Research currently being conducted at Virginia Tech has produced a viable pesticide wastewater clean-up procedure using biologically based materials. The system consists of two steps. First, pesticide laden wastewater is mixed with a lignocellulosic sorbent such as peat moss in a demulsification and sorption step. Second, the wastewater filtered rinsate is then passed through a packed column containing the same sorbent in a column sorption and filtration step. Although the system yielded adequate results and performed the tasks commensurate with its design, the original batch demulsification and sorption phase of the process had several operational problems. Namely, the system was large and not very portable, the recirculation system was not very reliable, and the entire step one process was laborious and time consuming to execute. These problems and limitations prompted the redesign of the step one process into a new, more efficient system. The new system consists of a steel frame similar to the old system. However, the new frame is smaller, lighter, more portable, and one person can easily operate and maneuver the new system. In addition, the new system provides a safer work environment for the operator. The frame surrounds the barrel and liner like a cage so that if the hand winch or the cable fails when lifting the liner out of the barrel, the operator is protected. The recirculation system has been automated such that it 1s self priming, and turns off when all the rinsate contained between the inner liner and outer barrel has been pumped dry. The come-a-long has been replaced with a hand wench that makes loading and unloading of the inner liner easier. Finally, the same pump that is used to recirculate the rinsate is used to transfer the rinsate onto the column for the second sorption and filtration step. There is now no need for additional pumps. After the system was designed and developed, it was tested on two pesticides, atrazine and metolachlor, to determine if the new system yielded results comparable to the old system. The atrazine concentration in the rinsate was reduced from an initial 721 mg/kg (ppm) in step one to 2 mg/kg in the step two column sorption and filtration phase. This represents an atrazine reduction of 99.72% over the entire two step process. The metolachlor concentration in the rinsate was reduced from an initial 704 mg/kg in step one to 30 mg/kg in the step two column sorption and filtration phase. This represents a metolachlor reduction of 95.74% over the entire two step process. The percent reduction of pesticide from the rinsate in the new system is comparable to the 99% reductions obtained using the old system. / Master of Science

The objective of this study was to investigate what effect the concentration of microorganisms would have on substrate removal, microbial substrate storage, and oxygen utilization at a constant food-to-microorganism ratio. Batch experiments were conducted, under aerated and completely mixed conditions, using a domestic wastewater, a paper mill wastewater, and a food processing wastewater. A series of three batch experiments were run for each of these wastes. The food-to-microorganism ratio for each series was kept constant while the mixed liquor suspended solids concentration was varied for each of the experiments within the series. The following analyses were conducted on samples that were withdrawn at specified time intervals: filtered and settled COD, oxygen uptake, mixed liquor volatile suspended solids (MLVSS), pH, protein concentration and carbohydrate concentration. No significant uptake and subsequent release of organic substrate was observed for any of the wastewaters studied. For the same F/M ratio, the rate of removal of organic substrate and the degree to which it was removed in the activated sludge system was found to be a direct function of the MLVSS concentration. The change in the cellular carbohydrate to cellular protein ratio in the activated sludge during substrate metabolism was a function of the MLVSS concentration. As the MLVSS concentration increased, the carbohydrate to protein ratio, which is an indicator of substrate storage, also increased, even though the F/M ratio was held constant. / Master of Science

M.S.

Industries producing highly polluted waste waters are having to purify their effluents to meet with ever increasing requirements laid down by water authorities. The South African Water Act of 1956 has prescribed a very high standard to which waste waters must conform before discharge into a South African water course. Enforcement of these standards falls under the jurisdiction of government authorities such as the Department of Water Affairs. Similarly, municipalities and other local authorities set standards with which trade effluents must comply before discharge into public sewers for treatment in a municipal sewage works. These local authorities are empowered to recover from the trader the additional costs incurred in treating trade effluents. Costs are usually levied in respect of volume, oxygen demand, settleable solids and the production of secondary sludge. In recent years, these standards have been enforced to an extent where the survival of several industries has become dependant on whether these industries are able to purify or dispose of their effluents in a manner acceptable to the water authorities. Chap. 1, p. 1.

The anaerobic digestion of tannery wastewaters was investigated with a view to using this form of treatment in the tanning industry. As these wastewaters are extremely complex and contain high concentrations of both inorganic and organic compounds, the effect of these individual compounds on the anaerobic digestion process was investigated in detail, in order to ascertain the fate of these compounds during the digestion process. The experiments comprising the initial toxicity study were carried out as adaptation experiments using a synthetic wastewater. It was found that the heavy metals such as chrome, aluminium and iron precipitated and accumulated in the sludge bed of the digesters . The soluble ions such as sodium and chloride were not retained and passed through the digesters. Approximately 20 % of the calcium ions were removed through precipitation, with the remainder being present in the digester effluent . Under the anaerobic conditions, ammonification of the organic nitrogen occurred, and influent sulphates were reduced to sulphides . These sulphides were present as either H2S, HS or insoluble sulphides. As these compounds under investigation on caused no inhibition of the anaerobic digestion process at the concentrations found in tannery wastewaters, the anaerobic treatment of these wastewaters appeared to be possible, provided the bacteria were given sufficient time to adapt to the potentially toxic compounds. However, despite the findings of the synthetic study, the successful anaerobic digestion of the tannery effluents could not be achieved. Although the use of acid was found to be essential in order to control the digester pH in the optimum range, the metabolism of the methanogenic bacteria was inhibited by the presence or absence of unknown compounds. Neither the addition of essential trace nutrients, nor the prevention of the competition between the methanogens and the sulphate-reducing bacteria were able to reverse this inhibition. As tannery effluents contain very low concentrations of phosphorous, it is possible that the methanogens were inhibited by a lack of phosphorous, which is essential during methanogenesis. In contrast to the results obtained from the effluent experiments, the anaerobic digestion of tannery sludge was found to be possible. Of the organic solids present in the sludge, 60 % were degraded and converted into biogas, which had a methane content greater than 70 %. The degradation of the organic solids ensured that COD and PV reductions of greater than 90 % were achieved, and the fate of the compounds in the digesters were in agreement with the findings of the v synthetic study. Efforts to improve the efficiency of the digestion process through the addition of trace nutrients and the use of a two-stage process were only successful in bringing about a minor improvement in digester performance. The overall results of this investigation show, therefore, that although the anaerobic treatment of the tannery effluent was not achieved, the successful anaerobic digestion of tannery sludge is possible at low loading rates. As many difficulties still need to be solved, a great deal of further research is necessary if anaerobic digestion is to be used on an industrial scale for the treatment and disposal of tannery wastewaters.

Thesis (M.Tech.: Biotechnology)-Dept. of Biotechnology, Durban Institute Of Technology, 2005 xxiii, 172 p. : ill. ; 30 cm / The provision of safe and sanitary water is a constitutional right and above all, a necessity of life. As a result of the rapid urbanisation and the past policies of apartheid, a large population of South Africa dwell in informal settlements, where there is very little hope of development, as the government does not possess the resources that are necessary for a full-scale sanitation programme. Therefore, on-site treatments have been considered to provide sanitation in these dense peri-urban areas. The anaerobic baffled reactor (ABR) is one such sanitation system. This reactor utilises the phenomenon of anaerobic digestion to degrade substrates. One of the major disadvantages of any anaerobic treatment processes is the extreme sensitivity of the bacterial communities, thus inducing slow recovery rates following toxic shocks. Therefore, an understanding of these microbial consortia is essential to effectively control, operate and optimise the anaerobic reactor. Fluorescence in situ hybridization, 4’,6-diamidino-2-phenylindole (DAPI) staining and DNA sequencing techniques were applied to determine the microbial consortium, as well as their reactions to daily operating conditions. With an understanding of these populations and their responses to perturbations within the system, it is possible to construct an anaerobic system that is successful in its treatment of domestic wastewater. In situ hybridizations were conducted for three operating periods, each characterised by specific flow rates. Results showed Eubacterial population dominance over the Archaeal population throughout both of the operating periods investigated. However, these cells cumulatively consisted of 50% of the total biomass fraction, as determined by DAPI staining. Group-probes utilised revealed a high concentration of fermentative acidogenic bacteria, which lead to a decrease in the pH values. It was noted that the ABR did not separate the acidogenic and methanogenic phases, as expected. Therefore, the decrease in pH further inhibited the proliferation of Archaeal acetoclastic methanogens, which were not present in the second operating period. DNA sequencing results revealed the occurrence of the hydrogenotrophic Methanobacterium and Methanococcus genera and confirmed the presence of Methanosarcina. Sequencing of the bacterial DNA confirmed the presence of the low G+ C Gram Positives (Streptococcus), the high G+C Gram Positives (Propionibacterium) and the sulfate reducing bacteria (Desulfovibrio vulgaris). However, justifications were highly subjective due to a lack of supportive analytical data, such as acetate, volatile fatty acids and methane concentrations. Despite this, findings served to add valuable information, providing details on the specific microbial groups associated with ABR treatment processes.

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 (MSc)--University of Stellenbosch, 2005. / ENGLISH ABSTRACT: Biological removal of sulphate from wastewater can be achieved by using a gas mixture consisting of 80% hydrogen and 20% carbon dioxide as energy and carbon sources. A novel reactor, including a venturi device for optimal hydrogen gas-liquid contact, and geotextile for immobilisation of the sulphate reducing bacterial community, was introduced. Efficient, relatively stable sulphate removal was obtained when the reactor was operated in continuous mode. The maximum sulphate removal rate obtained when the reactor was 8% packed with geotextile, was 1 g S04/(L.d) and 4 g S04/(L.d) when the reactor was 80% packed with geotextile. Kinetic batch studies showed that the highest sulphate removal rates were obtained at 29.5 °C; a pH of 7.5; initial sulphate concentration of 4000 mg/L; initial alkalinity of 1600 mg/L; cobalt concentration of 3 mg/L and when excess hydrogen gas was fed compared to what is stoichiometrically required (900 ml/min). Nickel addition showed inhibition at increased concentrations (>3 mg/L). The biofilm structure was observed on the geotextile with electron microscopy, while the viability of the biofilm was indicated with fluorescence microscopy. These observations indicated the suitability of the geotextile as a support material for biofilm formation in the sulphate reducing system. The stability of the sulphate reducing community was analysed, using the T-RFLP protocol. It was shown that the composition of the community changed after a period of 3 months, when the reactor was subjected to environmental changes. The reactor was also observed to be more efficient in terms of sulphate removal after the environmental changes, of which the temperature change from an average of 39 to 29.5 °C was the most prominent. Subsequently, it was speculated that the population shift was in favour of a more efficient system for sulphate removal. A dynamic, viable, mesophilic sulphate reducing community was therefore observed on the geotextile support, responsible for successful sulphate removal in a novel venturi-reactor. Defining optimal operating conditions, and a knowledge of biofilm structure and composition may contribute to the successful implementation of the biological sulphate removal component of the integrated chemical-biological process for the treatment of industrial wastewater, when hydrogen and carbon dioxide are supplied as the energy and carbon sources, respectively. / AFRIKAANSE OPSOMMING: Ongewenste industriële afval-water kan biologies behandel word deur 'n gasmengsel van 80% waterstof en 20% koolstofdioksied te gebruik vir sulfaat verwydering. 'n Reaktor wat 'n venturi apparaat bevat vir optimale waterstofgas-vloeistof kontak, asook geotekstiel vir die immobilisasie van die bakteriële sulfaatverwyderende gemeenskap, is bekend gestel. Effektiewe, relatief stabiele sulfaatverwydering is waargeneem sodra die reaktor op 'n kontinue basis gevoer is. Die optimale sulfaat verwyderingstempo wat bereik is as die reaktor 8% met geotekstiel gevul was, was 1 g S04/(L.d) en 4 g S04/(L.d) wanneer die reaktor 80% met geotekstiel gevul was. Kinetiese groepstudies het getoon dat die beste sulfaatverwydering bereik is by 'n gemiddelde temperatuur van 29.5 °C; pH van 7.5; aanvanklike sulfaatkonsentrasie van 4000 mg/L; aanvanklike sulfied konsentrasie van 268 mg/L; aanvanklike alkaliniteit van 1600 mg/L; kobalt konsentrasie van 3 mg/L, asook wanneer 'n oormaat waterstofgas gevoer is (900 ml/min), in vergelyking met wat stoichiometries benodig word. 'n Verhoogde byvoeging van nikkel by die voerwater (3 mg/L), het tekens van inhibisie getoon. Die biofilm struktuur is waargeneem op die geotekstiel met behulp van 'n elektronrnikroskoop, terwyl die lewensvatbaarheid van die biofilm aangedui is met behulp van fluoressensie mikroskopie. Hiermee is die bruikbaarheid van geotekstiel as 'n ondersteunings-matriks bevestig. Die stabiliteit van die sulfaatverwyderende gemeenskap is ondersoek deur die T-RFLP protokol te gebruik. Hiermee is aangedui dat die samestelling van die gemeenskap verander het na die 3 maande toets periode, toe die reaktor onderhewig was aan omgewings veranderinge. Die reaktor het ook 'n verbetering in sy sulfaatverwyderings vermoë getoon na hierdie tydperk van omgewingsveranderinge, waarvan 'n temperatuur verandering vanaf 'n gemiddeld van 39 na 29.5 °C die prominentste was. Dit is dus gespekuleer dat die populasie verskuiwing ten gunste was van 'n beter sisteem vir sulfaatverwydering. 'n Dinamiese, lewensvatbare, mesofiliese sulfaatreduserende gemeenskap, verantwoordelik vir die sulfaatverwydering in die venturi-reaktor, is dus waargeneem op die geotekstiel as 'n ondersteuningsmatriks. Met hierdie evaluasie kan die insig wat verkry is in die reaktor samestelling en die optimale kondisies vir die reaktor werking, bydra tot die suksesvolle implementasie van die biologiese komponent, in die geïntegreerde chemies-biologiese proses vir die behandeling van industriële afval water, wanneer 80% waterstof en 20% koolstofdioksied gas as energie en koolstofbron respektiewelik, gebruik word.

published_or_final_version / Civil Engineering / Master / Master of Philosophy