Search

Woodchip bioreactors are being successfully applied to remove nitrate from commercial land-based recirculating aquaculture system (RAS) effluents. In order to understand and optimize the overall function of these bioreactors, knowledge on the microbial communities, especially on the microbes with potential for production or mitigation of harmful substances (e.g. hydrogen sulfide; H 2 S) is needed. In this study, we quantified and characterized bacterial and fungal communities, including potential H 2 S producers and consumers, using qPCR and high throughput sequencing of 16S rRNA gene. We took water samples from bioreactors and their inlet and outlet, and sampled biofilms growing on woodchips and on the outlet of the three full-scale woodchip bioreactors treating effluents of three individual RAS. We found that bioreactors hosted a high biomass of both bacteria and fungi. Although the composition of microbial communities of the inlet varied between the bioreactors, the conditions in the bioreactors selected for the same core microbial taxa. The H 2 S producing sulfate reducing bacteria (SRB) were mainly found in the nitrate-limited outlets of the bioreactors, the main groups being deltaproteobacterial Desulfobulbus and Desulfovibrio. The abundance of H 2 S consuming sulfate oxidizing bacteria (SOB) was 5-10 times higher than that of SRB, and SOB communities were dominated by Arcobacter and other genera from phylum Epsilonbacteraeota, which are also capable of autotrophic denitrification. Indeed, the relative abundance of potential autotrophic denitrifiers of all denitrifier sequences was even 54% in outlet water samples and 56% in the outlet biofilm samples. Altogether, our results show that the highly abundant bacterial and fungal communities in woodchip bioreactors are shaped through the conditions prevailing within the bioreactor, indicating that the bioreactors with similar design and operational settings should provide similar function even when conditions in the preceding RAS would differ. Furthermore, autotrophic denitrifiers can have a significant role in woodchip biofilters, consuming potentially produced H 2 S and removing nitrate, lengthening the operational age and thus further improving the overall environmental benefit of these bioreactors., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Woodchip bioreactors are viable low-cost nitrate (NO 3 - ) removal applications for treating agricultural and aquaculture discharges. The active microbial biofilms growing on woodchips are conducting nitrogen (N) removal, reducing NO 3 - O) production through either incomplete denitrification or dissimilatory NO 3 - removal rates, and the abundance and community composition of microbes involved in N transformation processes in the three different full-scale woodchip bioreactors treating recirculating aquaculture system (RAS) effluents. We confirmed denitrification producing di‑nitrogen gas (N 2 O) production through either incomplete denitrification or dissimilatory NO 3 - reduction to ammonium (DNRA). Here, we combined stable isotope approach, amplicon sequencing, and captured metagenomics for studying the potential NO 3 - removal rates, and the abundance and community composition of microbes involved in N transformation processes in the three different full-scale woodchip bioreactors treating recirculating aquaculture system (RAS) effluents. We confirmed denitrification producing di‑nitrogen gas (N 2 removal rates were linked to the denitrifying community diversity. The same core proteobacterial groups were driving the denitrification, while Bacteroidetes dominated the DNRA carrying microbes in all the three bioreactors studied. Altogether, our results suggest that woodchip bioreactors have a high genetic potential for NO 3 - removal pathway, but found that 6% of NO 3 - could be released as N 2 O under high NO 3 - concentrations and low amounts of bioavailable C, whereas DNRA rates tend to increase with the C amount. The abundance of denitrifiers was equally high between the studied bioreactors, yet the potential NO 3 - removal rates were linked to the denitrifying community diversity. The same core proteobacterial groups were driving the denitrification, while Bacteroidetes dominated the DNRA carrying microbes in all the three bioreactors studied. Altogether, our results suggest that woodchip bioreactors have a high genetic potential for NO 3 - removal through a highly abundant and diverse denitrifying community, but that the rates and dynamics between the NO 3 - removal pathways depend on the other factors (e.g., bioreactor design, operating conditions, and the amount of bioavailable C in relation to the incoming NO 3 - concentrations)., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Molecular monitoring of bacterial communities can explain and predict the stability of bioprocesses in varying physicochemical conditions. To study methanol-fed denitrification biofilters of municipal wastewater treatment plants, bacterial communities of two full-scale biofilters were compared through fingerprinting and sequencing of the 16S rRNA genes. Additionally, 16S rRNA gene fingerprinting was used for 10-week temporal monitoring of the bacterial community in one of the biofilters. Combining the data with previous study results, the family Methylophilaceae and genus Hyphomicrobium were determined as suitable target groups for monitoring. An increase in the relative abundance of Hyphomicrobium-related biomarkers occurred simultaneously with increases in water flow, NO x load, and methanol addition, as well as a higher denitrification rate, although the dominating biomarkers linked to Methylophilaceae showed an opposite pattern. The results indicate that during increased loading, stability of the bioprocess is maintained by selection of more efficient denitrifier populations, and this progress can be analyzed using simple molecular fingerprinting.- load, and methanol addition, as well as a higher denitrification rate, although the dominating biomarkers linked to Methylophilaceae showed an opposite pattern. The results indicate that during increased loading, stability of the bioprocess is maintained by selection of more efficient denitrifier populations, and this progress can be analyzed using simple molecular fingerprinting.

Which bacterial taxonomic groups can be used in monitoring saline water methanol-utilizing denitrification and whether nitrate is transformed into N 2 in the process are unclear. Therefore, methylotrophic bacterial communities of two efficiently functioning (nitrate/nitrite reduction was 63-96 %) tropical and cool seawater reactors at a public aquarium were investigated with clone library analysis and 454 pyrosequencing of the 16S rRNA genes. Transformation of nitrate into N 2 was confirmed using 15 N labeling in incubation of carrier material from the tropical reactor. Combining the data with previous study results, Methylophaga and Hyphomicrobium were determined to be suitable target genera for monitoring the function of saline water methanol-fed denitrification systems. However, monitoring was not possible at the single species level. Interestingly, potential nitrate-reducing methylotrophs within Filomicrobium and closely related Fil I and Fil II clusters were detected in the reactors suggesting that they also contributed to methylotrophic denitrification in the saline environment.

Mid-summer N(2) profiles were analyzed from nine oxygen-stratified, humic-acid-rich lakes using a continuous flow isotope ratio mass spectrometer and a Gasbench II device. Sample preparation steps were performed under water to avoid air contamination. The instrument precision for the δ(15)N measurement was high (0.03‰), but for the whole sampling and analysis procedure the mean deviation between replicate samples was 0.13‰ for the δ(15)N measurements and 5.5% for the N(2) gas concentration analysis. The results show that the Gasbench peripheral was suitable for measurement of the (15)N natural abundance of dissolved nitrogen gas, with denitrification indicated by the oversaturation and slightly (<1‰) depleted δ(15)N values of the dissolved N(2) gas in the suboxic zones of some of the study lakes. Calculated values for the denitrified (excess) N(2) varied between -5.3 and 0.7‰. The denitrification potential was determined using the (15)N tracer method, with results showing nitrate-inducible denitrification and no signs of anaerobic ammonium oxidation (anammox)., (Copyright © 2011 John Wiley & Sons, Ltd.)

Despite high performance, municipal wastewater treatment plants (WWTPs) still discharge significant amounts of organic material and nitrogen and even microbes into the receiving water bodies, altering physico-chemical conditions and microbial functions. In this study, we examined how nitrified wastewater affects the microbiology of boreal lake sediments. Microbial community compositions were assessed with next generation sequencing of the 16S rRNA gene, and a more detailed view on nitrogen transformation processes was gained with qPCR targeting on functional genes (nirS, nirK, nosZI, nosZII, amoAarchaea, and amoAbacteria). In both of the two studied lake sites, the microbial community composition differed significantly between control point and wastewater discharge point, and a gradual shift toward natural community composition was seen downstream following the wastewater gradient. SourceTracker analysis predicted that ~2% of sediment microbes were of WWTP-origin on the study site where wastewater was freely mixed with the lake water, while when wastewater was specially discharged to the sediment surface, ~6% of microbes originated from WWTP, but the wastewater-influenced area was more limited. In nitrogen transformation processes, the ratio between nitrifying archaea (AOA) and bacteria (AOB) was affected by wastewater effluent, as the AOA abundance decreased from the control point (AOA:AOB 28:1 in Keuruu, 11:1 in Petäjävesi) to the wastewater-influenced sampling points, where AOB dominated (AOA:AOB 1:2-1:15 in Keuruu, 1:3-1:19 in Petäjävesi). The study showed that wastewater can affect sediment microbial community through importing nutrients and organic material and altering habitat characteristics, but also through bringing wastewater-originated microbes to the sediment, and may thus have significant impact on the freshwater biogeochemistry, especially in the nutrient-poor boreal ecosystems. 9 [ABSTRACT FROM AUTHOR]

Denitrification is an important microbial process in aquatic ecosystems that can reduce the effects of eutrophication. Here, quantification and pyrosequencing of nirS, nirK, and nosZ genes encoding for nitrite and nitrous oxide reductases was performed in sediment samples from four boreal lakes to determine the structure and seasonal stability of denitrifying microbial populations. Sediment quality and nitrate concentrations were linked to the quantity and diversity of denitrification genes, the abundance of denitrifying populations (nirS and nosZ genes) correlated with coupled nitrificationdenitrification (Dn), and the denitrification of the overlying water NO3- (Dw) correlated with the nirS/nirK ratio. The number of core nirS, nirK, and nosZ operational taxonomical units (OTUs) was low (6, 7, and 3, respectively), and most of these core OTUs were shared among the lakes. Dominant nirK sequences matched best with those of the order Rhizobiales, which was one of the main bacterial orders present in the sediment microbiomes, whereas the dominant nirS sequences were affiliated with the order Burkholderiales. Over half of the nosZ sequences belonged to a single OTU of the order Burkholderiales, but coupled nitrification-denitrification rate correlated with another dominant nosZ OTU assigned to the order Rhodospirillales. The study indicates that a few core proteobacterial clusters may drive denitrification in boreal lake sediments, as the same Alpha- and Betaproteobacteria denitrifier clusters were present in different lakes and seasons. [ABSTRACT FROM AUTHOR]

Abstract This study investigated the effect of salinity on microbial composition and denitrification capacity of woodchip bioreactors treating recirculating aquaculture system (RAS) effluents. Twelve laboratory-scale woodchip bioreactors were run in triplicates at 0, 15, 25, and 35 ppt salinities, and water chemistry was monitored every third day during the first 39 days of operation. Microbial communities of the woodchips bioreactors were analyzed at the start, after one week, and at the end of the trial. Woodchip bioreactors removed nitrate at all salinities tested. The highest NO 3 -N removal rate of 22.0 ± 6.9 g NO 3 -N/m3/d was obtained at 0 ppt, while 15.3 ± 4.9, 12.5 ± 5.4 and 11.8 ± 4.0 g NO 3 -N/m3/d were obtained at salinities of 15, 25 and 35 ppt, respectively. Nitrate removal rates thus decreased with salinity, being 54–69% lower than at 0 ppt. Leaching of total ammonia nitrogen (TAN) and orthophosphate (PO 4 -P) from woodchips was initially higher at saline treatments compared to 0 ppt, while initial leaching of BOD 5 appeared to be similar across all treatments. Production of alkalinity per g NO 3 -N removed was higher at 0 (3.6 ± 0.5 gCaCO 3 /gNO 3 -N) and 15 ppt (3.5 ± 0.8) than at the more saline treatments (25 ppt: 2.0 ± 0.9, 35 ppt: 1.12 ± 0.5 gCaCO 3 /gNO 3 -N), indicating that heterotrophic denitrification was the dominant nitrate removing process at 0 and 15 ppt, while autotrophic denitrification processes probably interfered with the alkalinity balance at 25 and 35 ppt. In the woodchip reactors, Gammaproteobacteria was the most abundant taxa. However, salinity shaped the woodchip microbiome, resulting in an increase in the abundance of sulfide oxidizing autotrophic denitrifiers, but decrease in the overall abundance of denitrifying microbes at higher salinities, which presumably explained the reduced nitrate removal rates at elevated salinities. This study demonstrates that woodchip bioreactors can be applied to remove nitrate from saline RAS effluents albeit at lower nitrate removal rates compared to freshwater installations. Highlights • Denitrifying woodchip bioreactors can be applied to remove nitrates from saline RAS effluents. • Increasing salinity decreased nitrate removal in denitrifying woodchip bioreactors. • The woodchip microbiome was affected by salinity. • Salinity increased the abundance of autotrophic denitrifiers but decreased the overall abundance of denitrifiers. [ABSTRACT FROM AUTHOR]

Sediment microbes have a great potential to transform reactive N to harmless N 2 , thus decreasing wastewater nitrogen load into aquatic ecosystems. Here, we examined if spatial allocation of the wastewater discharge by a specially constructed sediment diffuser pipe system enhanced the microbial nitrate reduction processes. Full-scale experiments were set on two Finnish lake sites, Keuruu and Petäjävesi, and effects on the nitrate removal processes were studied using the stable isotope pairing technique. All nitrate reduction rates followed nitrate concentrations, being highest at the wastewater-influenced sampling points. Complete denitrification with N 2 as an end-product was the main nitrate reduction process, indicating that the high nitrate and organic matter concentrations of wastewater did not promote nitrous oxide (N 2 O) production (truncated denitrification) or ammonification (dissimilatory nitrate reduction to ammonium; DNRA). Using 3D simulation, we demonstrated that the sediment diffusion method enhanced the contact time and amount of wastewater near the sediment surface especially in spring and in autumn, altering organic matter concentration and oxygen levels, and increasing the denitrification capacity of the sediment. We estimated that natural denitrification potentially removed 3–10% of discharged wastewater nitrate in the 33 ha study area of Keuruu, and the sediment diffusion method increased this areal denitrification capacity on average 45%. Overall, our results indicate that sediment diffusion method can supplement wastewater treatment plant (WWTP) nitrate removal without enhancing alternative harmful processes. [ABSTRACT FROM AUTHOR]

Aquaculture needs practical solutions for nutrient removal to achieve sustainable fish production. Passive denitrifying bioreactors may provide an ecological, low-cost and low-maintenance approach for wastewater nitrogen removal. However, innovative organic materials are needed to enhance nitrate removal from the low carbon effluents in intensive recirculating aquaculture systems (RAS). In this study, we tested three additional carbon sources, including biochar, dried Sphagnum sp. moss and industrial potato residues, to enhance the performance of woodchip bioreactors treating the low carbon RAS wastewater. We assessed nitrate (NO 3 −) removal and microbial community composition during a one-year in situ column test with real aquaculture wastewater. We found no significant differences in the NO 3 − removal rates between the woodchip-only bioreactor and bioreactors with a zone of biochar or Sphagnum sp. moss (maximum removal rate 31–33 g NO 3 −-N m−3 d−1), but potato residues increased NO 3 − removal rate to 38 g NO 3 −-N m−3 d−1, with stable annual reduction efficiency of 93%. The readily available carbon released from potato residues increased NO 3 −-N removal capacity of the bioreactor even at higher inflow concentrations (>52 mg L−1). The microbial community and its predicted functional potential in the potato residue bioreactor differed markedly from those of the other bioreactors. Adding potato residues to woodchip material enabled smaller bioreactor size to be used for NO 3 − removal. This study introduced industrial potato by-product as an alternative carbon source for the woodchip denitrification process, and the encouraging results may pave the way toward growth of blue bioeconomy using the RAS. • Woodchip bioreactors removed 31–38 g NO 3 −-N m−3 d−1 from intensive aquaculture effluent. • Additional potato residues to woodchip material increased 13% of nitrate removal rate. • The potato residue bioreactor hosted a distinctly different microbial community. [ABSTRACT FROM AUTHOR]