Characteristics of Nitrate Reduction Using Fe (II) as Electron Donor in Activated Sludge

Static experiments were conducted to investigate the effects of environmental factors on nitrate (NO₃^?-N)-removal efficiency, such as NO₃^?-N loading, pH value, C/N ratio and temperature in activated sludge using Fe (II) as electron donor. The results demonstrated that the average denitrification rate increased from 1.25 to 2.23 mg NO₃^?-N/(L·h) with NO₃^?-N loading increased from 30 to 60 mg/L. When pH increased from 7 to 8, the concentration of NO₃^?-N and nitrite (NO₂^?-N) in effluent were all maintained at quite low levels. C/N ratio had little impact on denitrification process, i.e., inorganic carbon (C) source could still be enough for denitrification process with C/N ratio as low as 5. Temperature had a significant effect on the denitrification efficiency, and NO₃^?-N removal efficiency of 92.03%, 96.77%, 97.67% and 98.23% could be obtained with temperature of 25°C, 30°C, 35°C and 40°C, respectively. SEM, XRD and XRF analysis was used to investigate microscopic surface morphology and chemical composition of the denitrifying activated sludge, and mechanism of the nitrate-dependent anaerobic ferrous oxidation (NAFO) bacterias could be explored with this research.     

Bioaugmented sulfur-oxidizing denitrification system with <Emphasis Type="Italic">Alcaligenes defragrans</Emphasis> B21 for high nitrate containing wastewater treatment

The performance of enriched sludge augmented with the B21 strain of Alcaligenes defragrans was compared with that of enriched sludge, as well as with pure Alcaligenes defragrans B21, in the context of a sulfur-oxidizing denitrification (SOD) process. In synthetic wastewater treatment containing 100–1,000 mg NO₃⁻-N/L, the single strain-seeded system exhibited superior performance, featuring higher efficiency and a shorter startup period, provided nitrate loading rate was less than 0.2 kg NO₃⁻-N/m³ per day. At nitrate loading rate of more than 0.5 kg NO₃⁻-N/m³ per day, the bioaugmented sludge system showed higher resistance to shock loading than two other systems. However, no advantage of the bioaugmented system over the enriched sludge system without B21 strain was observed in overall efficiency of denitrification. Both the bioaugmented sludge and enriched sludge systems obtained stable denitrification performance of more than 80% at nitrate loading rate of up to 2 kg NO₃⁻-N/m³ per day.     

Simultaneous oxidation of ammonium and p-cresol linked to nitrite reduction by denitrifying sludge

The metabolic capability of denitrifying sludge to oxidize ammonium and p-cresol was evaluated in batch cultures. Ammonium oxidation was studied in presence of nitrite and/or p-cresol by 55 h. At 50 mg/L NH₄⁺-N and 76 mg/L NO₂⁻-N, the substrates were consumed at 100% and 95%, respectively, being N₂ the product. At 50 mg/L NH₄⁺-N and 133 mg/L NO₂⁻-N, the consumption efficiencies decreased to 96% and 70%, respectively. The increase in nitrite concentration affected the ammonium oxidation rate. Nonetheless, the N₂ production rate did not change. In organotrophic denitrification, the p-cresol oxidation rate was slower than ammonium oxidation. In litho-organotrophic cultures, the p-cresol and ammonium oxidation rates were affected at 133 mg/L NO₂⁻-N. Nonetheless, at 76 mg/L NO₂⁻-N the denitrifying sludge oxidized ammonium and p-cresol, but at different rate. Finally, this is the first work reporting the simultaneous oxidation of ammonium and p-cresol with the production of N₂ from denitrifying sludge.     

Simultaneous biological removal of sulfur, nitrogen and carbon using EGSB reactor

High-rate biological conversion of sulfide and nitrate in synthetic wastewater to, respectively, elemental sulfur (S⁰) and nitrogen-containing gas (such as N₂) was achieved in an expanded granular sludge bed (EGSB) reactor. A novel strategy was adopted to first cultivate mature granules using anaerobic sludge as seed sludge in sulfate-laden medium. The cultivated granules were then incubated in sulfide-laden medium to acclimate autotrophic denitrifiers. The incubated granules converted sulfide, nitrate, and acetate simultaneously in the same EGSB reactor to S⁰, N-containing gases and CO₂ at loading rates of 3.0 kg S m⁻³ d⁻¹, 1.45 kg N m⁻³ d⁻¹, and 2.77 kg Ac m⁻¹ d⁻¹, respectively, and was not inhibited by sulfide concentrations up to 800 mg l⁻¹. Effects of the C/N ratio on granule performance were identified. The granules cultivated in the sulfide-laden medium have Pseudomonas spp. and Azoarcus sp. presenting the heterotrophs and autotrophs that co-work in the high-rate EGSB-SDD (simultaneous desulfurization and denitrification) reactor.     

Effects of operating parameters on performances of nitrification and denitrification processes

Nitrification and denitrification of synthetic wastewater was studied by using two reactors in series. An activated sludge unit was used for nitrification followed by a downflow biofilter (packed column) for denitrification. A glucose solution was fed to the denitrification column to supply carbon source. Effects of important process variables such as sludge age, hydraulic residence time and feed ammonium concentration on system's performance were investigated. Effluent ammonium-nitrogen (NH4-N) concentration decreased with increasing sludge age and hydraulic residence time and remained constant for sludge age and hydraulic residence times greater than 12 d and 15 h, respectively. Feed ammonium-nitrogen concentration above 200 mg/l resulted in significant levels of NH4-N in the effluent at Š_c = 15 d and Š_H = 12 h in nitrification. Performance of denitrification stage was not satisfactory for feed NO₃-N concentrations above 150 mg N/l resulting in significant effluent NO₃-N levels at hydraulic residence time of Š_H = 6 h.     

Microbial community structure and dynamics in a mixotrophic nitrogen removal process using recycled spent caustic under different loading conditions

A laboratory-scale Bardenpho process was established to investigate the proper nitrogen loading rate (NLR) when modified spent caustic (MSC) is applied as electron donor and alkalinity source for denitrification. MSC injection induced autotrophic nitrogen removal with sulfur as electron donor and heterotrophic denitrification. The nitrogen removal rate (NRR) did not increase proportionally to NLR. Based on the total nitrogen concentration in the effluent observed in the trials with MSC, the NLR in the influent should not exceed 0.15 kg N/m³ d in order to satisfy water quality regulations. Microbial communities in the anoxic reactors were characterized by pyrosequencing of 16S rRNA gene sequences amplified by the polymerase chain reaction of DNA extracted from sludge samples. Microbial diversity was lower as MSC dosage was increased, and the injection of MSC caused an increase in SOB belonging to the genus Thiobacillus which is responsible for denitrification using sulfur.     

Denitrification of high nitrate wastewater in a cloth strip bioreactor with immobilized sludge

Denitrification of synthetic high nitrate wastewater containing 40,000?ppm NO₃ (9,032?ppm NO₃-N) was achieved using immobilized activated sludge in a column reactor. Active anoxic sludge adsorbed onto Terry cloth was used in the denitrification of high nitrate wastewater. The operational stability of the immobilized sludge system was studied both in a batch reactor and in a continuous reactor. The immobilized sludge showed complete degradation of different concentrations of NO₃-N (1,129, 1,693, 3,387, 6,774, and 9,032?ppm) in a batch process. The reactors were successfully run for 90?days without any loss in activity. The immobilized cell process has yielded promising results in attaining high denitrifying efficiency.     

Characterization of microbial community and kinetics for spent sulfidic caustic applied autotrophic denitrification

Spent sulfidic caustic was applied to sulfur utilizing autotrophic denitrification as the simultaneous source of electron donor and alkalinity. The two experiment set-up of upflow anoxic hybrid growth reactor (UAHGR) and upflow anoxic suspended growth reactor (UASGR) was adopted and nitrate removals were similar in both reactors. Approximately 90% of the initial nitrate was denitrified at nitrate loading rate of 0.15∼0.40 kgNO₃ ⁻/m³·d. The experimental stoichiometric ratio of sulfate production to nitrate removal was ranged from 1.5 to 2.1 mgSO₄ ²⁻/mgNO₃ ⁻. During the operation period, denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction (PCR)-amplified 16S rDNA fragments for the sludge sample of both reactors showed the change of microbial communities. Thiobacillus denitrificans-like microorganism occupied 28.5% (18 clones) of the 63 clones by cloning the PCR products from the sludge sample of UAHGR. Acidovorax avenae, which can reduce nitrate to nitrogen gas while oxidizing phenol (heterotrophic denitrifier), was also found in 7 clones (11.1%). Although an organic carbon source was not added to the medium, a microorganism (Kaistella koreensis) capable of oxidizing organic compounds was found in 7 clones (11.1%). Therefore, the microbial community of spent sulfidic caustic applied autotrophic denitrification process well corresponds to the substrate components of spent sulfidic caustic. Through the batch cultivation of microorganisms in UAHGR, the microbial kinetic coefficients of spent sulfidic caustic applied autotrophic denitrification were estimated to be μ _max = 0.097 h⁻¹, k _d = 0.0021 h⁻¹, K _s = 200 mgNO₃ ⁻/L, and Y = 0.31 mgMLVSS/mgNO₃ ⁻.     

Stimulating denitrification in a stream mesocosm with elemental sulfur as an electron donor

The heavy use of fertilizers in agricultural lands can result in significant nitrate (NO₃⁻) loadings to the aquatic environment. We hypothesized that biological denitrification in agricultural ditches and streams could be enhanced by adding elemental sulfur (S^o) to the sediment layer, where it could act as a biofilm support and electron donor. Using a bench-scale stream mesocosm with a bed of S^o granules, we explored NO₃⁻ removal fluxes as a function of the effluent NO₃⁻ concentrations. With effluent NO₃⁻ ranging from 0.5 mg N L⁻¹ to 4.1 mg N L⁻¹, NO₃⁻ removal fluxes ranged from 228 mg N m⁻² d⁻¹ to 708 mg N m⁻² d⁻¹. This is as much as 100 times higher than for agricultural drainage streams. Sulfate (SO₄²⁻) production was high due to aerobic sulfur oxidation. Molecular studies demonstrated that the S^o amendment selected for Thiobacillus species, and that no special inoculum was required for establishing a S^o-based autotrophic denitrifying community. Modeling studies suggested that denitrification was diffusion limited, and advective flow through the bed would greatly enhance NO₃⁻ removal fluxes. Our results indicate that amendment with S^o is an effective means to stimulate denitrification in a stream environment. To minimize SO₄²⁻ production, it may be better to place S^o deeper in the sediment layer.     

Process of inorganic nitrogen transformation and design of kinetics model in the biological aerated filter reactor

As one of the plug-flow reactors, biological aerated filter (BAF) reactor was divided into four sampling sectors to understand the characteristics of elemental nitrogen transformation during the reaction process, and then the different characteristics of elemental nitrogen transformation caused by different NH₃-N loadings, biological quantities and activities in each section were obtained. The results showed that the total transformation ratio in the nitrifying reactor was more than 90% in the absence of any organic carbon resource, at the same time, more than 65% NH₃-N in the influent were nitrified at the filter height of 70 cm below under the conditions of the influent runoff 9-19 L/h, the gas-water ratio 4-5:1, the dissolved oxygen 3.0-5.8 mg/L and the NH₃-N load 0.28-0.48 kg NH₃-N/m³ d. On the base of the Eckenfelder mode, the kinetics equation of the NH₃-N transformation along the reactor was S_e = S₀ exp(−0.0134D/L^1.2612).     

Estimation of Sediment Denitrification Rates at In Situ Nitrate Concentrations

The denitrification rates in a marine sediment, estimated by using ¹⁵N-nitrate, V_max, K_m, and sediment nitrate concentrations, were 12.5 and 2.0 nmol of N₂-N cm⁻³ day⁻¹ at 0 to 1 and 1 to 3 cm, respectively, at 12°C. The total rate was 165 nmol of N₂-N m⁻² day⁻¹.     

A new method of autotrophic denitrification with spent sulfidic caustic as substrate and alkalinity source

A new method based on sulfide utilizing autotrophic denitrification was adopted to remove nitrate from wastewater and to reuse spent sulfidic caustic containing high sulfide and alkalinity levels. The experiments were performed using a bench-scale upflow anoxic hybrid growth reactor (UAHGR) and an upflow anoxic suspended growth reactor (UASGR) to characterize the stoichiometric relationship between sulfur and nitrate in the process as well as the performance of the reactors. The level of nitrate removal from the UAHGR and UASGR were maintained at over 90% at a nitrate loading rate ranging from 0.15∼0.40 kgNO₃ ⁻/m³·d and no significant nitrite accumulation was observed in either reactor. Although the influent pH values were higher than the optimum range of autotrophic denitrification at 8.7∼10.1, the effluent pH was stable at 7.2∼7.9 due to the production of hydrogen ions during operation. The stoichiometric ratio of sulfate production to nitrate removal was 1.5∼2.1 mgSO₄ ²⁻/mgNO₃ ⁻ in both reactors. A comparison of the reactor performance revealed that the chemical parameters of the UAHGR operation corresponded to a plug flow like type reactor while the chemical parameters of the UASGR operation corresponded to a completely stirred tank reactor like type reactor. UAHGR did not require sludge recycling due to the packed media while UASGR required 300∼700% sludge recycling. Therefore, spent sulfidic caustic could be used in the sulfur utilizing autotrophic denitrification processes as substrate and alkalinity sources.     

Enhancing biological nitrogen removal from tannery effluent by using the efficient Brachymonas denitrificans in pilot plant operations

Summary Laboratory scale and pilot plant reactors were inoculated with an efficient denitrifier, Brachymonas denitrificans(CCUG 45880), in order to evaluate whether a bio-augmentation approach can be used to enhance biological nitrogen removal from tannery effluents. To determine the effectiveness of the introduced strain, denitrifying activity in the activated sludge was monitored by nitrate uptake rate (NUR) measurement of NO₃-N. Fluorescent in situ hybridization (FISH) technique was used to monitor the growth of the augmented species. The laboratory scale nitrate removal efficiency with the introduced B. denitrificans (3.7±0.6 mg NO₃-N gVSS ⁻¹ h ⁻¹) was higher than that of the activated sludge without the addition of the bacteria (3.5±0.7 mg NO₃-N gVSS ⁻¹ h ⁻¹); the NUR in the pilot plant after and before the introduction of the strain was also of the magnitude of 12.0±1.4 and 10.6±1.4 mg NO₃-N gVSS ⁻¹ day ⁻¹ , respectively. In situ hybridization results revealed that the introduced denitrifying bacteria significantly facilitated the development of a dense denitrifying bacterial population in the activated sludge, which enhanced in situ denitrification activity. FISH data indicated that once introduced, B. denitrificans remained abundant throughout the experimental period. The ability to seed a bioreactor with bacterial strain capable of removing target pollutants from tannery effluents in a mixed microbial community suggests that this approach could have commercial applications.     

Use of Stable-Isotope Probing, Full-Cycle rRNA Analysis, and Fluorescence In Situ Hybridization-Microautoradiography To Study a Methanol-Fed Denitrifying Microbial Community

A denitrifying microbial consortium was enriched in an anoxically operated, methanol-fed sequencing batch reactor (SBR) fed with a mineral salts medium containing methanol as the sole carbon source and nitrate as the electron acceptor. The SBR was inoculated with sludge from a biological nutrient removal activated sludge plant exhibiting good denitrification. The SBR denitrification rate improved from less than 0.02 mg of NO₃⁻-N mg of mixed-liquor volatile suspended solids (MLVSS)⁻¹ h⁻¹ to a steady-state value of 0.06 mg of NO₃⁻-N mg of MLVSS⁻¹ h⁻¹ over a 7-month operational period. At this time, the enriched microbial community was subjected to stable-isotope probing (SIP) with [¹³C]methanol to biomark the DNA of the denitrifiers. The extracted [¹³C]DNA and [¹²C]DNA from the SIP experiment were separately subjected to full-cycle rRNA analysis. The dominant 16S rRNA gene phylotype (group A clones) in the [¹³C]DNA clone library was closely related to those of the obligate methylotrophs Methylobacillus and Methylophilus in the order Methylophilales of the Betaproteobacteria (96 to 97% sequence identities), while the most abundant clone groups in the [¹²C]DNA clone library mostly belonged to the family Saprospiraceae in the Bacteroidetes phylum. Oligonucleotide probes for use in fluorescence in situ hybridization (FISH) were designed to specifically target the group A clones and Methylophilales (probes DEN67 and MET1216, respectively) and the Saprospiraceae clones (probe SAP553). Application of these probes to the SBR biomass over the enrichment period demonstrated a strong correlation between the level of SBR denitrification and relative abundance of DEN67-targeted bacteria in the SBR community. By contrast, there was no correlation between the denitrification rate and the relative abundances of the well-known denitrifying genera Hyphomicrobium and Paracoccus or the Saprospiraceae clones visualized by FISH in the SBR biomass. FISH combined with microautoradiography independently confirmed that the DEN67-targeted cells were the dominant bacterial group capable of anoxic [¹⁴C]methanol uptake in the enriched biomass. The well-known denitrification lag period in the methanol-fed SBR was shown to coincide with a lag phase in growth of the DEN67-targeted denitrifying population. We conclude that Methylophilales bacteria are the dominant denitrifiers in our SBR system and likely are important denitrifiers in full-scale methanol-fed denitrifying sludges.     

Denitrification kinetics of simulated fish processing wastewater at different ratios of nitrate to biomass

Denitrification was studied in anoxic batch cultures of a simulated fish processing wastewater at 37 r C and pH 7.5, using a denitrifying enrichment culture from fishery wastewater. Different initial nitrate to biomass ratios (So/Xo) were used: nitrate and biomass varied from 7.5 to 94.7 mg NO₃-N l^–1, and from 20 to 4300 mg volatile suspended solids l^–1, respectively. The specific maximum denitrification rate (r _m) and the cell yield (Y _{X / S}) depended on the So/Xo ratio under anoxic conditions: r _m increased from 1.2 to 1584 mg NO₃-N g^–1 VSS h^–1 and Y _{X / S} decreased from 42 to 0.03 mg VSS mg^–1 NO₃-N when So/Xo varied from 5.5 10^{– 3} to 9.3 mg NO₃-N/mg VSS. Nomenclature C_{NO3 – N} nitrate concentration, mg NO₃-N l^–1 K _S saturation constant, mg NO₃-N l^–1 r _m specific maximum denitrification rate, mg NO₃-N g^–1 VSS h^–1 So initial substrate concentration, mg l^–1 t time, h TOC total organic carbon VSS volatile suspended solids x biomass concentration, g VSS l^–1 Xo initial biomass concentration, g VSS l^–1 Y _X/S substrate to biomass cell yield, mg VSS/mg N Greek symbols: _m maximum specific growth rate of the anoxic microbial population, 1 h^–1     

Spatial and Temporal Variability in Sediment Denitrification Within an Agriculturally Influenced Reservoir

Reservoirs are intrinsically linked to the rivers that feed them, creating a river–reservoir continuum in which water and sediment inputs are a function of the surrounding watershed land use. We examined the spatial and temporal variability of sediment denitrification rates by sampling longitudinally along an agriculturally influenced river–reservoir continuum monthly for 13 months. Sediment denitrification rates ranged from 0 to 63 μg N₂O g ash free dry mass of sediments (AFDM)⁻¹ h⁻¹ or 0–2.7 μg N₂O g dry mass of sediments (DM)⁻¹ h⁻¹ at reservoir sites, vs. 0–12 μg N₂O gAFDM⁻¹ h⁻¹ or 0–0.27 μg N₂O gDM⁻¹ h⁻¹ at riverine sites. Temporally, highest denitrification activity traveled through the reservoir from upper reservoir sites to the dam, following the load of high nitrate (NO₃⁻-N) water associated with spring runoff. Annual mean sediment denitrification rates at different reservoir sites were consistently higher than at riverine sites, yet significant relationships among theses sites differed when denitrification rates were expressed per gDM vs. per gAFDM. There was a significant positive relationship between sediment denitrification rates and NO₃⁻-N concentration up to a threshold of 0.88 mg NO₃⁻ -N l⁻¹, above which it appeared NO₃⁻-N was no longer limiting. Denitrification assays were amended seasonally with NO₃⁻-N and an organic carbon source (glucose) to determine nutrient limitation of sediment denitrification. While organic carbon never limited sediment denitrification, all sites were significantly limited by NO₃⁻-N during fall and winter when ambient NO ₃⁻-N was low.     

Physiology and kinetics of autotrophic denitrification by Thiobacillus denitrificans

Summary A strain of Thiobacillus denitrificans was isolated after enrichment under anaerobic conditions by the continuous culture technique using thiosulfate as energy source and nitrate as electron acceptor and nitrogen source. The isolate was an active denitrifyer, the optimal conditions being 30°C and pH 7.5–8.0. Denitrification was inhibited by sulfate (the reaction product) above 5 g SO ₄ ⁼ /l, whereas high concentrations of the substrates nitrate and thiosulfate were less harmful; nitrite affected denitrification above 0.2 g NO ₂ ^– /l. During the time course of denitrification in a batch culture growth and substrate consumption slowed down already after only half the substrate was utilized due to product inhibition. The following parameters were determined in continuous culture under nitrate limitation: _max=0.11 h^–1, K _S=0.2 mg NO ₃ ^– /l, maximum denitrification rate=0.78 g NO ₃ ^– /g cells·h, g cells/g NO ₃ ^– , g cells/g S₂O ₃ ⁼ . Nitrite did not accumulate during steady state denitrification; the denitrification gas was almost pure N₂. The concentrations of N₂O and NO were below 1 ppm.     

Effects of combining biological treatment and activated carbon on hexavalent chromium reduction

The objectives of the present work were: (a) to analyze the Cr(VI) removal by combining activated sludge (AS) with powdered activated carbon (PAC), (b) to analyze the effect of PAC and Cr(VI) on the growth kinetics of activated sludge, and (c) to determine if the combined method (AS-PAC) for Cr(VI) removal can be considered additive or synergistic with respect to the individual processes. Chromate removal was improved by increasing PAC concentrations in both PAC and AS-PAC systems. Cr(VI) removal using the AS-PAC system was higher than using AS or PAC. The increase of Cr(VI) caused longer lag phase and lower observed specific growth rate (μ_obs), biomass yield (Y_X/S), and specific growth substrate consumption rate (q_S) of activated sludge; additionally, PAC did not enhance the growth kinetic parameters (μ_obs, Y_X/S, q_S). Cr(VI) reduction in AS-PAC system was the result of the additive effect of each individual Cr(VI) removal process.     

Pyridine degradation characteristics of a newly isolated bacterial strain and its application with a novel reactor for the further treatment in pyridine wastewater

A pyridine-degrading strain Gemmobacter sp. ZP-12, isolated from an activated sludge, was able to use pyridine as the sole carbon and nitrogen source for the growth. The strain could effectively degrade pyridine and remove TOC over a wide range of initial pyridine concentrations. The pyridine degradation rate for 100, 500, 1000, 1500 and 2000 mg/L was 2.90 ± 0.17; 13.72 ± 0.21, 20.40 ± 0.24, 31.09 ± 0.26, 27.63 ± 0.17 mg/L/h, respectively. During the pyridine degraded, a large amount of NH₄⁺-N was released and accumulated. The accumulation of NH₄⁺-N increased with the increase of pyridine concentration. For further removing the NH₄⁺-N producing in pyridine degradation, an aerobic-moving bed biofilm reactor coupled with intermittent-aeration membrane biological reactor (a-MBBR-IMBR) was constructed, in which the strain and the aerobic / anoxic mixed sludge combined to remove the pollutants in the wastewater containing 500 mg/L pyridine. After 96 h of operation, the final TOC removal efficiency was 96.5 ± 1.05 %. The average residual concentration of NO₃⁻-N and NH₄⁺-N was respectively 9.09 ± 4.13 mg/L and 7.85 ± 3.88 mg/L. The study provides a viable option for treating pyridine wastewater.     

Denitrifying sulfide removal and carbon methanogenesis in a mesophilic, methanogenic culture

The inhibitory effects of 90-189 mg l⁻¹ of sulfide and 25-75 mg-N l⁻¹ of nitrate on methanogenesis were investigated in a mixed methanogenic culture using butyrate as carbon source. In the initial phase of 90 mg l⁻¹ S²⁻ test, autotrophic denitrification of nitrate occurred with sulfide as the electron donor. Then the sulfate-reducing strains converted the produced sulfur back to sulfide via heterotrophic oxidation pathway. Methanogenesis was not markedly inhibited when 90 mg l⁻¹ of sulfide was dosed alone. When 25-75 mg-N l⁻¹ of nitrate was presented, initiation of methanogenesis was seriously delayed. Nitrogen oxides (NO_x), the intermediates for nitrate reduction via denitrification pathway, inhibited methanogenesis. The 90 mg l⁻¹ of sulfide favored heterotrophic dissimilatory nitrate reduction to ammonia (DNRA) pathway for nitrate reduction. Possible ways of maximizing methane production from an organic carbon-rich wastewater with high levels of sulfide and nitrate were discussed.