Structural and functional bacterial biodiversity in a copper, zinc and nickel amended bioreactor: shotgun metagenomic study.

BACKGROUND: At lower concentrations copper (Cu), zinc (Zn) and nickel (Ni) are trace metals essential for some bacterial enzymes. At higher concentrations they might alter and inhibit microbial functioning in a bioreactor treating wastewater. We investigated the effect of incremental concentrations of Cu, Zn and Ni on the bacterial community structure and their metabolic functions by shotgun metagenomics. Metal concentrations reported in previous studies to inhibit bacterial metabolism were investigated. RESULTS: At 31.5 µM Cu, 112.4 µM Ni and 122.3 µM Zn, the most abundant bacteria were Achromobacter and Agrobacterium. When the metal concentration increased 2 or fivefold their abundance decreased and members of Delftia, Stenotrophomonas and Sphingomonas dominated. Although the heterotrophic metabolic functions based on the gene profile was not affected when the metal concentration increased, changes in the sulfur biogeochemical cycle were detected. Despite the large variations in the bacterial community structure when concentrations of Cu, Zn and Ni increased in the bioreactor, functional changes in carbon metabolism were small. CONCLUSIONS: Community richness and diversity replacement indexes decreased significantly with increased metal concentration. Delftia antagonized Pseudomonas and members of Xanthomonadaceae. The relative abundance of most bacterial genes remained unchanged despite a five-fold increase in the metal concentration, but that of some EPS genes required for exopolysaccharide synthesis, and those related to the reduction of nitrite to nitrous oxide decreased which may alter the bioreactor functioning.

Fine-Scale Spatial Variability of Greenhouse Gas Emissions From a Subantarctic Peatland Bog.

Peatlands are recognized as crucial greenhouse gas sources and sinks and have been extensively studied. Their emissions exhibit high spatial heterogeneity when measured on site using flux chambers. However, the mechanism by which this spatial variability behaves on a very fine scale remains unclear. This study investigates the fine-scale spatial variability of greenhouse gas emissions from a subantarctic Sphagnum peatland bog. Using a recently developed skirt chamber, methane emissions and ecosystem respiration (as carbon dioxide) were measured at a submeter scale resolution, at five specific 3 × 3 m plots, which were examined across the site throughout a single campaign during the Austral summer season. The results indicated that methane fluxes were significantly less homogeneously distributed compared with ecosystem respiration. Furthermore, we established that the spatial variation scale, i.e., the minimum spatial domain over which notable changes in methane emissions and ecosystem respiration occur, was <0.56 m2. Factors such as ground height relative to the water table and vegetation coverage were analyzed. It was observed that Tetroncium magellanicum exhibited a notable correlation with higher methane fluxes, likely because of the aerenchymatous nature of this species, facilitating gas transport. This study advances understanding of gas exchange patterns in peatlands but also emphasizes the need for further efforts for characterizing spatial dynamics at a very fine scale for precise greenhouse gas budget assessment.

Toilet effluent separation and brown water treatment: Survey and initial feasibility testing in Mexico.

Separation of domestic effluents at the source and the utilization of low-flush toilets offer alternative approaches for developing efficient wastewater treatment systems while promoting energy generation through anaerobic digestion. This study focused on assessing toilet usage in Mexico and exploring the potential of anaerobic co-digestion of brown water (feces) and toilet paper as influential factors in wastewater treatment systems. A survey was conducted on a representative sample of Mexicans to gather information on toilet usage frequency, toilet paper use and disposal practices, as well as the type and quantity of commercial disinfectants and pharmaceutical compounds they use or consume. The survey revealed that per capita toilet paper consumption is 2.9 kg annually, that 58 % of respondents do not dispose used paper in the toilet, and that about 47 % use two to three cleaning and disinfection products. Notably, 97 % of the sampled Mexican population expressed a willingness to transition to more eco-friendly toilet options. Subsequently, in a second step, the anaerobic co-digestion of brown water with toilet paper was evaluated, demonstrating a relatively high production of volatile fatty acids but low methane production. This suggests an efficient hydrolysis/acidogenesis process coupled with restrained methanogenesis, probably due to pH decrease caused by acidogenesis. This study underscores that toilet paper and brown water are potential suitable substrates for anaerobic co-digestion. Furthermore, it sheds light on the behaviors of Mexican society regarding bathroom use and cleaning, contributing to the establishment of foundations for wastewater treatment systems with effluent separation at the source.

Impacts of leaks and gas accumulation on closed chamber methods for measuring methane and carbon dioxide fluxes from tree stems.

Accurate measurements of methane (CH4) and carbon dioxide (CO2) fluxes from tree stems are important for understanding greenhouse gas emissions. Closed chamber methods are commonly employed for this purpose; however, leaks between the chamber and the atmosphere as well as gas accumulation, known as the concentration buildup effect, can impact flux measurements significantly. In this study, we investigated the impacts of concentration buildup and leaks on semi-rigid closed chamber methods. Field measurements were conducted on six tree species, including three species from a Mexican mangrove ecosystem and three species from a Magellanic sub-Antarctic forest. Systematic observations revealed significant leak flow rates, ranging from 0.00 to 465 L h-1, with a median value of 1.25 ± 75.67 L h-1. We tested the efficacy of using cement to reduce leaks, achieving a leak flow rate reduction of 46-98 % without complete elimination. Our study also demonstrates a clear and substantial impact of concentration buildup on CH4 flux measurements, while CO2 flux measurements were relatively less affected across all tree species studied. Our results show that the combined effects of leaks and concentration buildup can lead to an underestimation of CH4 emissions by an average of 40 ± 20 % and CO2 emissions by 22 ± 22 %, depending on the bark roughness. Based on these findings, we recall a straightforward yet effective method to minimize experimental errors associated with these phenomena, previously established, and reiterated in the current context, for calculating emissions that considers effects of leaks and concentration buildup, while eliminating the need for separate determinations of these phenomena. Overall, the results, combined with a literature review, suggest that our current estimates of GHG flux from tree stems are currently underestimated.

Methane and carbon dioxide cycles in lakes of the King George Island, maritime Antarctica.

Freshwater ecosystems are important contributors to the global greenhouse gas budget and a comprehensive assessment of their role in the context of global warming is essential. Despite many reports on freshwater ecosystems, relatively little attention has been given so far to those located in the southern hemisphere and our current knowledge is particularly poor regarding the methane cycle in non-perennially glaciated lakes of the maritime Antarctica. We conducted a high-resolution study of the methane and carbon dioxide cycle in a lake of the Fildes Peninsula, King George Island (Lat. 62°S), and a succinct characterization of 10 additional lakes and ponds of the region. The study, done during the ice-free and the ice-seasons, included methane and carbon dioxide exchanges with the atmosphere (both from water and surrounding soils) and the dissolved concentration of these two gases throughout the water column. This characterization was complemented with an ex-situ analysis of the microbial activities involved in the methane cycle, including methanotrophic and methanogenic activities as well as the methane-related marker gene abundance, in water, sediments and surrounding soils. The results showed that, over an annual cycle, the freshwater ecosystems of the region are dominantly autotrophic and that, despite low but omnipresent atmospheric methane emissions, they act as greenhouse gas sinks.

Spatial and seasonal dynamics of the methane cycle in a tropical coastal lagoon and its tributary river.

Coastal aquatic ecosystems such as estuaries and coastal lagoons are important atmospheric methane sources that must be better constrained. This work presents a detailed characterization of the methane cycle in a tropical coastal lagoon (La Mancha, Veracruz, Mexico) and its tributary river over three distinct seasons, along a transect from the river to the sea connection. In addition to several physicochemical parameters, the dissolved methane, carbon dioxide, and oxygen concentrations were measured with high resolution in the sediments and the water column, combined with production/uptake rates. Methane and carbon dioxide cycles were further constrained by determining atmospheric flux over the entire river and lagoon sections. The results indicate that La Mancha is a highly contrasted ecosystem. The river section is characterized by a strong pycnocline, relatively high methane concentration, and active methanogenesis and methanotrophy, discharging into a relatively homogeneous lagoon section where the methane and carbon cycles are less active. Overall, both the river and the lagoon were a net source of methane and carbon dioxide, with an annual emission of 2.9 metric tons of methane and 2757 metric tons of carbon dioxide. The spatial structure of the main components of the methane, carbon dioxide, and oxygen cycles was established, and it was observed that depthwise heterogeneities predominated in the river section. In contrast, lengthwise heterogeneities dominated in the lagoon section.

Temperature differently affected methanogenic pathways and microbial communities in sub-Antarctic freshwater ecosystems.

Freshwater ecosystems are responsible for an important part of the methane (CH4) emissions which are likely to change with global warming. This study aims to evaluate temperature-induced (from 5 to 20 °C) changes on microbial community structure and methanogenic pathways in five sub-Antarctic lake sediments from Magallanes strait to Cape Horn, Chile. We combined in situ CH4 flux measurements, CH4 production rates (MPRs), gene abundance quantification and microbial community structure analysis (metabarcoding of the 16S rRNA gene). Under unamended conditions, a temperature increase of 5 °C doubled MPR while microbial community structure was not affected. Stimulation of methanogenesis by methanogenic precursors as acetate and H2/CO2, resulted in an increase of MPRs up to 127-fold and 19-fold, respectively, as well as an enrichment of mcrA-carriers strikingly stronger under acetate amendment. At low temperatures, H2/CO2-derived MPRs were considerably lower (down to 160-fold lower) than the acetate-derived MPRs, but the contribution of hydrogenotrophic methanogenesis increased with temperature. Temperature dependence of MPRs was significantly higher in incubations spiked with H2/CO2 (c. 1.9 eV) compared to incubations spiked with acetate or unamended (c. 0.8 eV). Temperature was not found to shape the total microbial community structure, that rather exhibited a site-specific variability among the studied lakes. However, the methanogenic archaeal community structure was driven by amended methanogenic precursors with a dominance of Methanobacterium in H2/CO2-based incubations and Methanosarcina in acetate-based incubations. We also suggested the importance of acetogenic H2-production outcompeting hydrogenotrohic methanogenesis especially at low temperatures, further supported by homoacetogen proportion in the microcosm communities. The combination of in situ-, and laboratory-based measurements and molecular approaches indicates that the hydrogenotrophic pathway may become more important with increasing temperatures than the acetoclastic pathway. In a continuously warming environment driven by climate change, such issues are crucial and may receive more attention.

Overall spatiotemporal dynamics of greenhouse gasses and oxygen in two subtropical reservoirs with contrasting trophic states.

The impact of cultural eutrophication on carbon cycling in subtropical reservoirs was assessed using high-resolution measurements of dissolved gas concentration, atmospheric exchange, and uptake/production rates of methane, carbon dioxide, and oxygen. Seasonal measurements were performed in two reservoirs that pertain to the same hydrological basin but are drastically different in terms of allochthonous carbon input. These results were used to feed a mass balance model, from which a large number of overall parameters were determined to explicitly describe the dynamics and spatial attributes of the carbon cycle in the reservoirs. A single graphical representation of each reservoir was created to facilitate an overall appraisal of the carbon cycle. The impact of cultural eutrophication was profound and resulted in a complete redistribution of how the various bioprocesses participated in the methane, carbon dioxide, and oxygen cycles. Among several identified impacts of eutrophication, it was observed that while eutrophication triggered increased methane production, this effect was followed by a similar increase in methane emissions and methanotrophic rates, while gross primary production was depleted.

Spatial and temporal distribution of methane emissions from a covered landfill equipped with a gas recollection system.

A previously developed surface probe method, which allows for instantaneous methane (CH4) flux measurement, was used to establish CH4 emission maps of a municipal landfill with a final clay cover and equipped with a gas recollection system. In addition to spatial variations, the method was applied at 7 different times over a total timeframe of 65 h and under similar weather conditions to determine the intrinsic temporal variations of CH4 emissions; i.e., the temporal variation related to the dynamic of the landfill rather than the one driven by external factors. Furthermore, continuous CH4 fluxes, with a data acquisition frequency of 1 Hz, were measured during 12 h at a single position, and for one hour at 22 locations of the landfill, spanning a large range of CH4 emission magnitudes. A simple model for the numerical characterization of spatiotemporal variability of the landfill emission was used and allowed us to separately quantify the temporal and spatial variability. This model showed that, in the landfill tested, the temporal distribution of CH4 emissions resulted more homogeneous than the spatial distribution. Other attributes of the temporal and spatial distributions of CH4 emissions were also established including the anisotropic nature of the spatial distribution and, contrastingly, the stochastic temporal variability of such emissions.

Sub-oxycline methane oxidation can fully uptake CH4 produced in sediments: case study of a lake in Siberia.

It is commonly assumed that methane (CH4) released by lakes into the atmosphere is mainly produced in anoxic sediment and transported by diffusion or ebullition through the water column to the surface of the lake. In contrast to that prevailing idea, it has been gradually established that the epilimnetic CH4 does not originate exclusively from sediments but is also locally produced or laterally transported from the littoral zone. Therefore, CH4 cycling in the epilimnion and the hypolimnion might not be as closely linked as previously thought. We utilized a high-resolution method used to determine dissolved CH4 concentration to analyze a Siberian lake in which epilimnetic and hypolimnetic CH4 cycles were fully segregated by a section of the water column where CH4 was not detected. This layer, with no detected CH4, was well below the oxycline and the photic zone and thus assumed to be anaerobic. However, on the basis of a diffusion-reaction model, molecular biology, and stable isotope analyses, we determined that this layer takes up all the CH4 produced in the sediments and the deepest section of the hypolimnion. We concluded that there was no CH4 exchange between the hypolimnion (dominated by methanotrophy and methanogenesis) and the epilimnion (dominated by methane lateral transport and/or oxic production), resulting in a vertically segregated lake internal CH4 cycle.

Microbial community dynamics during aerobic granulation in a sequencing batch reactor (SBR).

Microorganisms in aerobic granules formed in sequencing batch reactors (SBR) remove contaminants, such as xenobiotics or dyes, from wastewater. The granules, however, are not stable over time, decreasing the removal of the pollutant. A better understanding of the granule formation and the dynamics of the microorganisms involved will help to optimize the removal of contaminants from wastewater in a SBR. Sequencing the 16S rRNA gene and internal transcribed spacer PCR amplicons revealed that during the acclimation phase the relative abundance of Acinetobacter reached 70.8%. At the start of the granulation phase the relative abundance of Agrobacterium reached 35.9% and that of Dipodascus 89.7% during the mature granule phase. Fluffy granules were detected on day 43. The granules with filamentous overgrowth were not stable and they lysed on day 46 resulting in biomass wash-out. It was found that the reactor operation strategy resulted in stable aerobic granules for 46 days. As the reactor operations remained the same from the mature granule phase to the end of the experiment, the disintegration of the granules after day 46 was due to changes in the microbial community structure and not by the reactor operation.

Methane dynamics in the subsaline ponds of the Chihuahuan Desert: A first assessment.

The Cuatro Cienegas Basin (CCB) in the Chihuahuan desert is characterized by the presence of over 500 ponds located in an endorheic basin. These ponds are subsaline ecosystems characterized by a low productivity and a particularly high sulfate concentration, comparable to marine environments. This study focused on assessing the main physicochemical parameters in these ponds along with the characterization of the CH4 dynamics through the determination of fluxes, dissolved CH4 concentrations, and net methanotrophic and methanogenic activity. Despite a sulfate concentration ranging from 1.06 to 4.73 g L-1, the studied ponds showed moderate but clear CH4 production and emission, which suggests that methanogenesis is not completely outcompeted by sulfate reduction. CH4 fluxes ranged from 0.12 to 0.98 mg m-2 d-1, which falls within the higher range of marine emissions and within the lower range reported for coastal saline lagoons and saline ponds. During summer, significant CH4 production in the oxic water column was observed. In addition to CH4, CO2 fluxes were determined at levels from 0.2 to 53 g m-2 d-1, which is within the range recorded for saline lakes in other parts of the world. Our results provide additional evidence that subsaline/saline aquatic ecosystems play an important role in the emission of greenhouse gases to the atmosphere.

Continuous Measurement of Diffusive and Ebullitive Fluxes of Methane in Aquatic Ecosystems by an Open Dynamic Chamber Method.

An open dynamic chamber for the continuous monitoring of diffusive and ebullitive fluxes of methane (CH4) in aquatic ecosystems was designed and developed. This method is based on a standard floating chamber in which a well-defined carrier gas flows. The concentration of CH4 is measured continuously at the outlet of the chamber, and the flux is determined from a mass balance equation. The method was carefully tested in a laboratory and was subsequently applied to two lakes, in Mexico, with contrasting trophic states. We show here that the method allows for the continuous quantification of CH4 diffusive flux higher than 25 × 10-6 g m-2 h-1, the determination of ebullitive flux, and the individual characterization of bubbles larger than 1.50-1.72 mm in diameter. The method was also applied to determine carbon dioxide emissions (CO2). In that case, the method was less sensitive but allowed for the characterization of diffusive fluxes higher than 10 mg CO2 m-2 h-1 and of bubbles larger than 5.3-8.4 mm in diameter. This high-throughput method can be adapted to any gas detector at low cost, making it a convenient tool to better constrain greenhouse gas emission from freshwater ecosystems.

Electron shuttling mediated by humic substances fuels anaerobic methane oxidation and carbon burial in wetland sediments.

Key pathways for the anaerobic oxidation of methane (AOM) have remained elusive, particularly in organic rich ecosystems. In this work, the occurrence of AOM driven by humus-catalyzed dissimilatory iron reduction was investigated in sediments from a coastal mangrove swamp. Anoxic sediment incubations supplied with both goethite (α-FeOOH) and leonardite (humic substances (HS)) displayed an average AOM rate of 10.7 ±â€¯0.8 µmol CH4 cm-3 day-1, which was 7 and 3 times faster than that measured in incubations containing only goethite or HS, respectively. Additional incubations performed with 13C-methane displayed Pahokee Peat HS-mediated carbonate precipitation linked to 13CH4 oxidation and ferrihydrite reduction (~1.3 µmol carbonate cm-3 day-1). These results highlight the role of HS on mitigating greenhouse gases released from wetlands, not only by mediating the AOM process, but also by enhancing carbon sequestration as inert minerals (calcite, aragonite and siderite).

Multiparameter analysis of activated sludge inhibition by nickel, cadmium, and cobalt.

Activated sludge processes are often inhibited by nickel, cadmium, and cobalt. The inhibitory effect of these heavy metals on a synthetic wastewater treatment process was tested through pulse microrespirometry; i.e., pulse of substrate injected in a microreactor system. The inhibitory effect was tested under different conditions including the heavy metals, substrate and biomass concentrations, and exposure time. The inhibitory effect was quantified by the percentage of inhibition, half saturation constant (KS), inhibition constant (KI), and maximum oxygen uptake rate (OURmax). The results indicated that, in a range of concentration from 0 to 40 mg L-1, the three heavy metals exerted an uncompetitive and incomplete inhibitory effect, with a maximum inhibition of 67, 57, and 53% for Ni, Co, and Cd, respectively. An increase of the biomass concentration by 620% resulted in a decrease of the inhibition by 47 and 69% for Co and Cd, respectively, while no effect was observed on Ni inhibition. An increase of the substrate concentration by 87% resulted in an increase of the inhibition by 24, 70, and 47% for Ni, Co and Cd, respectively. In the case of nickel and cadmium, an increase in the exposure time to the heavy metals also increased the inhibition.

Microbial community structure in aerobic and fluffy granules formed in a sequencing batch reactor supplied with 4-chlorophenol at different settling times.

Toxic compounds, such as 4-chlorophenol (4-CP), which is a common pollutant in wastewater, are removed efficiently from sequencing batch reactors (SBRs) by microorganisms. The bacterial community in aerobic granules formed during the removal of 4-CP in a SBR was monitored for 63days. The SBR reactor was operated with a constant filling and withdrawal time of 7 and 8min and decreasing settling time (30, 5, 3 and 2min) to induce the formation of aerobic granules. During the acclimation period lasting 15days (30min settling time) had a strong effect on the bacterial community. From day 18 onwards, Sphingobium and Comamonadaceae were detected. Rhizobiaceae were dominant from day 24 to day 28 when stable aerobic granules were formed. At day 35, fluffy granules were formed, but the bacterial community structure did not change, despite the changes in the reactor operation to inhibit filamentous bacteria growth. This is the first report on changes in the bacterial community structure of aerobic and fluffy granules during granulation process in a reactor fed with 4-CP and the prediction of its metabolic pathways.

Initial report on methane and carbon dioxide emission dynamics from sub-Antarctic freshwater ecosystems: A seasonal study of a lake and a reservoir.

The sub-Antarctic Magellanic ecoregion is a part of the world where ecosystems have been understudied and where the CH4 cycling and emissions in lakes has not ever been reported. To fill that knowledge gap, a lake and a reservoir located at 53°S were selected and studied during three campaigns equally distributed over one year. Among the parameters measured were CH4 and CO2 emissions, as well their dissolved concentrations in the water column, which were determined with high spatial resolution. No ebullition was observed and the CH4 flux ranged from 0.0094 to 4.47mmolm-2d-1 while the CO2 flux ranged from -22.95 to 35.68mmolm-2d-1. Dissolved CH4 concentrations varied over more than four orders of magnitude (0.025-128.75µmolL-1), and the dissolved carbon dioxide ranged from below the detection limit of our method (i.e., 0.15µmolL-1) to 379.09µmolL-1. The high spatial resolution of the methods used enabled the construction of bathymetric maps, surface contour maps of CH4 and CO2 fluxes, and transect contour maps of dissolved oxygen, temperature, and dissolved greenhouse gases. Overall, both lakes were net greenhouse gas producers and were not significantly different from temperate lakes located at a similar northern latitudes (53°N), except that ebullition was never observed in the studied sub-Antarctic lakes.

Bacterial community structure within an activated sludge reactor added with phenolic compounds.

Biodegradation of phenolic compounds in bioreactors is well documented, but the changes in the bacterial populations dynamics during degradation were not that often. A glass bubble column used as reactor was inoculated with activated sludge, spiked with 2-chlorophenol, phenol and m-cresol after 28 days and maintained for an additional 56 days, while the 16S rRNA gene from metagenomic DNA was monitored. Proteobacteria (68.1%) dominated the inoculum, but the bacterial composition changed rapidly. The relative abundance of Bacteroidetes and Firmicutes decreased from 4.8 and 9.4 to <0.1 and 0.2% respectively, while that of Actinobacteria and TM7 increased from 4.8 and 2.0 to 19.2 and 16.1% respectively. Phenol application increased the relative abundance of Proteobacteria to 94.2% (mostly Brevundimonas 17.6%), while that of Bacteroidetes remained low (1.2%) until day 42. It then increased to 47.3% (mostly Leadbetterella 46.9%) at day 84. It was found that addition of phenolic compounds did not affect the relative abundance of the Alphaproteobacteria initially, but it decreased slowly while that of the Bacteroidetes increased towards the end.

Hotspot detection and spatial distribution of methane emissions from landfills by a surface probe method.

A surface probe method previously developed was used to detect hotspots and to determine spatial variation of methane (CH4) emissions from three landfills located in Mexico, with an intermediate or a final cover, as well as with or without a landfill gas collection system. The method was effective in the three landfills and allowed mapping of CH4 emissions with a resolution of 24-64 measurements per hectare, as well as the detection and quantification of hotspots, with a moderate experimental effort. In the three selected landfills, CH4 emissions were quantified to 10, 72, and 575gm(-2)d(-1). Two straightforward parameters describing the spatial distribution of CH4 emissions were also developed. The first parameter provides the percentage of area responsible for a given percentage of total emissions, while the second parameter assigns a numerical value to flux homogeneity. Together, the emissions map and the spatial distribution parameters offer an appropriate tool to landfill operators willing to begin recovering CH4 emissions or to improve the effectiveness of an existing recovery system. This method may therefore help to reduce the greenhouse gas footprint of landfills, which are still the primary option for waste management in developing countries.

Kinetic Constants for Biological Ammonium and Nitrite Oxidation Processes Under Sulfide Inhibition.

Inhibition of nitrification by sulfide was assessed using sludge obtained from a steady-state nitrifying reactor. Independent batch activity assays were performed with ammonium and nitrite as substrate, in order to discriminate the effect of sulfide on ammonium and nitrite oxidation. In the absence of sulfide, substrate affinity constants (K S,NH4 = 2.41 ± 0.11 mg N/L; K s, NO2 = 0.74 ± 0.03 mg N/L) and maximum specific rates (q max,NH4 = 0.086 ± 0.008 mg N/mg microbial protein h; q max,NO2 = 0.124 ± 0.001 mg N/mg microbial protein h) were determined. Inhibition of ammonium oxidation was no-competitive (inhibition constant (K i , NH4 ) of 2.54 ± 0.12 mg HS(-)-S/L) while inhibition of nitrite oxidation was mixed (competitive inhibition constant (K' i , NO2 ) of 0.22 ± 0.03 mg HS(-)-S/L and no-competitive inhibition constant (K i , NO2 ) of 1.03 ± 0.06 mg HS(-)-S/L). Sulfide has greater inhibitory effect on nitrite oxidation than ammonium oxidation, and its presence in nitrification systems should be avoided to prevent accumulation of nitrite. By simulating the effect of sulfide addition in a continuous nitrifying reactor under steady-state operation, it was shown that the maximum sulfide concentration that the sludge can tolerate without affecting the ammonium consumption efficiency and nitrate yield is 1 mg HS(-)-S/L.