Retrofitting a full-scale multistage landfill leachate treatment plant by introducing coagulation/flocculation/sedimentation and ultrafiltration process steps.

Considering that landfilling still remains among the most commonly used methods for the confrontation of solid wastes, effective methods should be applied to treat the leachate generated, due to its recalcitrant nature. In this work, a full-scale system consisting of two SBRs operating in parallel (350 m3 each) and two activated carbon (AC) columns operating in series (3 m3 each) was retrofitted by introducing a coagulation/flocculation/sedimentation (C/F/S) unit of 7.8 m3 and an ultrafiltration (UF) membrane of 100 m2 to effectively treat landfill leachate. The raw leachate was characterized by high COD and NH4+-N concentration, i.e., 3095 ± 706 mg/L and 1054 ± 141 mg/L respectively, a BOD/COD ratio of 0.22, and high concentrations of certain heavy metals. Leachate processing in this retrofitted multistage treatment system resulted in total COD removal efficiency of 89.84%, with biological treatment, C/F, UF, and AC contributing 46.31%, 4.68%, 15.98%, and 22.87% to the overall organic content removal. The retrofitted scheme achieved an overall NH4+-N and TKN removal of 92.03% and 91.75% respectively, attributed mostly to the activity of an effective nitrifying community. Color number (CN) was reduced by 26.96%, 10.29%, 15.94%, and 5.39% after the activated sludge, the C/F, the UF, and the AC adsorption process respectively, corresponding to a 58.91% overall decrease. Regarding heavy metal removal, all elements examined, apart from Ni, i.e., effluent As, Cd, Co, Cr, Cu, Hg, Mg, Mn, and Pb, were below the legislative limits set by the national authorities for restricted or unrestricted irrigation. Lastly, total operating expenses (OPEX) were estimated as equal to 72,687 €/year or 6.64 €/m3.

Unraveling the important role of comammox Nitrospira to nitrification in the coastal aquaculture system.

Increasing nitrogen (N) input to coastal ecosystems poses a serious environmental threat. It is important to understand the responses and feedback of N removal microbial communities, particularly nitrifiers including the newly recognized complete ammonia-oxidizers (comammox), to improve aquaculture sustainability. In this study, we conducted a holistic evaluation of the functional communities responsible for nitrification by quantifying and sequencing the key functional genes of comammox Nitrospira-amoA, AOA-amoA, AOB-amoA and Nitrospira-nxrB in fish ponds with different fish feeding levels and evaluated the contribution of nitrifiers in the nitrification process through experiments of mixing pure cultures. We found that higher fish feeding dramatically increased N-related concentration, affecting the nitrifying communities. Compared to AOA and AOB, comammox Nitrospira and NOB were more sensitive to environmental changes. Unexpectedly, we detected an equivalent abundance of comammox Nitrospira and AOB and observed an increase in the proportion of clade A in comammox Nitrospira with the increase in fish feeding. Furthermore, a simplified network and shift of keystone species from NOB to comammox Nitrospira were observed in higher fish-feeding ponds. Random forest analysis suggested that the comammox Nitrospira community played a critical role in the nitrification of eutrophic aquaculture ponds (40-70 µM). Through the additional experiment of mixing nitrifying pure cultures, we found that comammox Nitrospira is the primary contributor to the nitrification process at 200 µM ammonium. These results advance our understanding of nitrifying communities and highlight the importance of comammox Nitrospira in driving nitrification in eutrophic aquaculture systems.

Biological treatment of N-methylpyrrolidone, cyclopentanone, and diethylene glycol monobutyl ether distilled residues and their effects on nitrogen removal in a full-scale wastewater treatment plant.

Manufacturing processes in semiconductor and photonics industries involve the use of a significant amount of organic solvents. Recycle and reuse of these solvents produce distillate residues and require treatment before being discharged. This study aimed to evaluate the performance of the biological treatment system in a full-scale wastewater treatment plant that treats wastewater containing distillate residues from the recycling of electronic chemicals. Batch experiments were conducted to investigate the optimal operational conditions for the full-scale wastewater treatment plant. To achieve good nitrogen removal efficiency with effluent ammonia and nitrate concentrations below 20 mg N/L and 50 mg N/L, respectively, it was suggested to control the ammonia concentration and pH of the influent below 500 mg N/L and 8.0, respectively. In addition, the biodegradability of N-methylpyrrolidone, diethylene glycol monobutyl ether, and cyclopentanone distillate residues from the electronic chemicals manufacturing process were evaluated under aerobic, anoxic, and anaerobic conditions. N-methylpyrrolidone and cyclopentanone distillate residues were suggested to be treated under anoxic condition. However, substrate inhibition occurred when using cyclopentanone distillate residue as a carbon source with chemical oxygen demand (COD) levels higher than 866 mg/L and nitrate levels higher than 415 mg N/L. Under aerobic condition, the COD from both N-methylpyrrolidone and cyclopentanone distillate residues could be easily degraded. Nevertheless, a negative effect on nitrification was observed, with a prolonged lag time for ammonia oxidation as the initial COD concentration increased. The specific ammonia oxidation rate and nitrate production rate decreased under high COD concentration contributed by N-methylpyrrolidone and cyclopentanone distillate residues. Furthermore, the biodegradability of diethylene glycol monobutyl ether distillate residue was found to be low under aerobic, anoxic, and anaerobic conditions. With respect to the abundance of nitrogen removal microorganisms in the wastewater treatment plant, results showed that Comammox may have an advantage over ammonia oxidizing bacteria under high pH conditions. In addition, Comammox may have higher resistance to environmental changes. Dominance of Comammox over ammonia oxidizing bacteria under high ammonia condition was first reported in this study.

High-throughput assays to identify archaea-targeting nitrification inhibitors.

Nitrification is a microbial process that converts ammonia (NH3) to nitrite (NO2 -) and then to nitrate (NO3 -). The first and rate-limiting step in nitrification is ammonia oxidation, which is conducted by both bacteria and archaea. In agriculture, it is important to control this process as high nitrification rates result in NO3 - leaching, reduced nitrogen (N) availability for the plants and environmental problems such as eutrophication and greenhouse gas emissions. Nitrification inhibitors can be used to block nitrification, and as such reduce N pollution and improve fertilizer use efficiency (FUE) in agriculture. Currently applied inhibitors target the bacteria, and do not block nitrification by ammonia-oxidizing archaea (AOA). While it was long believed that nitrification in agroecosystems was primarily driven by bacteria, recent research has unveiled potential significant contributions from ammonia-oxidizing archaea (AOA), especially when bacterial activity is inhibited. Hence, there is also a need for AOA-targeting nitrification inhibitors. However, to date, almost no AOA-targeting inhibitors are described. Furthermore, AOA are difficult to handle, hindering their use to test or identify possible AOA-targeting nitrification inhibitors. To address the need for AOA-targeting nitrification inhibitors, we developed two miniaturized nitrification inhibition assays using an AOA-enriched nitrifying community or the AOA Nitrosospaera viennensis. These assays enable high-throughput testing of candidate AOA inhibitors. We here present detailed guidelines on the protocols and illustrate their use with some examples. We believe that these assays can contribute to the discovery of future AOA-targeting nitrification inhibitors, which could complement the currently applied inhibitors to increase nitrification inhibition efficiency in the field and as such contribute to a more sustainable agriculture.

Fertilizing-induced changes in the nitrifying microbiota associated with soil nitrification and crop yield.

Ammonia oxidizing archaea (AOA) and bacteria (AOB), nitrite-oxidizing bacteria (NOB), and comammox Nitrospira (CMX) play pivotal roles in global nitrogen-cycling network. Despite its importance, the driving forces for niche specialization of these nitrifiers, as well as their relative contributions to nitrification and crop yield have not been fully understood. Here, we investigated the niche specialization and environmental prevalence of nitrifying communities, and their importance for the nitrification rate and crop yield across a gradient of nitrogen inputs in a two-decade old field experiment. The results of 15N-tracer and quantitative PCR revealed that AOB and NOB jointly determined the gross nitrification rates across mineral fertilizer treatments, whereas AOA and AOB contributed more than other nitrifiers to nitrification under with organic fertilizer amendments. Linear regression model revealed that crop yield could be linked with AOB and NOB under inorganic farming but closely associated with CMX under organic management. Amplicon sequencing of these functional genes further demonstrated that mineral and organic fertilizers have distinct influences on the ß-diversity and niche breadth of these nitrifying communities, indicating that fertilization triggered niche specialization of nitrifying guilds in agricultural soils. Notably, organic fertilization enhanced the network complexity of these nitrifiers by harboring keystone taxa. Random forest analysis provide robustly evidence for the hypothesis that abundance of functional genes contributed more than a- and ß-diversity of these nitrifiers for driving nitrification rates and crop yields. Collectively, these findings provide the empirical evidence for the environmental adaptation and niche specialization of nitrifying communities, and their contributions in nitrification and crop yield when confronted with long-term nitrogen inputs.

Niche differentiation among comammox (Nitrospira inopinata) and other metabolically distinct nitrifiers.

Due to global change, increasing nutrient input to ecosystems dramatically affects the nitrogen cycle, especially the nitrification process. Nitrifiers including ammonia-oxidizing archaea (AOAs), ammonia-oxidizing bacteria (AOBs), nitrite-oxidizing bacteria (NOBs), and recently discovered complete ammonia oxidizers (comammoxs) perform nitrification individually or in a community. However, much remains to be learned about their niche differentiation, coexistence, and interactions among those metabolically distinct nitrifiers. Here, we used synthetic microbial ecology approaches to construct synthetic nitrifying communities (SNCs) with different combinations of Nitrospira inopinata as comammox, Nitrososphaera gargensis as AOA, Nitrosomonas communis as AOB, and Nitrospira moscoviensis as NOB. Our results showed that niche differentiation and potential interactions among those metabolically distinct nitrifiers were determined by their kinetic characteristics. The dominant species shifted from N. inopinata to N. communis in the N4 community (with all four types of nitrifiers) as ammonium concentrations increased, which could be well explained by the kinetic difference in ammonia affinity, specific growth rate, and substrate tolerance of nitrifiers in the SNCs. In addition, a conceptual model was developed to infer niche differentiation and possible interactions among the four types of nitrifiers. This study advances our understanding of niche differentiation and provides new strategies to further study their interactions among the four types of nitrifiers.

Nitrite Oxidizer Activity and Community Are More Responsive Than Their Abundance to Ammonium-Based Fertilizer in an Agricultural Soil.

Autotrophic nitrification is mediated by ammonia oxidizing bacteria (AOB) or ammonia oxidizing archaea (AOA) and nitrite oxidizing bacteria (NOB). Mounting studies have examined the impact of nitrogen (N) fertilization on the dynamic and diversity of AOA and AOB, while we have limited information on the response of the activity, abundance, and diversity of NOB to N fertilization. We investigated the influence of organic and inorganic N fertilizers on soil NOB in silage corn field plots that received contrasting nitrogen (N) treatments: control (no additional N), ammonium sulfate (AS 100 and 200 kg N ha-1), and compost (200 kg N ha-1). Nitrifying community was examined using a universal marker (16S rRNA gene), functional gene markers (AOB amoA and Nitrospira nxrB), and metagenomics. The overall nitrifying community was not altered after the first fertilization but was significantly shifted by 4-year repeated application of ammonium fertilizers. Nitrospira were the dominant NOB (>99.7%) in our agricultural soil. Both community compositions of AOB and Nitrospira were significantly changed by ammonium fertilizers but not by compost after 4 years of repeated applications. All nitrifiers, including comammox, were recovered in soil metagenomes based on a gene-targeted assembly, but their sequence counts were very low. Although N treatment did not affect the abundance of Nitrospira nxrB determined by real-time quantitative PCR, ammonium fertilizers significantly promoted rates of potential nitrite oxidation determined at 0.15 mM nitrite in soil slurries. Understanding the response of both ammonia oxidizers and nitrite oxidizers to N fertilization may initiate or improve strategies for mitigating potential environmental impacts of nitrate production in agricultural ecosystems.

Environmental variables better explain changes in potential nitrification and denitrification activities than microbial properties in fertilized forest soils.

Because of increases in atmospheric nitrogen (N) deposition worldwide, nutrient imbalances and phosphorus (P) limitations in soil are aggravated, with the result that P fertilizer applications to terrestrial ecosystems worldwide may increase. Nitrification and denitrification in soil are major sources of nitrous oxide emissions, especially in soils treated with fertilizers. However, few researchers have studied how forest soils respond to nutrient additions, so we are not sure how the potential nitrification and denitrification activities (PNA and PDA, respectively) and microbial communities involved in these processes might respond when N and P are added to temperate and subtropical forest soils. We investigated how the PNA, PDA, the abundances and community compositions of nitrifiers and denitrifiers, and environmental properties, including soil pH, soil total and dissolved organic carbon, total and available N and phosphorus P, changed when N and/or P were added to subtropical and temperate forest soils. We quantified the abundance, and analyzed the composition, of functional marker genes of nitrifiers (ammonia-oxidizing bacteria and archaea amoA) and denitrifiers (nirK and nirS) using quantitative PCR and sequencing, respectively. We found that the PNA and PDA in the subtropical soil increased when P was added and PNA in the temperate forest soil increased when either N or P was added. The PNA and PDA were positively correlated with the abundance of ammonia-oxidizing bacteria and nirK-denitrifiers, respectively, in the subtropical forest soil but were not correlated with changes in corresponding community compositions in either of the forest soils. The soil total N to total P ratio explained most of the variabilities in the PNA and PDA in the subtropical forest soils, and the soil exchangeable ammonium concentrations and pH were the main controls on the PNA and PDA, respectively, in the temperate forest soils. Our results indicate that soil environmental conditions have more influence on variations in the PNA and PDA in forest soils fertilized with N and P than the corresponding microbial properties.

Understanding the relationships between grazing intensity and the distribution of nitrifying communities in grassland soils.

Nitrifying microbes are of critical importance in regulating efficient nitrogen (N) cycling, which plays a crucial role in plant productivity and maintaining soil sustainability. Long-term different intensities of grazing can strongly influence the microbial communities, while our understanding of the complex nitrifying community in the grazed grassland soil environment is still limited. To investigate whether and how long-term grazing with different intensities influence soil nitrifying communities, high-throughput sequencing and quantitative PCR analyses were performed on soil samples from permanent grassland soils under four grazing intensities: 0 (G0), 1.5 (G1), 6 (G2) and 9 (G3) sheepha-1. Results showed that the G3 treatment significantly reduced the soil nutrient content and increased the soil bulk density, changes that are not sustainable in the long run. The G1 treatment, on the other hand, significantly increased the soil nutrient content and would improve soil fertility. Some functional microbes were specifically enriched after long term grazing, like Nitrospirae (phylum) to Nitrospira (class) in the G2 samples and Chromatiales (order) to Nitrosococcus (genus) in the G3 soils. The numerically dominant Nitrosococcus watsonii lineage of ammonia oxidizing bacteria (AOB) was observed in this grassland soil. The redundancy analysis (RDA) together with the structural equation modeling (SEM) analysis showed that grazing intensity was important in mediating the distribution of soil microorganisms and affected nitrifying communities by impacting soil physicochemical characteristics (e.g., bulk density, NH4+-N). These results showed the shifts of nitrifying communities across different grazing intensities, and could aid in the determination of an optimal grazing intensity for these grazed grassland soils.

Nitrite oxidizing bacteria (NOB) dominating in nitrifying community in full-scale biological nutrient removal wastewater treatment plants.

Nitrification activities and microbial populations of ammonium oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB) were investigated in 10 full-scale biological nutrient removal wastewater treatment plants in Xi'an, China. Aerobic batch tests were used to determine the nitrifying activities while fluorescence in situ hybridization was used to quantify the fractions of AOB and NOB in the activated sludge. The results showed that nitrifying bacteria accounted for 1-10% of the total population. Nitrosomonas and Nitrospira were the dominant bacteria for AOB and NOB respectively. Moreover, the average percentage of AOB was 1.27% and that of NOB was 4.02%. The numerical ratios of NOB/AOB varied between 1.72 and 5.87. The average ammonium uptake rate and nitrite uptake rate were 3.25 ± 0.52 mg (NH4+-N)/g(VSS) h and 4.49 ± 0.49 mg (NO2--N)/g(VSS) h, respectively. Correspondingly, the activity of NOB was 1.08-2.00 times higher than that of AOB. Thus, NOB was the dominating bacteria in nitrifying communities. The year-round data of Dianzicun (W6) also expressed a similar trend. Since NOB had higher activities than that of AOB, a large nitrite oxidation pool could be formed, which guaranteed that no nitrite would be accumulated. Therefore, stable nitrification could be achieved. A conceptual model was proposed to describe the population variation of AOB and NOB in a nitrifying community.

Linking nitrifying biofilm characteristics and nitrification performance in moving-bed biofilm reactors for polluted raw water pretreatment.

Biofilm physiology was characterized by four biofilm constituents, i.e., polysaccharides, proteins (PN), humic-like substances and phospholipids (PL), for the first time to explore the relationships between biofilm characteristics and nitrification performance in moving-bed biofilm reactors (MBBRs) designed for pretreatment of polluted raw surface water for potable supply. The biofilm compositions depended highly on the balance of microbial decay and nitrification processes. The increased ammonia loading greatly regulated the community structure, promoting the dominance of nitrifiers and their proportions in the nitrifying biofilm. Nitrification rate and activity correlated linearly with the fractions of volatile solids (VS), PN and PL, which were related to nitrification processes in the biofilm. The specific biofilm activity demonstrated an exponential-asymptotic relationship with ratios of PN/VS and PL/VS. Thus, analyzing biofilm characteristics can be valid for estimating nitrification performance in MBBRs, and may offer engineers with basis to optimize MBBR design and operation.