Identification of a Novel Group of Bacteria in Sludge from a Deteriorated Biological Phosphorus Removal Reactor

1999 ◽  
Vol 65 (3) ◽  
pp. 1251-1258 ◽  
Author(s):  
Alex T. Nielsen ◽  
Wen-Tso Liu ◽  
Carlos Filipe ◽  
Leslie Grady ◽  
Søren Molin ◽  
...  
Keyword(s):  
Phosphorus Removal ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Weight Ratio ◽  
Microscopic Analysis ◽  
The Novel ◽  
Biological Phosphorus Removal ◽  
Close Relationship ◽  
Gradient Gel Electrophoresis ◽  
Biological Phosphorus

ABSTRACT The microbial diversity of a deteriorated biological phosphorus removal reactor was investigated by methods not requiring direct cultivation. The reactor was fed with media containing acetate and high levels of phosphate (P/C weight ratio, 8:100) but failed to completely remove phosphate in the effluent and showed very limited biological phosphorus removal activity. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA was used to investigate the bacterial diversity. Up to 11 DGGE bands representing at least 11 different sequence types were observed; DNA from the 6 most dominant of these bands was further isolated and sequenced. Comparative phylogenetic analysis of the partial 16S rRNA sequences suggested that one sequence type was affiliated with the alpha subclass of theProteobacteria, one was associated with theLegionella group of the gamma subclass of theProteobacteria, and the remaining four formed a novel group of the gamma subclass of the Proteobacteria with no close relationship to any previously described species. The novel group represented approximately 75% of the PCR-amplified DNA, based on the DGGE band intensities. Two oligonucleotide rRNA probes for this novel group were designed and used in a whole-cell hybridization analysis to investigate the abundance of this novel group in situ. The bacteria were coccoid and 3 to 4 μm in diameter and represented approximately 35% of the total population, suggesting a relatively close agreement with the results obtained by the PCR-based DGGE method. Further, based on electron microscopy and standard staining microscopic analysis, this novel group was able to accumulate granule inclusions, possibly consisting of polyhydroxyalkanoate, inside the cells.

scholarly journals Performance of enhanced biological phosphorus removal and population dynamics of phosphorus accumulating organisms in sludge-shifting sequencing batch reactors

2018 ◽  
Vol 78 (4) ◽  
pp. 886-895 ◽  
Author(s):  
Yang Pan ◽  
Wenquan Ruan ◽  
Yong Huang ◽  
Qianqian Chen ◽  
Hengfeng Miao ◽  
...  
Keyword(s):  
Removal Efficiency ◽  
Phosphorus Removal ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Batch Reactor ◽  
Batch Reactors ◽  
Enhanced Biological Phosphorus Removal ◽  
Biological Phosphorus Removal ◽  
High Concentration ◽  
Gradient Gel Electrophoresis ◽  
Biological Phosphorus

Abstract The sludge-shifting sequencing batch reactor (SBR) is an enhanced biological phosphorus removal (EBPR) process for wastewater treatment. In this study, the enrichment of phosphorus accumulating organisms (PAOs) will be attempted by using different high concentration of substrates. In sludge-shifting SBR, activated sludge can be continuously shifted from the bottom of SBR to anaerobic zone/selector, which contains high concentration of substrates, through an orderly reflux between the paralleled SBRs. Denaturing gradient gel electrophoresis (DGGE) methods were used to monitor microbial diversity in sludge. Fluorescence in situ hybridization (FISH) was used to determine the microbial population profile and distribution map under different sludge shifting volumes. The synthesis of intracellular polymers in this process was also analyzed. Phosphorus removal efficiency as high as 96% ± 1.3% was achieved under a sludge shifting ratio of 30%. Synthetic efficiencies of polyhydroxybutyrate (PHB) by PAOs were improved at high sludge shifting ratios. FISH results demonstrated that the population of PAOs in the process increased under properly sludge shifting ratio and it significantly improved phosphorus removal efficiency. Sequencing results indicated that determined sequences (11 OTUs) belonged to Proteobacterium, Actinobacteria and Firmicutes, Pseudomonas kuykendallii, which played an important role in the process of P removal.

Role of “G-bacteria” in anaerobic substrate uptake in a SBR with no phosphorus removal

2002 ◽  
Vol 46 (1-2) ◽  
pp. 171-178 ◽  
Author(s):  
Y.H. Kong ◽  
M. Beer ◽  
R.J. Seviour ◽  
K.C. Lindrea ◽  
G.A. Rees
Keyword(s):  
16S Rdna ◽  
Phosphorus Removal ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Phylogenetic Group ◽  
Substrate Uptake ◽  
Biological Phosphorus Removal ◽  
Gram Positive Bacteria ◽  
Gradient Gel Electrophoresis ◽  
Biological Phosphorus

Biomass from an SBR running with no enhanced biological phosphorus removal (EBPR) but which exhibited anaerobic assimilation of glucose and acetate, was dominated by “G-bacteria”, cocci in tetrads and clusters. Extracted 16S rDNA was amplified by PCR and then analysed using Denaturing Gradient Gel Electrophoresis (DGGE). Major bands were extracted and their sequences determined. Clone libraries were also prepared, the 16S rDNA extracted, PCR performed and the resultant fragments run by DGGE to aid in identifying the DGGE bands and provide fuller sequences than available by DGGE alone. The two approaches together allowed several bands to be identified. Probes for FISH analyses were designed for some of these in attempts to see to which phylogenetic group “G-bacteria” belonged, and whether they represented the dominant bands detected by DGGE. Then FISH/Microautoradiography (MAR) was used in attempts to see which bacteria there were assimilating substrates anaerobically. Results indicated that the “G-bacteria” were phylogenetically diverse, but mainly α-proteobacteria and members of the high G+C% Gram-positive bacteria. Not all of these could assimilate glucose and/or acetate anaerobically, and Amaricoccus, the original “G-bacteria” of Cech and Hartman, was not detected.

Applicability of experience from laboratory reactors with biological phosphorus removal in full-scale plants

2006 ◽  
Vol 54 (1) ◽  
pp. 267-275 ◽  
Author(s):  
E. Tykesson ◽  
L.L. Blackall ◽  
Y. Kong ◽  
P.H. Nielsen ◽  
J. la Cour Jansen
Keyword(s):  
Phosphorus Removal ◽  
Wastewater Treatment Plants ◽  
In Situ Hybridisation ◽  
Full Scale ◽  
Fluorescence In Situ Hybridisation ◽  
Biological Phosphorus Removal ◽  
Batch Tests ◽  
P Release ◽  
Biological Phosphorus

Enhanced biological phosphorus removal (EBPR) has been used at many wastewater treatment plants all over the world for many years. In this study a full-scale sludge with good EBPR was tested with P-release batch tests and combined FISH/MAR (fluorescence in situ hybridisation and microautoradiography). Proposed models of PAOs and GAOs (polyphosphate- and glycogen-accumulating organisms) and microbial methods suggested from studies of laboratory reactors were found to be applicable also on sludge from full-scale plants. Dependency of pH and the uptake of both acetate and propionate were studied and used for calculations for verifying the models and results from microbial methods. All rates found from the batch tests with acetate were higher than in the batch tests with propionate, which was explained by the finding that only those parts of the bacterial community that were able to take up acetate anaerobically were able to take up propionate anaerobically.

Morphology, In-Situ characterization with rRNA targetted probes and respiratory quinone profiles of enhanced biological phosphorus removal sludge

1998 ◽  
Vol 38 (8-9) ◽  
pp. 69-76 ◽  
Author(s):  
I. M. Sudiana ◽  
T. Mino ◽  
H. Satoh ◽  
T. Matsuo
Keyword(s):  
Carbon Source ◽  
Activated Sludge ◽  
Microbial Communities ◽  
Phosphorus Removal ◽  
Enhanced Biological Phosphorus Removal ◽  
Biological Phosphorus Removal ◽  
In Situ Characterization ◽  
Respiratory Quinone ◽  
Biological Phosphorus

The microbial communities in activated sludge acclimated with either acetate or glucose as the major carbon source under phosphorus limited or rich conditions were investigated morphologically, phylogenetically and chemotaxonomically. The sludge with a minimized polyphosphate content was dominated by tetrad shaped bacteria, which were suspected to be ‘glycogen accumulating bacteria (GAOs) or G bacteria’ The sludge containing high polyphosphate was dominated by cluster forming coccus bacteria. Quinone analyses suggested that all the sludge tested contained various ubiquinones and menaquinones, of which the ubiquinones Q-8 and Q-10 were dominant. Analyses with rRNA targeted probes showed that beta sub class of Proteobacteria was most predominant in all sludges tested. Morphological, phylogenetic and chemotaxonomic investigation all indicated that both high and low P sludges are microbiologically diverse.

Towards exposure of elusive metabolic mixed-culture processes: the application of metaproteomic analyses to activated sludge

2006 ◽  
Vol 54 (1) ◽  
pp. 217-226 ◽  
Author(s):  
P. Wilmes ◽  
P.L. Bond
Keyword(s):  
Protein Expression ◽  
Activated Sludge ◽  
Phosphorus Removal ◽  
Batch Reactor ◽  
Biological Phosphorus Removal ◽  
Microbial Community Structures ◽  
Proteomic Investigation ◽  
Two Phases ◽  
Biological Phosphorus

Protein expression is a direct reflection of specific microbial activities in any ecosystem. In order to assess protein expression in mixed microbial communities, the feasibility of applying proteomic techniques to activated sludge samples has recently been demonstrated. We report the application of metaproteomics to two activated sludges from a laboratory-scale sequencing batch reactor with dissimilar phosphorus removal performances. Fluorescence in situ hybridization (FISH) revealed that the sludge with good enhanced biological phosphorus removal performance (EBPR) was dominated by Betaproteobacteria (65% of EUBMIX binding cells) and gave positive signals for the Rhodocyclus-type PAO specific probe (59%). The non-EBPR sludge was dominated by tetrad-forming Alphaproteobacteria (75%). With regard to the proteomic investigation, 630 individual protein spots were matched across the replicate groups of the anaerobic and aerobic phases of the EBPR sludge with 9.4% of all spots being statistically different between the two phases. The non-EBPR metaproteomic maps exhibited 590 matched spots with 14.7% statistical differences between the two phases. Overall, the non-EBPR sludge expressed around 30% more significant differences than the EBPR sludge. The comparison of protein expression in the two sludges showed that their metaproteomes were substantially different and this was reflected in their microbial community structures and metabolic transformations.

Presence of Rhodocyclus in a full-scale wastewater treatment plant and their participation in enhanced biological phosphorus removal

2002 ◽  
Vol 46 (1-2) ◽  
pp. 123-128 ◽  
Author(s):  
J.L. Zilles ◽  
C.-H. Hung ◽  
D.R. Noguera
Keyword(s):  
Wastewater Treatment ◽  
Phosphorus Removal ◽  
Fluorescent In Situ Hybridization ◽  
Wastewater Treatment Plants ◽  
Full Scale ◽  
Enhanced Biological Phosphorus Removal ◽  
Biological Phosphorus Removal ◽  
Polyphosphate Accumulating Organisms ◽  
Biological Phosphorus

The objective of this research was to assess the relevance of organisms related to Rhodocyclus in enhanced biological phosphorus removal in full-scale wastewater treatment plants. The presence of these organisms in full-scale plants was first confirmed by fluorescent in situ hybridization. To address which organisms were involved in phosphorus removal, a method was developed which selected polyphosphate-accumulating organisms from activated sludge samples by DAPI staining and flow cytometry. Sorted samples were characterized using fluorescent in situ hybridization. The results of these analyses confirmed the presence of organisms related to Rhodocyclus in full-scale wastewater treatment plants and supported the involvement of these organisms in enhanced biological phosphorus removal. However, a significant fraction of the polyphosphate-accumulating organisms were not related to Rhodocyclus.

The effect of GAOs (glycogen accumulating organisms) on anaerobic carbon requirements in full-scale Australian EBPR (enhanced biological phosphorus removal) plants

2003 ◽  
Vol 47 (11) ◽  
pp. 37-43 ◽  
Author(s):  
A.M. Saunders ◽  
A. Oehmen ◽  
L.L. Blackall ◽  
Z. Yuan ◽  
J. Keller
Keyword(s):  
Phosphorus Removal ◽  
In Situ Hybridisation ◽  
Full Scale ◽  
Fluorescence In Situ Hybridisation ◽  
Enhanced Biological Phosphorus Removal ◽  
Biological Phosphorus Removal ◽  
Accumulibacter Phosphatis ◽  
Polyphosphate Accumulating Organisms ◽  
Biological Phosphorus

Glycogen-accumulating organisms (GAOs) were present in six full-scale plants investigated and in all but one made a significant contribution to the amount of volatile fatty acid (VFA) taken up anaerobically. While most plants surveyed contain GAOs, it was demonstrated that it is possible for a full-scale plant to operate with an insignificant GAO population.“Candidatus Accumulibacter phosphatis”were the significant polyphosphate-accumulating organisms (PAOs) in all plants surveyed. “Candidatus Competibacter phosphatis” were found in all plants along with other possible GAOs that were observed but not identified. A significant GAO population will increase the carbon requirements by removing VFA that could otherwise have been used by PAOs. Process optimization minimizing GAOs in full-scale plants would lead to a more efficient use of VFA. Enhanced biological phosphorus removal (EBPR), fluorescence in situ hybridisation (FISH), glycogen accumulating organism (GAO); polyphosphate accumulating organism (PAO);

Microbial succession of glycogen accumulating organisms in an anaerobic-aerobic membrane bioreactor with no phosphorus removal

2006 ◽  
Vol 54 (1) ◽  
pp. 29-37 ◽  
Author(s):  
M.-T. Wong ◽  
W.-T. Liu
Keyword(s):  
Phosphorus Removal ◽  
Membrane Bioreactor ◽  
Fragment Length Polymorphism ◽  
Operating Conditions ◽  
Biological Phosphorus Removal ◽  
Microbial Succession ◽  
Solid Residence Time ◽  
Biological Phosphorus ◽  
Quantitative Fluorescence

The succession of glycogen accumulating organisms (GAOs) has been observed in an acetate-fed, anaerobic-aerobic sequencing membrane bioreactor (MBR) operated for 260 days without enhanced biological phosphorus removal (EBPR) activity. Semi-quantitative fluorescence in situ hybridization results showed that a gammaproteobacterial lineage GB frequently observed in EBPR processes was initially the numerically dominant species (50–66% of total cells) of the GAO in the MBR from day 1 to day 38. During this period, succession of two different subgroups of group GB was also observed. On day 85 onward, a population shift from GB group to ‘Defluvicoccus’-related tetrad-forming organisms (TFO) occurred. This microbial succession was suspected to be related to the applied operating conditions (long hydraulic retention time and long solid residence time) which favored the proliferation of ‘Defluvicoccus’-related TFO rather than the GB group. Application of terminal restriction fragment length polymorphism on selected samples further revealed that the microbial diversity of the seeding sludge as determined by the number of terminal restriction fragments was higher than that of sludge samples taken after day 85.

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