Activated Sludge Microbial Community Research Articles

Contrasting distribution of antibiotic resistance genes and microbial communities in suspended activated sludge versus attached biofilms in an integrated fixed film activated sludge (IFAS) system

Suspended activated sludge (AS) and carrier-attached biofilms simultaneously exist in an integrated fixed film activated sludge (IFAS) system. However, the differentiation of antibiotic resistance genes (ARGs) and microbial communities in different types of biofilms is rarely reported. In this study, successions of ARGs and microbial communities of AS and two types of suspended carrier-attached biofilms over seasons were investigated in the IFAS system of one municipal wastewater treatment plant. Results showed that substantial differences were found in the distribution pattern of ARGs, bacterial communities, and predicted microbial function between AS and attached biofilms. The relative abundances of all detected ARGs in AS were significantly higher than those in attached biofilms. ARGs with higher relative abundances generally existed in K3 carrier (surface area ≥ 800 m2/m3) attached biofilms than those in K1 carrier (surface area ≥ 450 m2/m3) biofilms. The relative abundances of ARGs were negatively correlated with temperature and biochemical oxygen demand (BOD5) and positively correlated with ammonium nitrogen contents for AS but not for attached biofilms. No significant relationship was found between the extracellular polymeric substance (EPS) content and ARG abundance for all samples. Temperature, BOD5, and ammonium nitrogen contents were closely connected to microbial communities. The Bray–Curtis distance of bacterial communities between two adjacent sampling seasons for AS was larger than those of two attached biofilms. Network analysis indicated that the AS network had more positive links and intense connections than the attached biofilm networks, potentially facilitating the dissemination of ARGs. The differential distribution of ARGs among the three types of samples was significantly correlated with the microbial co-occurrence network topological properties. Bray–Curtis distance and network analysis suggest that microbial community is more robust in attached biofilms than in suspended AS. This work provides a more in-depth understanding of ARGs and microbial community distributions in wastewater biofilms.

Biogeography and Assembly of Microbial Communities in Wastewater Treatment Plants in China.

Scientific understanding of microbial biogeography and assembly is lacking for activated sludge microbial communities, even though the diversity of microbial communities in wastewater treatment plants (WWTPs) is thought to have a direct influence on system performance. Here, utilizing large-scale 16S rRNA gene data generated from 211 activated sludge samples collected from 15 cities across China, we show activated sludge microbes, whose growth and metabolism can be regulated followed with the metabolic theory of ecology with an apparent Ea value (apparent activation energy) of 0.08 eV. WWTPs at a lower latitude tend to harbor a more diverse array of microorganisms. In agreement with the general understanding, the activated sludge microbial assembly was mainly driven by deterministic processes and the mean annual temperature was identified as the most important factor affecting the microbial community structure. The treatment process types with similar microbial growth types and functions had a distinct impact on the activated sludge microbial community structure only when WWTPs were located near each other and received similar influent. Overall, these findings provide us with a deeper understanding of activated sludge microbial communities from an ecological perspective. Moreover, these findings suggest that, for a given set of performance characteristics (e.g., combined nitrification, denitrification, and phosphorus removal), it may be difficult to employ common engineering levers to control additional aspects of community structure due to the influence of natural environmental factors.

Comparison of Small Molecule Biotransformation Half-Lives between Activated Sludge and Soil: Opportunities for Read-Across?

Compartment-specific degradation half-lives are essential pieces of information in the regulatory risk assessment of synthetic chemicals. However, their measurement according to regulatory testing guidelines is laborious and costly. Despite the obvious ecological and economic benefits of knowing environmental degradability as early as possible, its consideration in the early phases of rational chemical design is therefore challenging. Here, we explore the possibility to use half-lives determined in highly time- and work-efficient biotransformation experiments with activated sludge and mixtures of chemicals to predict soil half-lives from regulatory simulation studies. We experimentally determined half-lives for 52 structurally diverse agrochemical active ingredients in batch reactors with three concentrations of the same activated sludge. We then developed bi- and multivariate models for predicting half-lives in soil by regressing the experimentally determined half-lives in activated sludge against average soil half-lives of the same chemicals extracted from regulatory data. The models differed in how we accounted for sorption-related bioavailability differences in soil and activated sludge. The best-performing models exhibited good coefficients of determination (R2 of around 0.8) and low average errors (<factor of 3 in half-life predictions) and were robust in cross-validation. From a practical perspective, these results suggest that it may indeed be possible to read across from half-lives determined in highly efficient biotransformation experiments in activated sludge to soil half-lives, which are obtained from much more work- and resource-intense regulatory studies, and that these predictions are clearly superior to predictions based on the output of BIOWIN, a publicly available quantitative structure-biodegradation relationship (QSBR) model. From a theoretical perspective, these results suggest that soil and activated sludge microbial communities, although certainly different in terms of taxonomic composition, may be functionally similar with respect to the enzymatic transformation of environmentally relevant concentrations of a diverse range of chemical compounds.

Activated Sludge Microbial Community and Treatment Performance of Wastewater Treatment Plants in Industrial and Municipal Zones.

Controlling wastewater pollution from centralized industrial zones is important for reducing overall water pollution. Microbial community structure and diversity can adversely affect wastewater treatment plant (WWTP) performance and stability. Therefore, we studied microbial structure, diversity, and metabolic functions in WWTPs that treat industrial or municipal wastewater. Sludge microbial community diversity and richness were the lowest for the industrial WWTPs, indicating that industrial influents inhibited bacterial growth. The sludge of industrial WWTP had low Nitrospira populations, indicating that influent composition affected nitrification and denitrification. The sludge of industrial WWTPs had high metabolic functions associated with xenobiotic and amino acid metabolism. Furthermore, bacterial richness was positively correlated with conventional pollutants (e.g., carbon, nitrogen, and phosphorus), but negatively correlated with total dissolved solids. This study was expected to provide a more comprehensive understanding of activated sludge microbial communities in full-scale industrial and municipal WWTPs.

Effects of quorum quenching on temporal succession of activated sludge microbial community in a membrane bioreactor.

Quorum quenching (QQ) is an attractive strategy for mitigating biofouling in membrane bioreactors (MBRs). However, the effects of QQ on the activated sludge (AS) process have not been adequately evaluated. This study investigated the long-term effects of QQ on a laboratory-scale anoxic-oxic MBR, focusing on AS performance and microbial community. Anoxic-oxic MBRs with and without QQ were operated for 91days. QQ did not affect COD and TN removal efficiencies over the experimental period, during which its activity remained >90%. QQ reduced floc size by approximately 8% but had no effect on biomass concentration. AS microbial communities were regularly analysed using massively parallel sequencing. AS bacterial communities were temporally dynamic irrespective of QQ presence, for example, a temporal increase in bacterial diversity and a temporal decay of community similarity. QQ counteracted the temporal change in diversity and the temporal distance-community decay. Community comparison revealed that QQ changed the successional trajectory of the AS community at a late period, because it decelerated temporal changes of specific members, such as Thiothrix and Sphingomonadaceae*. Correlation networks revealed that QQ increased network clustering, complexity and density. The combined results suggest that the tighter microbial association by QQ increased the community resistance. QQ can enhance the diversity and stability of the AS community in MBR by counteracting the innate temporal change in community structure. Our findings are useful for the further advancement of QQ-based strategies in engineered microbial environments.

Comparison of performance, microbial kinetics and microbial community between intelligent and conventional aeration-controlled Anoxic/Oxic systems

This work reports a comparison study between an intelligent aeration-controlled Anoxic/Oxic system and a conventional continuing aeration Anoxic/Oxic system for the removal of major containment, which includes the performance of major containments treatment, microbial kinetics and microbial communities by high-throughput sequencing tool. The results showed that at the end of low dissolved oxygen start-up period, there was few differences on the performance between the intelligent aeration-controlled Anoxic/Oxic system, with a 60% aeration saving, and conventional continuing aeration Anoxic/Oxic system. Moreover, the differences of activated sludge microbial community were significant. The kinetics study shows that the microbial kinetics for the intelligent aeration-controlled Anoxic/Oxic system was faster than for the comparison system, especially the yield coefficient for autotrophic biomass was much higher. The high-throughput sequencing tool also have provided a reasonable comparison of microbial community in aeration tank and indicated that the low dissolved oxygen demand species were more abundant. Meanwhile, the functional relative abundance had little statistical differences. However, the more abundance of low dissolved oxygen demand species gradually represented function of the less abundance species in the end of the start-up process. This study can provide a helpful information in the intelligent aeration-controlled Anoxic/Oxic system optimization to further removal the specific contaminates in the domestic wastewater treatment.

Long-term exposure of activated sludge in chemostats leads to changes in microbial communities composition and enhanced biodegradation of 4-chloroaniline and N-methylpiperazine.

Exposure history and adaptation of the inoculum to chemicals have been shown to influence the outcome of ready biodegradability tests. However, there is a lack of information about the mechanisms involved in microbial adaptation and the implication thereof for the tests. In the present study, we investigated the impact of a long-term exposure to N-methylpiperazine (NMP) and 4-chloroaniline (4CA) of an activated sludge microbial community using chemostat systems. The objective was to characterize the influence of adaptation to the chemicals on an enhanced biodegradation testing, following the OECD 310 guideline. Cultures were used to inoculate the enhanced biodegradability tests, in batch, before and after exposure to each chemical independently in chemostat culture. Composition and diversity of the microbial communities were characterised by 16s rRNA gene amplicon sequencing. Using freshly sampled activated sludge, NMP was not degraded within the 28 d frame of the test while 4CA was completely eliminated. However, after one month of exposure, the community exposed to NMP was adapted and could completely degrade it. This result was in complete contrast with that from the culture exposed for 3 months to 4CA. Long term incubation in the chemostat system led to a progressive loss of the initial biodegradation capacity of the community, as a consequence of the loss of key degrading microorganisms. This study highlights the potential of chemostat systems to induce adaptation to a specific chemical, ultimately resulting in its biodegradation. At the same time, one should be critical of these observations as the dynamics of a microbial community are difficult to maintain in chemostat, as the loss of 4CA biodegradation capacity demonstrates.

Evaluation of the Abundance of Fungi in Wastewater Treatment Plants Using Quantitative PCR (qPCR).

Assessment of the abundance of fungi in environmental samples by quantitative PCR (qPCR) of community DNA is often a difficult task due to biases introduced during PCR amplification, resulting from the differences associated with length polymorphism and the varying number of copies of the rRNA operon among fungal species, the lack of specificity of the primers targeting the different regions of the rRNA operon, or their insufficient coverage of the fungal lineages. To overcome those limitations, it is crucial to test and select the specific primers sets which provide the more accurate approximation to the quantification of the targeted fungal populations in a given set of samples. Fungi are a significant fraction of the microbiota in wastewater treatment plants (WWTPs), but the activated sludge microbial communities comprise many other eukaryotic microorganisms whose molecular markers are often coamplified by primers initially designed as fungal-specific. Here, the use of the FungiQuant primer set is recommended for the quantification of fungal molecular markers (18S rRNA genes) by qPCR in activated sludge samples and the full protocol is described.

Stress-responses of activated sludge and anaerobic sulfate-reducing bacteria sludge under long-term ciprofloxacin exposure

The activated sludge (AS) and sulfate-reducing bacteria (SRB) sludge systems were continuously operated for 200 days in laboratory to investigate the stress-responses of these two sludge systems under ciprofloxacin (CIP) exposure. It was found that CIP was effectively removed by SRB sludge via adsorption and biodegradation, but little biodegradation in AS system. The CIP biodegradation by SRB sludge made the SRB sludge system more sustainable and tolerant to long-term CIP exposure than AS system with significant (p < 0.05) CIP desorption and decrease of CIP removal. CIP shaped the microbial communities in AS and SRB sludge, and significantly (p < 0.05) inhibited the family Nitrosomonadaceae (ammonia-oxidizing bacteria (AOB)) and genus Nitrospira (nitrite-oxidizing bacteria (NOB)/complete ammonia oxidizer(comammox)) and the nitrogen removal in AS system. Moreover, CIP posed the increase of genus Zoogloea-like organisms and the non-filamentous bulking of AS, e.g. 313 ± 12 mL/g of sludge volume index (SVI) at phase V (influent CIP = 5000 μg/L). The genus Desulfobacter was enriched in SRB sludge system under long-term CIP exposure, and stimulated chemical oxygen demand (COD) removal and sulfate reduction. The increase of genera Zoogloea, Acinetobacter and Flavobacterium in AS, and Zoogloea and Acinetobacter in SRB sludge systems under CIP exposure promoted extracellular polymeric substances (EPS) production and CIP adsorption for self-protection of microbes against CIP toxicity. The functional groups of N–H, O–H, C–O–C and C=O in EPS of AS and SRB sludge provided adsorption sites for CIP and impeded CIP impact on microbial cells. The findings of this study provide an insight into the stress-responses of AS and SRB sludge under long-term CIP exposure, and exhibit the great potential of treating CIP-laden wastewater by SRB sludge system.

Состав микробного сообщества активного ила в процессах совместной биологической и реагентной очистки сточных вод

Состав микробного сообщества активного ила в процессах совместной биологической и реагентной очистки сточных вод

Removal of Chloroxylenol Disinfectant by an Activated Sludge Microbial Community

Chloroxylenol (CHL) is an antimicrobial ingredient that is frequently used in antiseptics/disinfectants for skin (e.g. hand soap) and non-living surfaces. CHL is an alternative to triclosan and triclocarban, the use of which has recently been banned in some countries. Accordingly, the more widespread use of CHL may significantly increase its occurrence and level in aquatic environments in the near future, eventually resulting in potential ecological risks. Wastewater treatment plants (WWTPs) may be a point source of CHL in natural environments due to extensive discharge through urban waste stream disposal. While the satisfactory removal of CHL in WWTPs is critical for maintaining healthy aquatic ecosystems, the extent of CHL removal and whether CHL causes system upset/failure in WWTPs currently remain unknown. In the present study, we conducted bioreactor operation and batch experiments to investigate the fate and effects of CHL and elucidate the mechanisms underlying degradation at various levels from environmentally relevant to high levels (0.5–5 mg L−1). Bioreactors partially removed CHL (44–87%) via a largely biological route. Microbial association networks constructed using 16S rRNA gene sequencing data revealed selective enrichment and a correlation between Sphingobium and CHL, implying its involvement in the biological breakdown of CHL through dehalogenation and ring hydroxylation pathways. The present results provide insights into the behavior and effects of CHL in activated sludge communities and important information for the sustainable management of CHL that may be an emerging issue in the urban water cycle.

Cometabolic biotransformation and impacts of the anti-inflammatory drug diclofenac on activated sludge microbial communities

This study evaluated the removal of diclofenac (DCF) in activated sludge and its long-term exposure effects on the function and structure of the microbial community. Activated sludge could remove <50% of 50 μg/L DCF. The removal decreased significantly to below 15% when DCF concentrations increased to 500 and 5000 μg/L. Quantitative assessment of the fate of DCF showed that its main removal routes were biodegradation (21%) and adsorption (7%), with other abiotic removals being insignificant (<5%). The biodegradation occurred through cometabolic mechanisms. DCF exposure in the range of 50–5000 μg/L did not disrupt the major functions of the activated sludge ecosystem (e.g. biomass yield and heterotrophic activity) over two months of DCF exposure. Consistently, 16S rRNA gene-based community analysis revealed that the overall community diversity (e.g. species richness and diversity) and structure of activated sludge underwent no significant alterations. The analysis did uncover a significant increase in several genera, Nitratireductor, Asticcacaulis, and Pseudacidovorax, which gained competitive advantages under DCF exposure. The enrichment of Nitratireductor, Asticcacaulis, and Pseudacidovorax genus might contribute to DCF biodegradation and emerge as a potential microbial niche for the removal of DCF.

Activated sludge bacterial communities of typical wastewater treatment plants: distinct genera identification and metabolic potential differential analysis

To investigate the differences in activated sludge microbial communities of different wastewater treatment plants (WWTPs) and understand their metabolic potentials, we sampled sludge from every biological treatment unit of 5 full-scale waste water treatment systems in 3 typical Chinese municipal WWTPs. The microbial communities and overall metabolic patterns were not only affected by influent characteristics but also varied between different biological treatment units. Distinct genera in different wastewater treatment systems were identified. The important microorganisms in domestic sewage treatment systems were unclassified SHA-20, Caldilinea, Dechloromonas, and unclassified genera from Rhodospirilaceae and Caldilineaceae. The important microorganisms in dyeing wastewater treatment systems were Nitrospira, Sphingobacteriales, Thiobacillus, Sinobacteraceae and Comamonadaceae. Compared with the obvious differences in microbial community composition, the metabolic potential showed no significant differences.

Activated sludge microbial communities of a chemical plant wastewater treatment facility with high-strength bromide ions and aromatic substances.

The prokaryotic and eukaryotic microbial communities of activated sludge in a chemical plant wastewater treatment facility, processing relatively oligotrophic wastewater containing aromatic compounds and high-strength bromide ions, were characterized by high-throughput sequencing of rRNA genes based on DNA and RNA extracts. The microbial community structure was distinct from those previously reported from domestic wastewater treatment plants. Several abundant OTUs in the RNA-based prokaryotic community were related to aromatic compound-degrading bacteria, which most likely contributed to the removal of recalcitrant chemicals from the wastewater. Furthermore, both prokaryotic and eukaryotic predators were highly abundant. These might promote stabilization of the microbial food chain and affect biomass in the activated sludge, maintaining the waste-removal function of the microbial community.

Removal of anti-inflammatory/analgesic pharmaceuticals from urban wastewater in a pilot-scale A2O system: Linking performance and microbial population dynamics to operating variables

In this study, the removal rates of eight anti-inflammatory and/or analgesic pharmaceuticals, AIAPs (acetaminophen, ibuprofen, naproxen, ketoprofen, diclofenac, codeine, indomethacin and propyphenazone) were assessed in a pilot-scale A2O system (including anaerobic/anoxic/aerobic zones), long term operated during two experimental phases using different sets of environmental conditions and operating parameters. qPCR was used to quantify the absolute abundances of total Bacteria, total Archaea, mycolic-acid containing filamentous Actinobacteria (Mycolata) and Fungi within the activated sludge microbial community developed in the system. Multivariate analyses and Spearman correlation coefficients were used in search of significant links among the removal rates of the AIAPs, the abundances of the targeted microbial groups in the activated sludge, and the changes of environmental/operating variables in the A2O system. Improved removal efficiencies of several of the AIAPs analyzed (acetaminophen, ibuprofen, naproxen, ketoprofen) were correlated to higher organic load in the influent water, higher concentration of mixed liquor suspended solids (MLSS), lower temperature and lower food-to-microorganisms ratio (F/M). Removal efficiencies of several pharmaceuticals correlated with increased abundances of Mycolata in the A2O system, pointing at this group of bacteria as candidate key players for AIAPs removal in activated sludge.

Disentangling the Drivers of Diversity and Distribution of Fungal Community Composition in Wastewater Treatment Plants Across Spatial Scales.

Activated sludge microbial community composition is a key bio-indicator of the sustainability of wastewater treatment systems. Therefore, a thorough understanding of the activated sludge microbial community dynamics is critical for environmental engineers to effectively manage the wastewater treatment plants (WWTPs). However, fungal communities associated with activated sludge have been poorly elucidated. Here, the activated sludge fungal community in 18 geographically distributed WWTPs was determined by using Illumina sequencing. The results showed that differences in activated sludge fungal community composition were observed among all WWTPs and also between oxidation ditch and anaerobic-anoxic-aerobic (A/A/O) systems. Ascomycota was the largest phyla, followed by Basidiomycota in all samples. Sporidiobolales and Pezizales were the most abundant order in oxidation ditch and A/A/O systems, respectively. The network analysis indicated cooperative and co-occurrence interactions between fungal taxa in order to accomplish the wastewater treatment process. Hygrocybe sp., Sporobolomyces sp., Rhodotorula sp., Stemphylium sp., Parascedosporium sp., and Cylindrocarpon sp., were found to have statistically significant interactions. Redundancy analysis revealed that temperature, total phosphorus, pH, and ammonia nitrogen were significantly affected the fungal community. This study sheds light on providing the ecological characteristics of activated sludge fungal communities and useful guidance for improving wastewater treatment performance efficiency.

Estimating the Transfer Range of Plasmids Encoding Antimicrobial Resistance in a Wastewater Treatment Plant Microbial Community

Wastewater treatment plants (WWTPs) have been suggested as reservoirs and sources of antibiotic resistance genes (ARGs) in the environment. In a WWTP ecosystem, human enteric and environmental bacteria are mixed and exposed to pharmaceutical residues, potentially favoring genetic exchange and thus ARG transmission. However, the contribution of microbial communities in WWTPs to ARG dissemination remains poorly understood. Here, we examined for the first time plasmid permissiveness of an activated sludge microbial community by utilizing an established fluorescent bioreporter system. The activated sludge microbial community was challenged in standardized filter matings with one of three multidrug resistance plasmids (pKJK5, pB10, and RP4) harbored by Escherichia coli or Pseudomonas putida. Different donor–plasmid combinations had distinct transfer frequencies, ranging from 3 to 50 conjugation events per 100000 cells of the WWTP microbial community. In addition, transfer was observed to a broad phylogenetic r...

Structure and function of the microbial consortia of activated sludge in typical municipal wastewater treatment plants in winter

To better understand the relationship between the environmental variables and microbial communities of activated sludge, we took winter samples from different biological treatment units (anaerobic, oxic, etc) from the WWTP’s of a number of Chinese cities. Differences in influent organic components and activated sludge microbial communities were identified by gas chromatography-mass spectrometry and high-throughput sequencing technology, respectively. Liquid nitrogen grinding pretreatment of samples was found to aid in the obtaining of a more bio-diversified sample. Influent type and dissolved oxygen concentration influenced the activated sludge microbial community structure. Nitrospira, Caldilineaceae and Anaerolineaceae were highly related to domestic wastewater treatment systems, whereas Thauera was the most abundant putative refractory aromatic hydrocarbon decomposer found in industrial wastewater treatment systems. Within the influent composition, we speculate that Thauera, Macellibacteroides and Desulfomicrobium are the key functional genera of the anaerobic environment of the textile dyeing industry wastewater treatment systems, whilst Thauera and Thiobacillus are key functional microbes in fine chemical wastewater treatment systems.

Elucidating the impact of microbial community biodiversity on pharmaceutical biotransformation during wastewater treatment.

SummaryIn addition to removing organics and other nutrients, the microorganisms in wastewater treatment plants (WWTPs) biotransform many pharmaceuticals present in wastewater. The objective of this study was to examine the relationship between pharmaceutical biotransformation and biodiversity in WWTP bioreactor microbial communities and identify taxa and functional genes that were strongly associated with biotransformation. Dilution‐to‐extinction of an activated sludge microbial community was performed to establish cultures with a gradient of microbial biodiversity. Batch experiments were performed using the dilution cultures to determine biotransformation extents of several environmentally relevant pharmaceuticals. With this approach, because the communities were all established from the same original community, and using sequencing of the 16S rRNA and metatranscriptome, we identified candidate taxa and genes whose activity and transcript abundances associated with the extent of individual pharmaceutical biotransformation and were lost across the biodiversity gradient. Metabolic genes such as dehydrogenases, amidases and monooxygenases were significantly associated with pharmaceutical biotransformation, and five genera were identified whose activity significantly associated with pharmaceutical biotransformation. Understanding how biotransformation relates to biodiversity will inform the design of biological WWTPs for enhanced removal of chemicals that negatively impact environmental health.

Identification of metabolites produced during the complete biodegradation of 1-butyl-3-methylimidazolium chloride by an enriched activated sludge microbial community

Ionic liquids (ILs) are highly polar solvents with unique physicochemical properties that make them promising green alternatives to volatile organic solvents. Since ILs can be toxic to organisms, the development of methods to degrade ILs into harmless molecules prior to disposal is critical to enhancing their green properties. In this study, metabolites generated during the biodegradation of 1-butyl-3-methylimidazolium chloride (BMIMCl) by an enriched, activated sludge microbial community were investigated. Biodegradation of BMIM and the metabolic products released into the growth media were examined using 1H-NMR spectroscopy and mass spectrometry. To the best of our knowledge, this is the first reported complete primary catabolism of the biodegradation-resistant BMIMCl ionic liquid. The bacterial community responsible for degradation was analyzed using a 16S-rRNA amplicon approach. Although the community was diverse, Bacteroidetes was the predominant phylum. The study provides a greater insight into imidazolium-based IL biodegradability and a means to proactively prevent the ecotoxicity of the BMIM cation and its metabolites, by complete primary biodegradation of the cation and removal of most resulting metabolites, prior to release into aquatic waste streams.