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In this study, we analysed the nitrifying microbial community (ammonium-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB)) within three different aerobic granular sludge treatment systems as well as within one flocculent sludge system. Granular samples were taken from one pilot plant run on municipal wastewater as well as from two lab-scale reactors. Fluorescent in situ hybridization (FISH) and quantitative PCR (qPCR) showed that Nitrobacter was the dominant NOB in acetate-fed aerobic granules. In the conventional system, both Nitrospira and Nitrobacter were present in similar amounts. Remarkably, the NOB/AOB ratio in aerobic granular sludge was elevated but not in the conventional treatment plant suggesting that the growth of Nitrobacter within aerobic granular sludge, in particular, was partly uncoupled from the lithotrophic nitrite supply from AOB. This was supported by activity measurements which showed an approximately threefold higher nitrite oxidizing capacity than ammonium oxidizing capacity. Based on these findings, two hypotheses were considered: either Nitrobacter grew mixotrophically by acetate-dependent dissimilatory nitrate reduction (ping-pong effect) or a nitrite oxidation/nitrate reduction loop (nitrite loop) occurred in which denitrifiers reduced nitrate to nitrite supplying additional nitrite for the NOB apart from the AOB.

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The long-and short-term effects of salt on biological nitrogen and phosphorus removal processes were studied in an aerobic granular sludge reactor. The microbial community structure was investigated by PCR-denaturing gradient gel electrophoresis (DGGE) on 16S rRNA and amoA genes. PCR products obtained from genomic DNA and from rRNA after reverse transcription were compared to determine the presence of bacteria as well as the metabolically active fraction of bacteria. Fluorescence in situ hybridization (FISH) was used to validate the PCR-based results and to quantify the dominant bacterial populations. The results demonstrated that ammonium removal efficiency was not affected by salt concentrations up to 33 g/liter NaCl. Conversely, a high accumulation of nitrite was observed above 22 g/liter NaCl, which coincided with the disappearance of Nitrospira sp. Phosphorus removal was severely affected by gradual salt increase. No P release or uptake was observed at steady-state operation at 33 g/liter NaCl, exactly when the polyphosphate-accumulating organisms (PAOs), "Candidatus Accumulibacter phosphatis" bacteria, were no longer detected by PCR-DGGE or FISH. Batch experiments confirmed that P removal still could occur at 30 g/liter NaCl, but the long exposure of the biomass to this salinity level was detrimental for PAOs, which were outcompeted by glycogen-accumulating organisms (GAOs) in the bioreactor. GAOs became the dominant microorganisms at increasing salt concentrations, especially at 33 g/liter NaCl. In the comparative analysis of the diversity (DNA-derived pattern) and the activity (cDNA-derived pattern) of the microbial population, the highly metabolically active microorganisms were observed to be those related to ammonia (Nitrosomonas sp.) and phosphate removal ("Candidatus Accumulibacter").Numerous wastewaters, like those generated in seafood canning, pickling, and cheese-processing industries, can contain significant amounts of inorganic dissolved salts (14). The use of seawater for toilet flushing in ships, offshore installations, and regions with problems related to water supply also introduces salts into freshwater. Wastewaters with a high salt content require the activity of salt-tolerant microorganisms which may not be present in high numbers in microbial inocula from sewage treatment plants. Salt ions exert high osmotic pressure on the microorganisms, and most of the freshwater-based microbial populations are unable to survive at these high osmotic pressures and either die or become dormant under these conditions (40). Besides that, inorganic salts can affect the structure and settling properties of microbial flocs as well as the maximum solubility of oxygen and its transfer to the liquid phase, which can lead to oxygen limitation. Consequently, high salt concentrations have a negative influence on existing biological wastewater treatment plants, affecting organic matter, nitrogen, and phosphorus removal (17). So far, most studies on chemical oxygen demand (COD), N, and P removal from saline wastewa...

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Salinity can adversely affect the performance of most biological processes involved in wastewater treatment. The effect of salt on the main conversion processes in an aerobic granular sludge (AGS) process accomplishing simultaneous organic matter, nitrogen, and phosphate removal was evaluated in this work. Hereto, an AGS sequencing batch reactor was subjected to different salt concentrations (0.2 to 20 g Cl(-) l(-1)). Granular structure was stable throughout the whole experimental period, although granule size decreased and a significant effluent turbidity was observed at the highest salinity tested. A weaker gel structure at higher salt concentrations was hypothesised to be the cause of such turbidity. Ammonium oxidation was not affected at any of the salt concentrations applied. However, nitrite oxidation was severely affected, especially at 20 g Cl(-) l(-1), in which a complete inhibition was observed. Consequently, high nitrite accumulation occurred. Phosphate removal was also found to be inhibited at the highest salt concentration tested. Complementary experiments have shown that a cascade inhibition effect took place: first, the deterioration of nitrite oxidation resulted in high nitrite concentrations and this in turn resulted in a detrimental effect to polyphosphate-accumulating organisms. By preventing the occurrence of the nitrification process and therefore avoiding the nitrite accumulation, the effect of salt concentrations on the bio-P removal process was shown to be negligible up to 13 g Cl(-) l(-1). Salt concentrations equal to 20 g Cl(-) l(-1) or higher in absence of nitrite also significantly reduced phosphate removal efficiency in the system.

In this study, the effect of different operational conditions on biofilm development and nitrification in three moving-bed biofilm reactors (MBBRs) was investigated: two reactors were operated in a continuously fed regime and one in sequencing-batch mode. The presence of organic carbon reduced the time required to form stable nitrifying biofilms. Subsequent stepwise reduction of influent COD caused a decrease in total polysaccharide and protein content, which was accompanied by a fragmentation of the biofilm, as shown by scanning electron microscopy, and by an enrichment of the biofilm for nitrifiers, as observed by fluorescent in situ hybridization (FISH) analysis. Polysaccharide and protein concentrations proved to be good indicators of biomass development and detachment in MBBR systems. Ammonium- and nitrite-oxidizing bacteria activities were affected when a pulse feeding of 4 g of NH(4)-N/(m(2)·day) was applied. Free nitrous acid and free ammonia were likely the inhibitors for ammonium- and nitrite-oxidizing bacteria.

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