Thesis (M.Sc. (Aquaculture)) -- University of Limpopo, 2014 / Population growth in urban areas is putting pressure on sewage treatment plants. The improper treatment of sewage entering the aquatic ecosystems causes deterioration of the water quality of the receiving water body. The effect of sewage effluent on the Sand River was assessed. Eight sampling sites were selected, site 1 and 2 were upstream of the of the sewage treatment plant along the urbanised area of Polokwane, whilst sites 3, 4, 5, 6, 7 and 8 were downstream. The physicochemical parameters and coliform counts in the water samples were determined. Macroinvertebrate abundances and diversity (H') was determined at the different sites during the dry and rainy season. The water quality status of the Sand River with respect to the South African scoring system (SASS) scores and average score per taxon (ASPT) was determined. A linear regression was performed to test the correlation of the SASS scores with abundance and H'. Heavy metal concentrations in water, sediment, grass (Ishaemum fasciculatum) and fish (Oreochromis mossambicus) at the sites were evaluated. The suitability of the Sand River and surrounding borehole water for irrigation was also determined. Hierarchical average linkage cluster analysis produced two clusters, grouping two sites above the sewage treatment works and six sites downstream of the sewage effluent discharge point. Principal component analysis (PCA) identified total nitrogen, total phosphorus, conductivity and salinity as the major factors contributing to the variability of the Sand River water quality. These factors are strongly associated with the downstream sites. Canonical correspondence analysis (CCA) indicated that Chironomidae family was found on the nitrogen gradient during the dry season. However during the rainy season, Chironomidae was found in the centre of the ordination which indicated that it was ubiquitous. Escherichia coli levels (1463.73 counts/100ml) in the maturation ponds of Polokwane wastewater treatment works could potentially lead to contamination of the Polokwane aquifer. High diversity was recorded at the sites before discharge and the sites further downstream. There was significant correlation (P<0.05) between the SASS scores and macroinvertebrate diversity during the dry season (R2=0.69) and the rainy season (R2=0.77). Fish samples had significantly higher (P<0.05) iron and copper concentrations, while the vi sediment had significantly higher lead concentration (P>0.05). The United States salinity laboratory (USSL) diagram indicated that the sodium hazard (SAR) and alkalinity hazard (conductivity) was low and the Sand River and borehole water was suitable for irrigation. The residual sodium carbonate (RSC) was below 1.24 meq/l, also indicating that both the Sand River and borehole water is still suitable for irrigation. The total phosphorus concentrations fluctuated across the different site. Total nitrogen concentrations showed a gradual decrease downstream from the point of discharge. The coliform levels also showed a gradual decrease downstream. This shows that the river still has a good self-purification capacity.

Almost 90% of the treated sewage effluent processed by the two treatment plants serving the greater Tucson area is available for passive recharge through the Santa Cruz River streambed north of Tucson. In the absence of any major disturbance of the effluent channel, the recharge capacity of the streambed materials decreases over time as microbial activity, and possibly suspended sediments settling out of solution, act to clog the surficial sediments under the effluent stream. Effluent stream transmission-loss measurements made over the period from November 1994 to August 1995 provided data used to determine the average vertical hydraulic conductivity of the low-flow channel in the study reach through simulations using the computer model known as KINEROS2. Saturated hydraulic conductivity (KSAT) served as the calibration parameter in the model. The appropriate KSAT value was chosen for each set of field data by matching the observed and simulated downstream hydrographs for the study reach. KSAT values were corrected for viscosity changes resulting from changing average daily surface water temperatures over the study period. Saturated hydraulic conductivity values for the effluent stream channel ranged from a maximum of 37 mm/hr in January, 1995, following several major winter storms, to a minimum of 11 mm/hr in August, 1995, after a nearly six-month interstorm period. The saturated hydraulic conductivity values decay exponentially with time after the last major winter storm. The mathematical model describing this decay may be used to estimate effluent recharge rates under similar future meteorological and climatological conditions.

Thesis (M. Sc.)--University of Hong Kong, 1999. / Includes bibliographical references.

Sewage effluents disposed into the marine environment from De Beers Marine Namibia diamond mining vessels have the potential to cause endocrine disruptive effects in marine organisms. Endocrine disruption refers to the alteration of the normal functioning of the endocrine system and various chemicals have the ability to mimic hormones, effecting endogenous hormone synthesis, transport, receptor interaction and intracellular signaling. The potential endocrine disruptive effects, caused by the release of different types of sewage effluents into the ocean, on fish species is a concern due to the commercial importance of fish species found in the mining area e.g. hake, sole, horse mackerel. Increased awareness of marine environmental degradation due to the presence of chemical contaminants has resulted in research being done on early warning systems, in the form of biomarkers. Cytochrome P450 monooxygenase 1A (CYP1A) and vitellogenin (vtg) are important proteins found in fish liver and blood, that have been used as biomarkers for the detection of pollutants in fish. CYP1A is a subfamily of the P450 superfamily of enzymes and catalyzes the oxidation, hydrolysis and reduction of exogenous and endogenous compounds (phase I reactions) and thus has the capacity to regulate the metabolism of several organic contaminants. CYP1A expression is altered by exposure to planar xenobiotic compounds e.g. polyaromatic hydrocarbons. Vtg is an important precursor for egg yolk proteins and plays a role in the growth and development of an oocyte. Expression of this protein is altered upon exposure to estrogenic compounds. The aim of this project was to isolate CYP1A from fish liver by differential centrifugation and optimize conditions for the CYP1A-mediated ethoxyresorufin-Odeethylase (EROD) assay and western blot analysis (to assess CYP1A expression). Another aim of this study was to evaluate the potential effects of biologically disruptive chemicals from sewage effluents, discharged into the marine environment, on the expression of CYP1A in two species of hake, Merluccius capensis and M. paradoxus (Cape hake). CYP1A in Cape hake is approximately a 60 kDa protein and the highest EROD activity was detected in the microsomal fraction after differential centrifugation. Optimal EROD assay conditions were observed at pH 7.5, a temperature of 25 °C, 10 μl of sample and a reaction time of 30 seconds. Enzyme stability assays indicated a drastic decrease in enzyme activity after 30 seconds. The EROD assay was not NADPH dependent but was limited by NADPH supply, with an increase of 300% in EROD activity being observed with the addition of 0.1 M exogenous NADPH. The addition of dicumarol (40 μM), a phase II enzyme inhibitor, showed a 232% increase in EROD activity. This is because dicumarol inhibited enzymes with the capacity to metabolize the product (resorufin) of the EROD reaction. With regard to western blot analysis, the optimal primary (rabbit antifish CYP1A peptide) and secondary (anti-mouse/rabbit antibody-horseradish peroxidase conjugate (POD)) antibody dilutions were determined to be 1:1000 and 1:5000, respectively. The comparison of CYP1A expression in Cape hake samples from De Beers Marine mining area and reference sites showed higher EROD activity (16.29 ± 0.91 pmol/min) in fish samples from the mining area in comparison to the reference site (10.42 ± 2.65 pmol/min). Western blot analysis was in agreement with the EROD assay results and a higher CYP1A expression was observed in fish from the mining sites. The increased CYP1A expression observed in fish from the mining area is not definitively an indication of a pollutant effect in the environment, as several environmental and biological factors (e.g. photoperiod and age) must also be considered before reaching this conclusion. Another aim of this study was to purify vtg from Cape hake blood samples. Cape hake vtg was purified from fish plasma by selective precipitation with MgCl2 and EDTA. Precipitated sample was subjected to anion exchange chromatography using fast protein liquid chromatography (FPLC). Vtg eluted as two broad peaks and had a molecular weight above 200 kDa. SDS-PAGE analysis also resolved smaller molecular weight proteins below 70 kDa, which were thought to be vitellogenin cleavage proteins, lipovitellin and phosphovitins. Western blot analysis was performed; however, it did not produce any conclusive results. The purification of vtg enables further studies in characterizing this protein and developing assay aimed at detecting estrogenic pollutants in the marine environment

The ecological position of micro-algae at the base of the aquatic food web makes them critical components of aquatic ecosystems. Their short generation time also makes them useful biological indicators because they respond quickly to changes in environmental condition, enabling timely identification and assessment of water quality changes. The inclusion of micro-algae as indicators in water resource regulation and management in South Africa has started recently, their more extensive use in biomonitoring and ecotoxicology programmes for water resource management would contribute to the South African policy if water resource protection. The standard algal growth inhibition assay with the species Pseudokirchneriella subcapitata is currently used for monitoring toxicity of in-stream and industrial wastewater discharges to freshwater micro-algae. The relevance of the data generated by standard toxicity bioassays has been questioned, since micro-algae in particular are extremely variable in their sensitivity to a range of contaminants and these standard species used may not occur in the local aquatic environment. As a result, international regulatory agencies, have recommended algal growth inhibition tests be changed from a single standard species to tests with a number of species. One recommendation, in addition to the use of standard toxicity tests, is the use of species isolated from the local environment which may be more relevant for assessing site specific impacts. This study investigated the value and application of locally isolated South African freshwater micro-algae in toxicity tests for water resource management and was carried out in three phases. The first phase involved isolating micro-algae from South African aquatic resources. Micro-algae suitable for toxicity testing were identified and selected using as set of criteria. Three (Scenedesmus bicaudatus, Chlorella sorokiniana and Chlorella vulgaris) out of eight successfully isolated species satisfied the prescribed selection criteria and these were selected as potential toxicity test species. The second phase focused on refining and adapting the existing algal toxicity test protocol (the algal growth inhibition assay) for use on the locally isolated algal species. The refinement of the test protocol was achieved by exposing the locally isolated species to reference toxicants in order to assess and compare their growth and sensitivity to the toxicants under the prescribed toxicity test conditions with that of the standard toxicity test species (Pseudokirchneriella subcapitata) and a commercial laboratory species (Chlorella protothecoides). During this phase, one of the three local species (Scenedesmus bicaudatus) was eliminated as a potential toxicity test species due to inconsistent growth. The third phase of the study involved assessing the sensitivity of the two remaining species (C. vulgaris and C. sorokiniana) to a range of toxicants (reference toxicants, salts, effluents and a herbicide) and comparing it to that of the standard toxicity test species P. subcapitata and C. protothecoides. The toxicants were selected based on their relative importance in the South African context, as well as the practicality of using these local micro-algae to routinely determine the impact of these toxicants on local aquatic resources. The growth of the four micro-algae was stimulated by the selected effluents. The standard toxicity test species P. subcapitata was ranked the most sensitive and of the four species to two reference toxicants and two inorganic salts. Chlorella sorokiniana was ranked the most sensitive of the three Chlorella species to two reference toxicants and two inorganic salts. The herbicide stimulated the growth of C. vulgaris while inhibiting the growth of the other species. Pseudokirchneriela subcapitata and C. sorokiniana showed high intra-specific variability in growth, which made it difficult to determine the effective concentrations of the herbicide and therefore compare the sensitivity of the species. This varied response of micro-algal species to toxicants may result in the biodiversity shifts in aquatic ecosystems, and also supports the recommendation of using a battery of different species to support more informed decisions in water resource management.

The Green Drop Certification Programme, launched in 2008 alongside the Blue Drop Certification Programme, aims to provide the Department of Water Affairs with a national overview of how municipalities and their individual wastewater treatment works (WWTW) are complying with licence conditions set by the National Water Act (NWA) (No. 36 of 1998; DWAF 1998) and the Water Services Act (No. 108 of 1997; DWAF 1998). By publishing the results of each municipality’s performance, the programme aims to ensure continuous improvement in the wastewater treatment sector through public pressure. The programme has been identified by this project as a necessary linking tool between the NWA and the Water Services Act to ensure protection and sustainable use of South Africa’s natural water resources. It does this through assisting municipalities to improve their wastewater treatment operations which in theory will lead to discharged effluent that is compliant with discharge licence conditions. These discharge licences form part of the NWA’s enforcement tool of Source Directed Controls (SDC) which help a water resource meet the ecological goals set for it as part of Resource Directed Measures (RDM). The link between meeting the required SDC and achieving the RDM goals has never been empirically tested. This project aimed to determine the present ecological condition of the Uie River, a tributary of the Sundays River which the Sundays River Valley Municipality (SRVM) discharges its domestic effluent into. It then determined whether the SRVM’s WWTW was complying with the General Standard licence conditions and what the impact of the effluent on the river was through the analysis of monthly biomonitoring, water chemistry and habitat data. Lastly, the project examined the effectiveness of the Green Drop Certification Programme in bringing about change in the SRVM’s wastewater treatment sector, which previously achieved a Green Drop score of 5.6 percent. It wanted to examine the underlying assumption that a WWTW which improves its Green Drop score will be discharging a better quality effluent that will help a water resource meets the RDM goals set for it. The Kirkwood WWTW did not have a discharge licence at the time of assessment and was thus assessed under the General Standard licence conditions. It was found that the Kirkwood WWTW was not complying with the General Standard discharge licence conditions in the Uie River. This was having a negative impact on the river health, mainly through high concentrations of Total Inorganic Nitrogen (TIN-N), orthophosphate and turbidity. The SRVM should see an improvement in its Green Drop score for the Kirkwood WWTW. However, the municipality showed no implementation of necessary programmes. Implementation of these programmes would help the SRVM meet the General Standard licence conditions (part of SDC) which would help the Uie River meet the RDM goals set for it.