-characterized syntrophs, including the metabolic pathways for phosphite oxidation, nitrite and sulfate reduction.

The Proteome of Tetrasphaera elongata is adapted to Changing Conditions in Wastewater Treatment Plants.

The activated sludge in wastewater treatment plants (WWTP) designed for enhanced biological phosphorus removal (EBPR) experiences periodically changing nutrient and oxygen availability. Tetrasphaera is the most abundant genus in Danish WWTP and represents up to 20–30% of the activated sludge community based on 16S rRNA amplicon sequencing and quantitative fluorescence in situ hybridization analyses, although the genus is in low abundance in the influent wastewater. Here we investigated how Tetrasphaera can successfully out-compete most other microorganisms in such highly dynamic ecosystems. To achieve this, we analyzed the physiological adaptations of the WWTP isolate T. elongata str. LP2 during an aerobic to anoxic shift by label-free quantitative proteomics and NMR-metabolomics. Escherichia coli was used as reference organism as it shares several metabolic capabilities and is regularly introduced to wastewater treatment plants without succeeding there. When compared to E. coli, only minor changes in the proteome of T. elongata were observed after the switch to anoxic conditions. This indicates that metabolic pathways for anaerobic energy harvest were already expressed during the aerobic growth. This allows continuous growth of Tetrasphaera immediately after the switch to anoxic conditions. Metabolomics furthermore revealed that the substrates provided were exploited far more efficiently by Tetrasphaera than by E. coli. These results suggest that T. elongata prospers in the dynamic WWTP environment due to adaptation to the changing environmental conditions.

The morphology and metabolic potential of the Chloroflexi in full-scale activated sludge wastewater treatment plants.

Filamentous bacteria belonging to the phylum Chloroflexi have received considerable attention in wastewater treatment systems for their suggested role in the operational problem of impaired sludge settleability known as bulking. Their consistently high abundance in full-scale systems, even in the absence of bulking, indicates that they make a substantial contribution to the nutrient transformations during wastewater treatment. In this study, extensive 16S rRNA amplicon surveys of Danish wastewater treatment plants (WWTPs) with nutrient removal were screened to identify numerically important Chloroflexi genera. Fluorescence in situ hybridization probes were designed for their in situ characterization. All abundant Chloroflexi phylotypes were putatively identified as facultative anaerobic chemoorganotrophs involved in sugar fermentation. They were all filamentous but differed in their morphology and spatial arrangement. 'Candidatus Villigracilis' was predominantly located within the activated sludge flocs, where they possibly have structural importance, and their abundance was relatively stable. Conversely, the abundance of 'Candidatus Amarolinea' was highly dynamic, relative to other genera, sometimes reaching abundances in excess of 30% of the biovolume, suggesting their likely role in bulking episodes. This study gives an important insight into the role of Chloroflexi in WWTPs, thus contributing to the broader goal of understanding the ecology of these biotechnologically important systems.

Quantification of microplastic mass and removal rates at wastewater treatment plants applying Focal Plane Array (FPA)-based Fourier Transform Infrared (FT-IR) imaging

This paper presents a method for microplastic (MP) mass quantification using a Focal Plane Array-based Fourier Transform Infrared imaging technique. It discusses the issue that particle number is not a conserved base quantity and hence less suited than mass to compare independent studies on MP in the environment. It concludes that MP mass should be included when quantifying MP pollution in the environment, supplementing the conventional approach of reporting particle numbers. Applying mass as the unit of MP measurement, the paper presents data showing that Danish wastewater treatment plants discharge around 3 t/year of MP in the size range 10–500 μm. This value corresponds to an annual per capita emission from these plants of 0.56 g MP/(capita year). The distribution of polymer types by mass and particle number differed because the size of MP particles of the different material types varied.

Effects of thermal drying on phosphorus availability from iron‐precipitated sewage sludge

AbstractThermal drying of sewage sludge implies sanitation and improves practical handling options of the sludge prior to land application. However, it may also affect its value as a fertilizer. The objective of this study was to assess whether thermal drying of sewage sludge, as well as drying temperature, affects plant P availability after application to soil. The experiment included dewatered sewage sludge (20% DM) and thermally dried sewage sludge (95% DM) collected at a Danish wastewater treatment plant, as well as laboratory oven‐dried (70, 130, 190, and 250°C; DM > 95%) subsamples of the dewatered sludge, and a triple superphosphate as a reference. Plant P availability was studied in a 197 d soil incubation experiment, with sampling for Diffusive Gradients in Thin films (DGT) and water extractable P (WEP) analyses over time, and in a pot experiment with spring barley (Hordeum vulgare L.). In both experiments, thermal drying reduced P availability, as shown by 37 and 23% lower DGT and WEP values, respectively, and a 16% lower P uptake by barley in the pot experiment. The specific drying temperature did not appear to have much effect. Overall, our results suggest that thermal drying of iron‐precipitated sewage sludge is not an optimal treatment option if the aim is to optimize plant P availability.

Can DNA sequencing show differences between microbial communities in Polish and Danish wastewater treatment plants?

The microbial populations in the activated sludge of two Polish wastewater treatment plants (WWTPs) were identified and quantified using Illumina sequencing of 16S ribosomal RNA amplicons over a 2-year period. Their dynamics over time were compared to Danish WWTPs (data collected in previous studies by Center for Microbial Communities, Aalborg University). The bacterial communities in Polish and Danish WWTPs were similar to each other, but the microbial diversity in Polish WWTPs was lower. The dominant genera in Polish WWTPs were more abundant than in Danish WWTPs; 30 of them constituted more than half the of activated sludge community. Polish WWTPs showed a higher abundance of bacteria involved in nitrogen and chemical oxygen demand removal (Proteobacteria and Bacteroidetes), while polyphosphate-acculumating bacteria were the dominant bacterial group in Danish plants. The microbial community structures in the examined Polish WWTPs were relatively similar to each other and showed strong seasonal variations which are not normally observed in Danish WWTPs.

Increasing thermal drying temperature of biosolids reduced nitrogen mineralisation and soil N2O emissions.

Previous studies found that thermally dried biosolids contained more mineralisable organic nitrogen (N) than the raw or anaerobically digested (AD) biosolids they were derived from. However, the effect of thermal drying temperature on biosolid N availability is not well understood. This will be of importance for the value of the biosolids when used to fertilise crops. We sourced AD biosolids from a Danish waste water treatment plant (WWTP) and dried it in the laboratory at 70, 130, 190 or 250°C to >95% dry matter content. Also, we sourced biosolids from the WWTP dried using its in-house thermal drying process (input temperature 95°C, thermal fluid circuit temperature 200°C, 95% dry matter content). The drying process reduced the ammonium content of the biosolids and reduced it further at higher drying temperatures. These findings were attributed to ammonia volatilisation. The percentage of mineralisable organic N fraction (min-N) in the biosolids, and nitrous oxide (N2O) and carbon dioxide (CO2) production were analysed 120days after addition to soil. When incubated at soil field capacity (pF 2), none of the dried biosolids had a greater min-N than the AD biosolids (46.4%). Min-N was lowest in biosolids dried at higher temperatures (e.g. 19.3% at 250°C vs 35.4% at 70°C). Considering only the dried biosolids, min-N was greater in WWTP-dried biosolids (50.5%) than all of the laboratory-dried biosolids with the exception of the 70°C-dried biosolids. Biosolid carbon mineralisation (CO2 release) and N2O production was also the lowest in treatments of the highest drying temperature, suggesting that this material was more recalcitrant. Overall, thermal drying temperature had a significant influence on N availability from the AD biosolids, but drying did not improve the N availability of these biosolids in any case.

Degradation of PPCPs in activated sludge from different WWTPs in Denmark.

Pharmaceuticals and Personal care products (PPCPs) are often found in effluents from wastewater treatment plants (WWTPs) due to insufficient removal during wastewater treatment processes. To understand the factors affecting the removal of PPCPs in classical activated sludge WWTPs, the present study was performed to assess the removal of frequently occurring pharmaceuticals (Naproxen, Fenoprofen, Ketoprofen, Dichlofenac, Carbamazepine) and the biocide Triclosan in activated sludge from four different Danish WWTPs. The respective degradation constants were compared to operational parameters previous shown to be of importance for degradation of micropollutants such as biomass concentration, and sludge retention time (SRT). The most rapid degradation, was observed for NSAID pharmaceuticals (55-90% for Fenoprofen, 77-94% for Ketoprofen and 46-90% for Naproxen), followed by Triclosan (61-91%), while Dichlofenac and Carbamazepine were found to be persistent in the systems. Degradation rate constants were calculated as 0.0026-0.0407 for NSAID pharmaceuticals and 0.0022-0.0065 for triclosan. No relationships were observed between degradation rates and biomass concentrations in the diverse sludges. However, for the investigated PPCPs, the optimal SRT was within 14-20 days (for these values degradation of these PPCPs was the most efficient). Though all of these parameters influence the degradation rate, none of them seems to be overall decisive. These observations indicate that the biological composition of the sludge is more important than the design parameters of the respective treatment plant.

The activated sludge ecosystem contains a core community of abundant organisms.

Understanding the microbial ecology of a system requires that the observed population dynamics can be linked to their metabolic functions. However, functional characterization is laborious and the choice of organisms should be prioritized to those that are frequently abundant (core) or transiently abundant, which are therefore putatively make the greatest contribution to carbon turnover in the system. We analyzed the microbial communities in 13 Danish wastewater treatment plants with nutrient removal in consecutive years and a single plant periodically over 6 years, using Illumina sequencing of 16S ribosomal RNA amplicons of the V4 region. The plants contained a core community of 63 abundant genus-level operational taxonomic units (OTUs) that made up 68% of the total reads. A core community consisting of abundant OTUs was also observed within the incoming wastewater to three plants. The net growth rate for individual OTUs was quantified using mass balance, and it was found that 10% of the total reads in the activated sludge were from slow or non-growing OTUs, and that their measured abundance was primarily because of immigration with the wastewater. Transiently abundant organisms were also identified. Among them the genus Nitrotoga (class Betaproteobacteria) was the most abundant putative nitrite oxidizer in a number of activated sludge plants, which challenges previous assumptions that Nitrospira (phylum Nitrospirae) are the primary nitrite-oxidizers in activated sludge systems with nutrient removal.

The Microbial Database for Danish wastewater treatment plants with nutrient removal (MiDas-DK) – a tool for understanding activated sludge population dynamics and community stability

Since 2006 more than 50 Danish full-scale wastewater treatment plants with nutrient removal have been investigated in a project called 'The Microbial Database for Danish Activated Sludge Wastewater Treatment Plants with Nutrient Removal (MiDas-DK)'. Comprehensive sets of samples have been collected, analyzed and associated with extensive operational data from the plants. The community composition was analyzed by quantitative fluorescence in situ hybridization (FISH) supported by 16S rRNA amplicon sequencing and deep metagenomics. MiDas-DK has been a powerful tool to study the complex activated sludge ecosystems, and, besides many scientific articles on fundamental issues on mixed communities encompassing nitrifiers, denitrifiers, bacteria involved in P-removal, hydrolysis, fermentation, and foaming, the project has provided results that can be used to optimize the operation of full-scale plants and carry out trouble-shooting. A core microbial community has been defined comprising the majority of microorganisms present in the plants. Time series have been established, providing an overview of temporal variations in the different plants. Interestingly, although most microorganisms were present in all plants, there seemed to be plant-specific factors that controlled the population composition thereby keeping it unique in each plant over time. Statistical analyses of FISH and operational data revealed some correlations, but less than expected. MiDas-DK (www.midasdk.dk) will continue over the next years and we hope the approach can inspire others to make similar projects in other parts of the world to get a more comprehensive understanding of microbial communities in wastewater engineering.