Methanol‐driven enhanced biological phosphorus removal with a syntrophic consortium

The presence of suitable carbon sources for enhanced biological phosphorus removal (EBPR) plays a key role in phosphorus removal from wastewater in urban WWTP. For wastewaters with low volatile fatty acids (VFAs) content, an external carbon addition is necessary. As methanol is the most commonly external carbon source used for denitrification it could be a priori a promising alternative, but previous attempts to use it for EBPR have failed. This study is the first successful report of methanol utilization as external carbon source for EBPR. Since a direct replacement strategy (i.e., supply of methanol as a sole carbon source to a propionic‐fed PAO‐enriched sludge) failed, a novel process was designed and implemented successfully: development of a consortium with anaerobic biomass and polyphosphate accumulating organisms (PAOs). Methanol‐degrading acetogens were (i) selected against other anaerobic methanol degraders from an anaerobic sludge; (ii) subjected to conventional EBPR conditions (anaerobic + aerobic); and (iii) bioaugmented with PAOs. EBPR with methanol as a sole carbon source was sustained in a mid‐term basis with this procedure. Biotechnol. Bioeng. 2013; 110: 391–400. © 2012 Wiley Periodicals, Inc.     

Identification and comparison of aerobic and denitrifying polyphosphate-accumulating organisms

Two laboratory-scale sequencing batch reactors (SBRs) were operated for enhanced biological phosphorus removal (EBPR) in alternating anaerobic-aerobic or alternating anaerobic-anoxic modes, respectively. Polyphosphate-accumulating organisms (PAOs) were enriched in the anaerobic-aerobic SBR and denitrifying PAOs (DPAOs) were enriched in the anaerobic-aerobic SBR. Fluorescence in situ hybridization (FISH) demonstrated that the well-known PAO, "Candidatus Accumulibacter phosphatis" was abundant in both SBRs, and post-FISH chemical staining with 4,6-diamidino-2-phenylindol (DAPI) confirmed that they accumulated polyphosphate. When the anaerobic-anoxic SBR enriched for DPAOs was converted to anaerobic-aerobic operation, aerobic uptake of phosphorus by the resident microbial community occurred immediately. However, when the anaerobic-aerobic SBR enriched for PAOs was exposed to one cycle with anoxic rather than aerobic conditions, a 5-h lag period elapsed before phosphorus uptake proceeded. This anoxic phosphorus-uptake lag phase was not observed in the subsequent anaerobic-aerobic cycle. These results demonstrate that the PAOs that dominated the anaerobic-aerobic SBR biomass were the same organisms as the DPAOs enriched under anaerobic-anoxic conditions.     

Effect of nitrite from nitritation on biological phosphorus removal in a sequencing batch reactor treating domestic wastewater

Although nitrite effect on enhanced biological phosphorus removal (EBPR) has been previously studied, very limited research has been undertaken about the effect of nitrite accumulation caused by nitritation on EBPR. This paper focused on nitrite effect from nitritation on EBPR in a sequencing batch reactor treating domestic wastewater. Results showed that nitrite of below 10 mg/L did not inhibit P-uptake and release; whereas EBPR deterioration was observed when nitrite accumulation reached 20 mg/L. Due to P-uptake prior to nitritation, nitrite of 20 mg/L has no effect on aerobic P-uptake. The main reason leading to EBPR deterioration was the competition of carbon source. Batch tests were conducted to investigate nitrite effect on anaerobic P-release. Under sufficient carbon source, nitrite of 30 mg/L had no impact on poly-β-hydroxyalkanoate (PHA) storage; contrarily, under insufficient carbon source, denitrifiers competing for carbon source with phosphorus accumulating organisms resulted in decrease of PHA synthesis and P-release.     

Effect of pH change on the performance and microbial community of enhanced biological phosphate removal process

An acetate-rich wastewater, containing 170 mg/L of total organic carbon (TOC), 13 mg/L of N, and 15 mg/L of P, was treated using the enhanced biological phosphate removal (EBPR) process operated in a sequencing batch reactor. A slight change of pH of the mixed liquor from 7.0 to 6.5 led to a complete loss of phosphate-removing capability and a drastic change of microbial populations. The process steadily removed 94% of TOC and 99.9% of P from the wastewater at pH 7.0, but only 93% TOC and 17% of P 14 days after the pH was lowered to pH 6.5. The sludge contained 8.8% P at pH 7.0, but only 1.9% at pH 6.5. Based on 16S rDNA analysis, 64.8% of the clones obtained from the sludge at pH 7.0 were absent in the pH 6.5 sludge. The missing microbes, some of which were likely responsible for the phosphate removal at pH 7.0, included beta-Proteobacteria, Actinobacteria, Bacteriodetes/Chlorobi group, plus photosynthetic bacteria and Defluvicoccus of the alpha-Proteobacteria. Among them, the last two groups, which represented 9.3% and 10.1% of the EBPR sludge at pH 7.0, have rarely been reported in an EBPR system.     

Enhanced biological phosphorus removal in a sequencing batch reactor using propionate as the sole carbon source

An enhanced biological phosphorus removal (EBPR) system was developed in a sequencing batch reactor (SBR) using propionate as the sole carbon source. The microbial community was followed using fluorescence in situ hybridization (FISH) techniques and Candidatus 'Accumulibacter phosphatis' were quantified from the start up of the reactor until steady state. A series of SBR cycle studies was performed when 55% of the SBR biomass was Accumulibacter, a confirmed polyphosphate accumulating organism (PAO) and when Candidatus 'Competibacter phosphatis', a confirmed glycogen-accumulating organism (GAO), was essentially undetectable. These experiments evaluated two different carbon sources (propionate and acetate), and in every case, two different P-release rates were detected. The highest rate took place while there was volatile fatty acid (VFA) in the mixed liquor, and after the VFA was depleted a second P-release rate was observed. This second rate was very similar to the one detected in experiments performed without added VFA.A kinetic and stoichiometric model developed as a modification of Activated Sludge Model 2 (ASM2) including glycogen economy, was fitted to the experimental profiles. The validation and calibration of this model was carried out with the cycle study experiments performed using both VFAs. The effect of pH from 6.5 to 8.0 on anaerobic P-release and VFA-uptake and aerobic P-uptake was also studied using propionate. The optimal overall working pH was around 7.5. This is the first study of the microbial community involved in EBPR developed with propionate as a sole carbon source along with detailed process performance investigations of the propionate-utilizing PAOs.     

Verification of enhanced phosphate removal capability in pure cultures of<Emphasis Type="Italic">Acinetobacter calcoaceticus</Emphasis> under anaerobic/aerobic conditions in an SBR

Laboratory experiments were conducted using pure cultures ofAcinetobacter under anaerobic/aerobic cyclic conditions to explain the release and uptake of soluble phosphate in an activated sludge process showing enhanced biological phosphate removal (EBPR). Under anaerobic/aerobic cyclic conditions in a Sequencing Batch Reactor (SBR), COD uptake concurrent with soluble phosphate release byAcinetobacter was not significant during the anaerobic periods, indicating that EBPR would not be established in pure cultures. However,Acinetobacter cells accumulated higher phosphate content (5.2%) in SBR than that obtained (4.3%) from batch experiments. These results suggest thatAcinetobacter sp. may not follow the proposed pattern of behavior of poly-P bacteria in EBPR activated sludge plants.     

‘Candidatus Competibacter'-lineage genomes retrieved from metagenomes reveal functional metabolic diversity

The glycogen-accumulating organism (GAO) ‘Candidatus Competibacter'' (Competibacter) uses aerobically stored glycogen to enable anaerobic carbon uptake, which is subsequently stored as polyhydroxyalkanoates (PHAs). This biphasic metabolism is key for the Competibacter to survive under the cyclic anaerobic-‘feast'': aerobic-‘famine'' regime of enhanced biological phosphorus removal (EBPR) wastewater treatment systems. As they do not contribute to phosphorus (P) removal, but compete for resources with the polyphosphate-accumulating organisms (PAO), thought responsible for P removal, their proliferation theoretically reduces the EBPR capacity. In this study, two complete genomes from Competibacter were obtained from laboratory-scale enrichment reactors through metagenomics. Phylogenetic analysis identified the two genomes, ‘Candidatus Competibacter denitrificans'' and ‘Candidatus Contendobacter odensis'', as being affiliated with Competibacter-lineage subgroups 1 and 5, respectively. Both have genes for glycogen and PHA cycling and for the metabolism of volatile fatty acids. Marked differences were found in their potential for the Embden–Meyerhof–Parnas and Entner–Doudoroff glycolytic pathways, as well as for denitrification, nitrogen fixation, fermentation, trehalose synthesis and utilisation of glucose and lactate. Genetic comparison of P metabolism pathways with sequenced PAOs revealed the absence of the Pit phosphate transporter in the Competibacter-lineage genomes—identifying a key metabolic difference with the PAO physiology. These genomes are the first from any GAO organism and provide new insights into the complex interaction and niche competition between PAOs and GAOs in EBPR systems.     

强化生物除磷系统主要微生物及其代谢机理研究进展

强化生物除磷(enhanced biological phosphorus removal,EBPR)工艺在废水除磷处理中应用广泛.主要功能微生物及其代谢机理的研究是有效调控EBPR工艺稳定运行与效能提升的基础.本文选取EBPR系统中最主要的两类微生物(聚磷菌和聚糖菌),从底物吸收机制、糖酵解途径、TCA途径的贡献以及聚磷菌和聚糖菌的代谢相似性等方面对这些微生物的代谢机理进行综述,评价了分子生物学技术在研究EBPR系统微生物学及其代谢机理方面的应用现状,在此基础上对EBPR系统今后的研究方向进行了展望.
     

Temperature effects on the aerobic metabolism of glycogen-accumulating organisms

Short-term temperature effects on the aerobic metabolism of glycogen-accumulating organisms (GAO) were investigated within a temperature range from 10 to 40 degrees C. Candidatus Competibacter Phosphatis, known GAO, were the dominant microorganisms in the enriched culture comprising 93 +/- 1% of total bacterial population as indicated by fluorescence in situ hybridization (FISH) analysis. Between 10 and 30 degrees C, the aerobic stoichiometry of GAO was insensitive to temperature changes. Around 30 degrees C, the optimal temperature for most of the aerobic kinetic rates was found. At temperatures higher than 30 degrees C, a decrease on the aerobic stoichiometric yields combined with an increase on the aerobic maintenance requirements were observed. An optimal overall temperature for both anaerobic and aerobic metabolisms of GAO appears to be found around 30 degrees C. Furthermore, within a temperature range (10-30 degrees C) that covers the operating temperature range of most of domestic wastewater treatment systems, GAOs aerobic kinetic rates exhibited a medium degree of dependency on temperature (theta = 1.046-1.090) comparable to that of phosphorus accumulating organisms (PAO). We conclude that GAO do not have metabolic advantages over PAO concerning the effects of temperature on their aerobic metabolism, and competitive advantages are due to anaerobic processes.