Micropollutant transformation and taxonomic composition in hybrid MBBR - A comparison of carrier-attached biofilm and suspended sludge.

The suspended sludge and carrier-attached biofilms of three different hybrid moving bed biofilm reactor (MBBR) systems were investigated with respect to their transformation potential for a broad range of micropollutants (MPs) as well as their microbial community composition. For this purpose, laboratory-scale batch experiments were conducted with the separated suspended sludge and the carrier-attached biofilm of every system in triplicate. For all batches the removal of 31 MPs as well as the composition of the microbial community were analyzed. The carrier-attached biofilms from two hybrid MBBR systems showed a significant higher overall transformation potential in comparison to the respective suspended sludge. Especially for the MPs trimethoprim, diclofenac, mecoprop, climbazole and the human metabolite 10,11-dihydro-10-hydroxycarbamazepine consistently higher pseudo-first-order transformation rates could be observed in all three systems. The analysis of the taxonomic composition revealed taxa showing higher relative abundances in the carrier-attached biofilms (e. g. Nitrospirae and Chloroflexi) and in the suspended biomasses (e. g. Bacteroidetes and Betaproteobacteria). Correlations of the biodiversity indices and the MP biotransformation rates resulted in significant positive associations for 11 compounds in suspended sludge, but mostly negative associations for the carrier-attached biofilms. The distinct differences in MP removal between suspended sludge and carrier-attached biofilm of the three different MBBR systems were also reflected by a statistically significant link between the occurrence of specific bacterial taxa (Acidibacter, Nitrospira and Rhizomicrobium) and MP transformation rates of certain MPs. Even though the identified correlations might not necessarily be of causal nature, some of the identified taxa might serve as suitable indicators for the transformation potential of suspended sludge or carrier-attached biofilms.

Biotransformation of organic micropollutants by anaerobic sludge enzymes.

Biotransformation of organic micropollutants (OMPs) in wastewater treatment plants ultimately depends on the enzymatic activities developed in each biological process. However, few research efforts have been made to clarify and identify the role of enzymes on the removal of OMPs, which is an essential knowledge to determine the biotransformation potential of treatment technologies. Therefore, the purpose of the present study was to investigate the enzymatic transformation of 35 OMPs under anaerobic conditions, which have been even less studied than aerobic systems. Initially, 13 OMPs were identified to be significantly biotransformed (>20%) by anaerobic sludge obtained from a full-scale anaerobic digester, predestining them as potential targets of anaerobic enzymes. Native enzymes were extracted from this anaerobic sludge to perform transformation assays with the OMPs. In addition, the effect of detergents to recover membrane enzymes, as well as the effects of cofactors and inhibitors to promote and suppress specific enzymatic activities were evaluated. In total, it was possible to recover enzymatic activities towards 10 out of these 13 target OMPs (acetyl-sulfamethoxazole and its transformation product sulfamethoxazole, acetaminophen, atenolol, clarithromycin, citalopram, climbazole, erythromycin, and terbutryn, venlafaxine) as well as towards 8 non-target OMPs (diclofenac, iopamidol, acyclovir, acesulfame, and 4 different hydroxylated metabolites of carbamazepine). Some enzymatic activities likely involved in the anaerobic biotransformation of these OMPs were identified. Thereby, this study is a starting point to unravel the still enigmatic biotransformation of OMPs in wastewater treatment systems.

Insights into the variability of microbial community composition and micropollutant degradation in diverse biological wastewater treatment systems.

The biological potential of conventional wastewater treatment plants to remove micropollutants mainly depends on process conditions and the predominant microbial community. To explore this dependence and to connect the occurrence of genera with operating conditions, five pilot-scale reactors with different process conditions were combined into two reactor cascades and fed with the effluent of the primary clarifier of a municipal WWTP. All reactors and the WWTP were analyzed for the removal of 33 micropollutants by LC-MS/MS and the presence of the microbial community using 16S rRNA gene sequencing. The overall removal of the micropollutants was slightly improved (ca. 20%) by the reactor cascades in comparison to the WWTP while certain compounds such as diatrizoate, venlafaxine or diclofenac showed an enhanced removal (ca. 70% in one or both cascades). To explore the diverse bacteria in more detail, the general community was divided into a core and a specialized community. Despite their profoundly different operating parameters (especially redox conditions), the different treatments share a core community consisted of 143 genera (9% of the overall community). Furthermore, the alpha- and beta-biodiversity as well as the occurrence of several genera belonging to the specialized microbial community could be linked to the prevalent process conditions of the individual treatments. Members of the specialized community also correlated with the removal of certain groups of micropollutants. Hence, the comparison of the specialized community with micropollutant removal and operating conditions via correlation analysis is a valuable tool for an extended evaluation of prevalent process conditions. Based on an extended data set this approach could also be used to identify organisms as indicators for operating conditions which are beneficial for an improved removal of specific micropollutants.

Anaerobic biodegradation of (emerging) organic contaminants in the aquatic environment.

Although strictly anaerobic conditions prevail in several environmental compartments, up to now, biodegradation studies with emerging organic contaminants (EOCs), such as pharmaceuticals and personal care products, have mainly focused on aerobic conditions. One of the reasons probably is the assumption that the aerobic degradation is more energetically favorable than degradation under strictly anaerobic conditions. Certain aerobically recalcitrant contaminants, however, are biodegraded under strictly anaerobic conditions and little is known about the organisms and enzymatic processes involved in their degradation. This review provides a comprehensive survey of characteristic anaerobic biotransformation reactions for a variety of well-studied, structurally rather simple contaminants (SMOCs) bearing one or a few different functional groups/structural moieties. Furthermore it summarizes anaerobic degradation studies of more complex contaminants with several functional groups (CMCs), in soil, sediment and wastewater treatment. While strictly anaerobic conditions are able to promote the transformation of several aerobically persistent contaminants, the variety of observed reactions is limited, with reductive dehalogenations and the cleavage of ether bonds being the most prevalent. Thus, it becomes clear that the transferability of degradation mechanisms deduced from culture studies of SMOCs to predict the degradation of CMCs, such as EOCs, in environmental matrices is hampered due the more complex chemical structure bearing different functional groups, different environmental conditions (e.g. matrix, redox, pH), the microbial community (e.g. adaptation, competition) and the low concentrations typical for EOCs.

Micropollutant degradation via extracted native enzymes from activated sludge.

A procedure was developed to assess the biodegradation of micropollutants in cell-free lysates produced from activated sludge of a municipal wastewater treatment plant (WWTP). This proof-of-principle provides the basis for further investigations of micropollutant biodegradation via native enzymes in a solution of reduced complexity, facilitating downstream protein analysis. Differently produced lysates, containing a variety of native enzymes, showed significant enzymatic activities of acid phosphatase, ß-galactosidase and ß-glucuronidase in conventional colorimetric enzyme assays, whereas heat-deactivated controls did not. To determine the enzymatic activity towards micropollutants, 20 compounds were spiked to the cell-free lysates under aerobic conditions and were monitored via LC-ESI-MS/MS. The micropollutants were selected to span a wide range of different biodegradabilities in conventional activated sludge treatment via distinct primary degradation reactions. Of the 20 spiked micropollutants, 18 could be degraded by intact sludge under assay conditions, while six showed reproducible degradation in the lysates compared to the heat-deactivated negative controls: acetaminophen, N-acetyl-sulfamethoxazole (acetyl-SMX), atenolol, bezafibrate, erythromycin and 10,11-dihydro-10-hydroxycarbamazepine (10-OH-CBZ). The primary biotransformation of the first four compounds can be attributed to amide hydrolysis. However, the observed biotransformations in the lysates were differently influenced by experimental parameters such as sludge pre-treatment and the addition of ammonium sulfate or peptidase inhibitors, suggesting that different hydrolase enzymes were involved in the primary degradation, among them possibly peptidases. Furthermore, the transformation of 10-OH-CBZ to 9-CA-ADIN was caused by a biologically-mediated oxidation, which indicates that in addition to hydrolases further enzyme classes (probably oxidoreductases) are present in the native lysates. Although the full variety of indigenous enzymatic activity of the activated sludge source material could not be restored, experimental modifications, e.g. different lysate filtration, significantly enhanced specific enzyme activities (e.g. >96% removal of the antibiotic erythromycin). Therefore, the approach presented in this study provides the experimental basis for a further elucidation of the enzymatic processes underlying wastewater treatment on the level of native proteins.