<p>Waste Stabilisation Ponds (WSPs) are characterised by low phosphorus (P) removal capacity. Heterotrophic bacteria are principal microbial agents in WSPs in addition to algae. As treatment proceeds in WSPs, algal growth increases and pH rises, this has lead to believe that P removal is mainly through sedimentation as organic P algal biomass and precipitation as inorganic P. In activated sludge treatment plants (AS), microbial P removal has been improved and is termed as enhanced biological phosphorus removal. There was a need to establish whether it was possible to enhance P removal in WSPs. A performance assessment of pond system at the University of Dare s Salaam (UDSM), Tanzania, has shown that 90% of the P removed was in the primary pond (facultative) and the rest in the maturation pond (aerobic).</p><p>In these studies, a pure strain A. hydrophyla was isolated from an activated sludge wastewater treatment plant in Sweden. This plant has a train that functions with enhanced biological phosphorus removal. The strain was tested for P uptake in minimal media supplemented with glucose, succinate or acetate, grown aerobically and anaerobically/aerobically. This strain was able to take up P without having been subjected to the anaerobic phase. It was observed that P uptake was enhanced after the anaerobic phase with media supplemented with glucose, but not with succinate or acetate. Phosphorus uptake repeatedly followed the bacterial growth pattern with correlation coefficients of more than 95%. Therefore P removal has a direct correlation with bacterial growth.</p><p>Two isolates Acinetobacter sp. (isolated from the primary facultative pond) and E .coli (isolated from the maturation pond) were obtained from a tropical WSP treatment system at the UDSM. They were subjected to aerobic P uptake experiment similar to those of A.hydrophyla. The uptake per unit absorbance of bacterial growth was found to be comparable to that of A.hydrophyla, isolated from AS. These results showed that heterotrophic activity is important in WSPs. It is possible to enhance P removal in these systems by designing the primary ponds for maximum heterotrophic activity and probably enrichment.</p>

<p>Analytical analyses conducted at the Himmerfjärden WWTP (285.000 PE connected) identified 70 pharmaceutical compounds belonging to different therapeutic classes. Such organic micropollutants at low detected concentration range of µg - ng l^-1 did not affect the treatment processes at WWTP. Results from analytical studies indicated continuous discharge of organic micropollutants to the surface water with a calculated load amounting to 1.51 kg day-1. Metoprolol, carbamazepine and naproxen were chosen for testing different removal methods. Oxygen Uptake Rate (OUR) tests were conducted to assess the bacterial activity of an activated sludge taken from a full scale aeration plant with the presence of selected target compounds.</p><p>A semi-technical scale membrane bioreactor ZeeWeed10™, treating final effluent from the Himmerfjärden WWTP (Sweden) was seeded with activated sludge from full scale biological stage. The membrane bioreactor (MBR) system placed after the final treatment appeared to be an insufficient technology for removal of residual amounts of organic micropollutants from WWTP effluents. Batch test studies with activated sludge taken from the membrane bioreactor and with application of granular activated carbon (GAC) filtration resulted in giving an overall assessment of removal efficiency. Metoprolol and carbamazepine tend to be resistant to the biodegradation process and in the dosed high concentration lead to bacterial cell decomposition in the activated sludge. Apparently, removal efficiency for naproxen exceeded the value of 46% with the spiked initial amount of 3.3 mg NAP g^-1MLSS. Application of the GAC filtration proved to be an efficient technique for removal of pharmaceutical compounds from treated wastewater.</p><p>Application of the statistical programme Modde7 was a time saving tool in studies of fouling occurrence. The effect of fouling phenomenon, which is a highly limiting factor for MBR performance, was minimised by adjusting the operational parameters as predicted by the Modde7 programme.</p>

With increasing water reuse applications and upcoming stringent regulations for treated wastewater effluent discharge, wastewater plants need to consider alternative technologies beyond conventional treatment processes. The new regulations, Numeric Nutrient Criteria (NNC), may regulate discharge nitrogen and phosphorus concentrations to as low as 0.5 mg/L as N and 10 μg/L as P respectively. To meet these target requirements, system retrofitting to incorporate chemical or advanced nutrient removal systems possibly with membrane technology will most likely be required. Although microfiltration/ultrafiltration membranes coupled with biological processes, otherwise known as membrane bioreactors (MBR), remove contaminants and suspended solids, nutrient removal is minimal to none. This emphasizes the importance of the biological process in MBRs. This study evaluated and tested the improvement of biological nutrient removal (BNR) in an MBR system which can meet NNC regulations along with the optimization of membrane operation for the reduction of fouling and energy consumption. A pilot study was conducted at the City of Tampa wastewater treatment plant and was divided into four phases of experimentation using two submerged MBR membranes operated with modified biological configurations. Laboratory analyses and data collection were conducted during the experiments and the performance evaluated for each configuration. System configurations were also optimized throughout each phase of testing for nutrient removal. Important factors used in the development of an appropriate configuration included isolation of the membrane tank from the biological reactors in the design, control of the dissolved oxygen (DO) concentrations or specifically the oxidation reduction potential (ORP) during operation and appropriate internal recirculation rates between the reactors. The results of this study provided information relevant for the assessment of both the BNR process and membrane performance. Membrane performance data indicated the importance and effect of air scouring (despite energy consumption) on membrane fouling for long-term stable flux operation as well as the cleaning frequency whether chemical enhanced backwash (CEB) or clean-in-place (CIP). This assessment also discussed how BNR systems can be enhanced through the incorporation of important design factors to eliminate the inhibiting factors of nitrogen and phosphorus removal such as dissolved oxygen. One of the biological processes tested in this study achieved effluent nitrogen and phosphorus concentrations below 5 mg/L and 1 mg/L respectively. Although the process tested did not meet NNC criteria, it can be applied with chemical precipitation. This, in turn, can reduce the operating and maintenance (O&M) costs associated with the chemical precipitation of phosphorus.

The recent development of culture-independent methods has revolutionized the study of complex microbial communities such as those present in activated sludge treatment systems. DNA probes that hybridize to genes coding for key enzymes that catalyze microbial processes have been widely used. Can such probes be used to quantify target genes and thus quantify the potential of a microbial community to carry out a reaction of interest? / Optimal conditions for DNA extraction, probe validation, hybridization, and activity measurements were determined for the pulp and paper treatment system environment under study. Using gene probes for key denitrification genes (nirS, nirK), the correlation between denitrifiers and denitrification activity in an enrichment culture and activated sludge samples was tested. The same correlation between nitrogen fixation and nitrogen-fixing bacteria in primary clarifiers was assessed using a probe for the gene encoding a component of the nitrogenase enzyme (nifH). This work was successful in establishing the correlation between gene numbers and their corresponding enzymatic activity and thus supports the quantitative hybridization approach for the monitoring of microbial communities. (Abstract shortened by UMI.)

The influence of nitrogen (N) and phosphorus (P) limitation on pulp and paper mill activated sludge (AS) floc properties was studied using a bioreactor fed with synthetic Kraft mill effluent. The bioreactor and synthetic effluent were designed and shown to perform like the real mill system providing the AS, establishing the in vivo relevance of the results. Limitation of either N or P produced inadequate effluent biotreatment, shown by poor BOD5 and suspended solids removal, and by decreased biomass health, performance, and floc settling. Greatly enhanced poly-beta-hydroxybutyrate (PHB) (but not carbohydrate or extracellular polymeric substances [EPS]) synthesis was the common response of the floc microbial community to N limitation over many days. In contrast, P-limitation increased total carbohydrate and EPS, but not PHB. / N limitation, but not P limitation, caused the net floc surface charge to be much more negative, while P-limitation, but not N-limitation, increased the floc bound water content and surface hydrophobicity. Thus, in real pulp and paper mill AS systems, careful manipulation of N or P additions may be useful to optimize the key process of charged polymer-assisted AS dewatering.

Thesis (M.Tech.)-Department of Biotechnology, Durban University of Technology, 2006 xvi, 126 leaves / The microbial community present in a laboratory-scale modified Ludzack-Ettinger activated sludge system was investigated using a combination of novel molecular techniques. The parent system was investigated for a duration of one year and samples were taken at regular intervals to determine the profile and structure of the microbial community present within the anoxic and aerobic zones of the MLE system. The combination of molecular techniques included fluorescent in situ hybridisation (FISH) and denaturing gradient gel electrophoresis (DGGE). FISH was performed using oligonucleotide probes, which were complementary to conserved regions of the rRNA for the alpha, beta and gamma subclasses of the gram negative family Proteobacteria as well as a group-specific HGC oligonucleotide probe as a representative of the gram positive actinomycetes branch. The total eubacteria present was determined using the EUB oligonucleotide probes, EUB388, EUB388-II and EUB388-III. The DGGE analysis of PCR-amplified 16S rDNA gene segments was used to examine the microbial community profile in the anoxic and aerobic zones. The profile for each of the zones revealed a number of consistent bands throughout the duration of the laboratory-scale process. However, the profiles obtained suggested that a diverse microbial community existed within the aerobic and anoxic zones. The bands also indicated the presence of dominant and less dominant species of bacteria. Hybridisations obtained from the FISH analyses indicated that the alpha and gamma subclasses were predominant within the anoxic zone and the aerobic zone showed a dominance of the beta subclass of Proteobacteria. The steady state behaviour of the MLE system was confirmed with the results obtained from COD, TKN, nitrates and OUR analytical tests. COD and nitrogen mass balances were conducted to confirm the acceptance of the results obtained for each batch as an indication of the system performance for the MLE model. Nitrogen mass balances indicated an upset in the nitrogen levels for batches two and seven.

Thesis (M.Tech.: Biotechnology)-Dept. of Biotechnology, Durban Institute of Technology, 2005 xiv, 114 leaves : ill. 30 cm / The activated sludge process is the mostl used biological treatment process. Engineers and microbiologists are constantly seeking ways to improve process efficiency, which can be attributed to the increasing demand for fresh water supplies and proper environmental management. Since the inception of the activated sludge process, bulking and foaming have been major problems affecting its efficiency. Filamentous bacteria have been identified as the primary cause of bulking and foaming. Numerous attempts have been made to resolve this problem. Some of these attempts were effective as interim measures but failed as long term control strategies. The identification of filamentous bacteria and the study of their physiology have been hampered by the unreliability of conventional microbiological techniques. This is largely due to their morphological variations and inconsistent characteristics within different environments. To fully understand their role in promoting bulking and foaming, filamentous bacteria need to be characterized on a molecular level. The aim of this study was, therefore, to identify filamentous bacteria in pure culture with the purpose of validating these findings to the physiological traits of the pure cultures when they were isolated. Fourteen different filamentous cultures were used for this study. The cultures were identified using specific oligonucleotide probes via fluorescent in situ hybridisation and nucleotide sequencing. Prior to sequencing, an agarose gel and a denaturing gradient gel Electrophoresis profile were determined for each isolate. The various techniques were optimised specifically for the filamentous isolates. The isolates were identified as Gordonia amarae, Haliscomenobacter hydrossis, Acinetobacter sp./Type 1863, Type 021N, Thiothrix nivea, Sphaerotilus natans and Nocardioform organisms.

Thesis (D.Tech.: Biotechnology)-Dept. of Biotechnology, Durban University of Technology, 2008. xxiv, 322 leaves / Activated sludge models now in use worldwide for the design and operation of treatment systems use hypothetical concentrations of active organisms. In order to validate and calibrate model outputs, concentrations and activities of organisms responsible for nitrification and denitrification need to be reflected by actual measurements. This research has been initiated by the observation of an increasing gap of suitable techniques that exist in the direct measurement and separation of active biomass components, responsible for COD removal and denitrification.

The effectiveness of partial ozonation of return activated sludge for enhancing denitrification and waste sludge minimization were examined. A pair of nitrifying sequencing batch reactors was operated in either aerobic or alternating anoxic/aerobic conditions, with one control and one ozonated reactor in each set. The amount of solids decreased with the ozone dose. Biomass in the anoxic/aerobic reactor was easier to destroy than in the aerobic one, generating approximately twice as much soluble chemical oxygen demand (COD) by cell lysis. Increased COD favoured production of extracellular polymers in ozonated reactors, enhancing flocculation and improving settling. Floc stability was also strengthened in prolonged operation in alternating treatment, resulting in declined solids destruction. Dewaterability was better in alternating reactors than in aerobic ones indicating that incorporation of an anoxic zone for biological nutrient removal leads to improvement in sludge dewatering. The negative impact of ozonation on dewaterability was minimal in terms of the long-term operation. Ozone successively destroyed indicator estrogenic compounds, contributing to total estrogen removal from wastewater. Denitrification rate improved up to 60% due to additional carbon released by ozonation. Nitrification rates deteriorated much more in the aerobic than in the alternating reactor, possibly as a result of competition created by growth of heterotrophs receiving the additional COD. Overall, ozonation provided the expected benefits and had less negative impacts on processes in the alternating treatment, although after prolonged operation, benefits could become less significant. The alternating anoxic/aerobic reactor achieved twice the nitrification rates of its aerobic counterpart. Higher removal rates of estrogens were associated with higher nitrification rates, supporting the contention that the nitrifying biomass was responsible for their removal. The alternating treatment offered the better estrogen biodegradation. Microbial populations in both reactors were examined with fluorescent in situ hybridization. Dominance of rapid nitrifiers like Nitrosomonas and Nitrobacter (79.5%) in the alternating reactor, compared to a dominance of slower nitrifiers like Nitrosospira and Nitrospira (78.2%) in the aerobic reactor were found. The findings are important to design engineers, as reactors are typically designed based on nitrifiers’ growth rate determined in strictly aerobic conditions.

The effect of engineered nanoparticles (ENPs) mixture consisting of silver oxide, (Agg0[Silver Oxide Nanopartical], 20 nm), titanium dioxide, (TiO2[Titanium dioxide], 30-40 nm) and zinc oxide, (ZnO, 20 nm) compared with their bulk metal salts was evaluated against unspiked activated sludge (control) using 3 parallel pilot-scale treatment plants. The total concentration of the ionic species of Ag+ Ti[Silver + Titanium] and Zn(2+) in the effluent of the ENP spiked activated sludge (AS) was below limits of detection and> 99% of the spiked ENP were found in the waste activated sludge (WAS), whereas 39 – 58 % of Ag0[Silver Oxide Nanopartical], 51 – 63 % and 58 – 74 % of ZnO ion concentrations were recovered in the anaerobic digestate (AD) cake suggesting higher affinity of ENPs to WAS than to anaerobic digestate. ENPs induced a 2-fold increase of the microbial community specific oxygen uptake rate (SOUR) compared with the control and > 98 % of ammonia and 80 % of COD were removed from the AS suggesting that the heterotrophic biomass retained their ability to nitrify and degrade organic matter at the spiked ENP concentration. The floc size and cultivable microbial abundance was reduced in the ENP spiked AS with no apparent disruption of the overall AS process efficiency. However, scanning electron microscopic analysis clearly showed damage to specific microbial cells. The lipid fingerprint and 16S rRNA gene-based pyrosequencing evidenced the dominance of Proteobacteria, Firmicutes, and Bacteriodetes with a clear temporal shift in microbial community structure. The prominent nano-tolerant bacterial species identified were Acidovorax, Rhodoferax, and Comamonas whereas Methanocorpusculum and Methanosarcina were recovered in AS and were the dominant Archaea in the AD with 99 and 98 % similarities to the closest culturable relative. Their presence in the AS suggests tolerance to ENPs and oxygen-dependent respiration. V. fisheri activity was not sensitive to the ionic concentrations of the ENP or metal salt mixture in the digestate samples and illustrates the need to develop bioassay using indigenous wastewater microorganisms to detect the potential effect of ENP. Overall, unlike other xenobiotic compounds, ENPs can hasten the natural selection of microbial species in activated sludge and anaerobic digestion processes.