M.Sc. / During the last decade, Cape Wagtails (Motacilla capensis) have been observed with several forms of feet abnormalities, such as missing toes and clubbed feet. The aim of this research was to find the reason for these abnormalities. The initial hypothesis was that ‘Feet abnormalities in the Cape Wagtail (M. capensis) are caused by the internal action of contaminants (e.g. pesticides and metals) through direct contact and/or secondary poisoning’. Wagtails were caught with mist nets and/or ground traps depending on the area and accessibility of the sampling sites. Wagtails with feet abnormalities, as well as those without, were caught at six localities around South Africa. Indicators of pollutant exposure and effect were studied. The bioaccumulation of certain metals (measures of exposure) in the feathers such as aluminium (Al), arsenic (As), cadmium (Cd), chromium (Cr), cobalt (Co), copper (Cu), iron (Fe), Pb, nickel (Ni), manganese (Mn), silver (Ag), strontium (Sr) and zinc (Zn) and Al, As, calcium (Ca), Cd, Cr, Co, Cu, Fe, Pb, Ni, Mn, Ag and Zn in the blood plasma were analyzed on the Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Biomarker testing (measures of effect) was carried out in the blood such as acetylcholinesterase (AChE) which tested for nerve transmission inhibition, ä-aminolevulinic acid dehydratase (ALA-D) which tested for lead (Pb) bioaccumulation, catalase (CAT) which tested for the invasion of oxyradicles and oxidative stress and DNA damage which tested for DNA band length alteration caused by stress due to xenobiotics. Biomarker outcomes and metal analysis results were carried out on a site comparison basis. Acetylcholinesterase, ALA-D and DNA damage were found not to be linked to the occurrence of feet deformities at these sites. However, CAT revealed stress at all the sites which may have indicated the presence of a stress (or stresses) which could have been caused by feet abnormalities or other factors. A non-metric multidimensional scaling graph (NMDS) ordination suggested that biomarkers were not grouped according to sites that had affected birds and those that did not, but were rather assorted. Consequently, few real differences in biomarker responses were noted between healthy and affected birds. Catalase enzyme activity and DNA damage were suggested to be indicative of feet abnormalities although results were not convincing. Secunda displayed the highest concentrations of metals in the feathers when compared to the other sites. Most metals were higher than reference values for contaminated areas and recent South Africanreported concentrations. Almost all of the metals tested at each site exceeded the Ecological Quality iii Objectives (EcoQOs) for metals in bird feathers, based on Weavers (Ploceus sp.) in the Gauteng region. Zinc levels at all the sites posed some concern. Chromium and Ni were suspected to possibly trigger feet and toe abnormalities in Cape Wagtails. Levels of various metals in the plasma were higher at sites that had no deformities. This suggested that these metal concentrations did not cause or influence feet or toe deformities. Calcium levels were found to be lower at sites with affected birds which indicated that the deformities may perhaps be caused by decreased Ca concentrations. Metal concentrations were higher than the uncontaminated reference levels, but were lower than contaminated site reference values. / Prof. V. Wepener

A static-renewal, seven day test was developed and utilized to study the effects of nutrition on the sensitivity of Mysidopsis intii Holmquist to zinc. It was found that juvenile growth was the most reliable and sensitive endpoint that could also reflect the subtle effects of low zinc concentrations and diet modifications. It was shown that even slight diet modifications could significantly alter growth over seven days. It was also shown that this small change in overall growth was accompanied by a heightened sensitivity to zinc. This heightened sensitivity was evident as a lowering of the Lowest Observed Effect Concentration of zinc in diets that suppressed growth. Though this shift in response to a toxicant is subject to variability, the fact that it was observed should make researchers and regulators aware of the potential significance of dietary influences on bioassays of pollutants. / Graduation date: 1995

PhD. (Chemistry) / This project assessed the toxicity of double-walled carbon nanotubes to three aquatic organisms belonging to different trophic levels, namely Pseudokirchneriella subcapitata (algae), Daphnia pulex (macro-invertebrate) and Poecilia reticulata (fish). Prior to the toxicity testing, the dry DWCNTs were characterised using scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray spectroscopy and Raman spectroscopy. Dynamic light scattering was used to characterise DWCNT suspensions. Natural water parameters such as increased ionic strength (Ca2+ and Na+) and increased humic acid affected the agglomeration potential of DWCNTs in aquatic medium. Increased ionic strength increased the agglomeration of DWCNTs while humic acid decreased agglomeration. The study explored the lethal/effective concentrations and sublethal effects of DWCNTs on the three organisms. The LC50/EC50 of DWCNTs for the three organisms differed in order of magnitude with D. pulex being the most sensitive and P. reticulata being the least sensitive. The LC50 for D. pulex was 2.81 and 4.45 mg/L for pristine and oxidised DWCNTs, respectively. Pseudokirchneriella subcapitata had an average EC50 of 10.01 mg/L and 10.93 mg/L for pristine and oxidised DWCNTs, respectively. Poecilia reticulata had an LC50 of 113.64 mg/L and 214.0 mg/L for pristine and oxidised DWCNTs, respectively. Exploring the effects of natural water parameters such as humic acid and ionic strength revealed that the acute toxicity of DWCNTs to D. pulex and P. reticulata was increased with increasing humic acid concentrations in exposure media, but increased ionic strength decreased the toxicity of both pristine and oxidised DWCNTs. However, these water parameters all decreased the toxicity of DWCNTs to P. subcapitata. The acute toxicity of DWCNTs was found to be directly linked to their agglomeration state in aquatic systems. Humic acid decreased the hydrodynamic sizes of DWCNT agglomerates making the engineered nanomatrials (ENMs) more available to the organisms while the cations increased the hydrodynamic sizes of DWCNT agglomerates, thereby reducing the probability of interactions with organisms. Time-based survival plots revealed that for P. reticulata and P. subcapitata, there were steady mortality/growth inhibitions throughout the duration of the exposures. For D. pulex, however, the plots revealed that there was a high initial die-off, whereafter mortalities proceeded at different rates. An assessment of whether DWCNTs cause oxidative stress in the three organisms revealed that DWCNTs caused significantly high oxidative stress in D. pulex and P. reticulata but not in P. subcapitata. In D. pulex and P. reticulata, DWCNTs were found to also cause DNA damage. The sublethal toxicity of DWCNTs was affected differently by the humic acid and increased cation concentration in exposure experiments. The sublethal effects were linked to the mode of interaction between DWCNTs and organisms. In P. subcapitata, the interaction was mainly physical with DWCNTs entrapping the algal cells in agglomerates and depriving the algal cells of light for normal photosynthesis to take place. For the other two organisms, the interaction was through intestinal cells as the organisms ingested DWCNTs and through accumulation of nanotubes on the exterior or organisms. The intestinal cell/DWCNT interaction resulted in the excessive generation of reactive oxygen species (ROS) and led to the death of the organism. Humic acid induced the highest antioxidant responses in both D. pulex and P. reticulata and this led to increased DNA damage in both organisms. Increased ionic strengths induced increased antioxidant responses at some DWCNT concentrations but the DNA damage was not significantly increased. These results suggested that with humic acid, the ROS production was excessive and sustained and had an effect on the DNA. The ROS production in increased ionic strengths was not excessive and was not prolonged, reducing their impact on DNA. The use of three organisms to assess the toxicity of DWCNTs provided comprehensive information on the potential effects of these ENMs in the aquatic food chain. Moreover, a multi-tier approach provided information on the potential effect of DWCNTs on populations at sublethal concentrations.

Thesis (PhD)--Stellenbosch University, 2002. / ENGLISH ABSTRACT: Metals occur naturally in the environment but since the industrial revolution the amounts ofbioavailable metals in the environment have increased significantly. There are various anthropogenic sources of metals in the environment. In recent years there has been an increasing interest in the use of biomarkers in terrestrial invertebrates for the assessment of the potential adverse effects of chemicals in soil ecosystems. Terrestrial isopods are one of the groups that play an important role in the decomposition process as fragmentors of dead plant material in soil ecosystems. They are also known to accumulate contaminants in their bodies, especially in the hepatopancreas. Cadmium has no known biological function and is deposited in the environment through a variety of sources. Zinc on the other hand is one of the essential metals and play an important role in the normal metabolism of animals. Zinc too, is deposited in the environment through a variety of anthropogenic sources. The aim of this study was to undertake an experimental and field evaluation of the role cellular change in the hepatopancreas of terrestrial isopods can play in conjunction with other sublethal responses to contribute to the assessment of the impact of cadmium and zinc in terrestrial ecosystems. Porcel/io laevis was used as test species in the laboratory experiments in this study, to conduct sublethal toxicity tests. Cadmium- and zinc sulphate were used as the contaminants, applied separately and as mixtures. The isopods were weighed before the beginning of the exposures and every week throughout the exposure period of six weeks. At the end of the exposure period they were dissected to remove the hepatopancreas. The zinc and cadmium content of the hepatopancreases and rest of the bodies of the isopods were determined by atomic absorption spectrophotometry. Hepatopancreas samples were also prepared for histological analysis, and measurements of the Percentage Cellular Area (PCA) were made using image analysis. Specimens of Porcellionides pruinosus were collected from contaminated- and uncontaminated field sites. Hepatopancreas- and rest of the body samples of P. pruinosus were also analysed for cadmium and zinc. Histological sections of the hepatopancreas were similarly prepared and analysed, as in the laboratory experiments. Exposure to cadmium- and zinc sulphate affected the mass of P. laevis negatively in the single metal exposures. However, these two metals "neutralised" each other's effects in the mixture toxicity exposures, pointing towards an antagonistic interaction. Administered cadmium- and zinc sulphate resulted in the accumulation of cadmium and zinc in the isopods, especially in the hepatopancreas. In the mixture toxicity experiments cadmium and zinc influenced the accumulation of each other. Cadmium- and zinc sulphate, whether administered separately or as mixtures, changed the structure of the hepatopancreas and this was quantitatively shown through PCA measurements. PCA of the hepatopancreas of terrestrial isopods was shown to be a good general biomarker of exposure and effects for cadmium as well as for zinc. It is suggested that PCA could serve as a general biomarker to predict possible impairment of growth or mass change of isopods exposed to cadmium and zinc mixtures. The findings of the field survey also suggest that PCA may be suitable to be used as a general biomarker to measure metal induced stress in woodlice from contaminated field sites. / AFRIKAANSE OPSOMMING: Metale kom natuurlik voor in die omgewing, maar sedert die industriële revolusie het die hoeveelheid bio-beskikbare metale in die omgewing betekenisvol toegeneem. Daar is verskeie antropogeniese bronne van metale in die omgewing. Die afgelope paar jaar was daar toenemende belangstelling in die gebruik van biomerkers by terrestriële invertebrate vir die assessering van potensiële negatiewe effekte van chemiese stowwe in grondekosisteme. Terrestriële Isopoda is een van die groepe wat 'n belangrike rol speel III die ontbindingsproses as fragmenteerders van dooie plantmateriaal III grondekosisteme. Hulle akkumuleer ook kontaminante in hul liggame, veral in die hepatopankreas. Kadmium het geen biologiese funksie nie, en word vrygestel in die omgewing deur verskeie bronne. Sink, aan die ander kant, is een van die essensiële metale en speel 'n belangrike rol in die metabolisme van diere. Sink word ook in die omgewing vrygestel deur 'n verskeidenheid antropogeniese bronne. Die doel van die studie was om deur eksperimentele en veldondersoek die rol van sellulêre veranderinge in die hepatopankreas van terrestriële Isopoda in samehang met ander subietale response te evalueer om by te dra tot die assessering van die impak van kadmium en sink op terrestriële ekosisteme. Porcellio laevis is as toets-spesie in die laboratorium eksperimente gebruik om subietale toksisiteitstoetse mee uit te voer. Kadmium- en sinksulfaat is as kontaminante gebruik, en is apart en gemeng toegedien. Die houtluise is geweeg voor die begin van die blootstellings en elke week deur die loop van die blootstellingsperiode van ses weke. Aan die einde van die blootstellingsperiode is hulle gedissekteer om die hepatopankreas te verwyder. Die sink en kadmium inhoude van die hepatopankreas en res van die liggame van die houtluise is bepaal met behulp van atoomabsorpsiespektrofotometrie. Monsters van die hepatopankreas is ook voorberei vir histologiese analise, en metings van die Persentasie Sellulêre Oppervlak (PSO) is gedoen met behulp van beeldanalise. Eksemplare van Porcellionides pruinosus is versamel van 'n gekontamineerde- en 'n ongekontamineerde gebied. Monster van die hepatopankreas en res van die liggaam van P. pruinosus is ook geanaliseer vir kadmium en sink. Histologiese sneë van die hepatopankreas was ook voorberei en geanaliseer, soos in die laboratorium eksperimente. Blootstelling aan kadmium- en sinksulfaat het die massa van P. laevis negatief beïnvloed in die enkelmetaal-eksperimente. Hierdie twee metale het egter mekaar se effekte in die mengseltoksisiteitstoetse "geneutraliseer". Dit dui op 'n antagonistiese interaksie. Toegediende kadmium- en sinksulfaat het gelei tot die akkumulasie van kadmium en sink in the houtluise, veral in die hepatopankreas. In die mengseltoksisiteitstoetse het kadmium en sink mekaar se akkumulasie beïnvloed. Kadmium- en sinksulfaat, apart of as mengsels toegedien, het die struktuur van die hepatopankreas verander, en dit is kwantitatief aangetoon deur die PSO metings. PSO van die hepatopankreas van houtluise is 'n bruikbare algemene biomerker van blootstelling en effek vir kadmium en sink. Dit word voorgestel dat PSO kan dien as 'n algemene biomerker om negatiewe effekte op die groei of massaverandering van houtluise te voorspel wat aan kadmium en sink mengsels blootgestel word. Die bevindinge van die veldondersoek dui daarop dat PSO gepas mag wees om gebruik te word as 'n algemene biomerker om metaal-geïnduseerde stres by houtluise vanaf 'n gekontamineerde gebied te meet.

M.Sc. / The increase in industrialisation, mining and agricultural activity along rivers could have a detrimental effect on aquatic environments unless the dangers of pollutants are not taken notice of. Metal ions and industrial effluent have become a source of pollution in the watercourses of South Africa. Pollutants generally have negative effects on the physiology of aquatic biota in polluted waters. The effects of copper and industrial effluent by the exposure of the clawed toad, Xenopus laevis and Tilapia sparrmanii are presented in this study. An experimental static-renewal system with an exposure time of 96 hours was followed at 25±1°C. After copper and effluent exposure, several physiological changes occurred in the two aquatic organisms. The sublethal effects that occurred include changes in hatching, survival, behaviour, growth impairment and developmental limitations. The results of the present study suggest that lethal endpoints can be used as indicators in detecting and evaluating the effects of aquatic pollution, caused by copper and effluent. Individual variation, however, could hamper the conclusions made but the study of aquatic organisms is of practical importance when conducting experimental studies in a laboratory and does not have the same impact as during field studies. Apart from the exposure to sublethal concentrations of copper and effluent, computational derivations of LC50, NOEC values and 95% confidence limits were made. The obtained concentrations were used as assumptions that pollutants should not exceed for the protection of aquatic life. Statistically different differences were found between the chosen derived variables of control and experimental organisms. The advantages of FETAX solution over borehole water can be attributed to the bioavailability of pollutants, which appears to be much less in those solutions. The predicted NOEC values provide some information regarding the concentrations at which no effects will be observed and the Target Water Quality Ranges (TWGR) for water were used to determine if the diluent was correct.

published_or_final_version / Zoology / Master / Master of Philosophy

In near-shore marine environments, two of the most prevalent anthropogenic disturbances affecting ecosystem diversity and function are the invasion of exotic species, and the release of toxic chemicals. Estuarine and harbour environments worldwide are dually subjected to high levels of toxic disturbance and ever increasing rates of invasion, via activities such as boating and shipping. It is not clear, however, whether the success of invasive species is influenced by the presence of toxic contaminants in these impacted environments. In particular, nonindigenous marine species (NIS) may be advantaged in chemically disturbed environments if they are more tolerant of toxicants. This thesis examines whether nonindigenous and native sessile invertebrate species have differential tolerance to a common aquatic contaminant, copper (Cu), which has links to both the invasion process (via its use in antifouling paints and the vector of hull fouling) and the modification of recipient environments (via human-mediated pollution). A series of laboratory- and field-based experiments showed that the cosmopolitan invasive bryozoans Watersipora subtorquata, Bugula neritina and Schizoporella errata are highly tolerant of copper pollution. The larvae and adults of W. subtorquata and B. neritina in particular were able to survive and grow in copper-polluted environments far exceeding most observed real-world levels (< 100 /??g 1-1). These nonindigenous species also showed very strong postexposure recovery. In contrast, the co-occurring native bryozoans Celleporaria nodulosa and Fenestrulina mutabilis demonstrated comparatively low tolerance to copper, with markedly reduced survival and fitness under Cu contaminated conditions, and inferior recovery abilities post-exposure. These findings were further supported by a long-term manipulative field experiment, examining the effect that varying levels of Cu exposure have on the structure and diversity of developing sessile invertebrate assemblages. Cu exposure decreased native species diversity by up to 50%, yet had no effect on NIS numbers. Consequently, in the presence of copper contamination, NIS dominated sessile communities, often resulting in substantial changes to community structure. Important insights were also gained into the nature of metal tolerance in NIS. Toxicity studies on larvae of the introduced bryozoan Bugula neritina revealed intraspecific differences in tolerance to Cu, associated with the level of pollution experienced by the adult source populations. There was also evidence of fitness costs related with increased Cu tolerance. In summary, we have found evidence that the success of invasive species is influenced by chemical contamination. NIS may be advantaged in contaminated estuaries relative to native species and this may assist in their establishment and persistence in new habitats.

The southeastern coast of South Africa is deemed unpolluted and with the exception of a few localized areas, is believed to be pristine. This study aimed to (1) assess the concentration of metals in the water and sharks (Squalus megalops, Mustelus mustelus) from different sites along the southeastern coast, (2) determine the effects that metals have on each other and investigate the accumulation and storage of metals in different shark tissues, (3) determine the effects of length and sex on metal concentration and the presence of metallothioneins in the liver. Metal concentrations in water from seven sites along the southeastern coast revealed concentrations below the detection limit of the extraction method, except for iron at St. Francis Bay which was below the recommended levels as supplied by DWAF. Met al concentrations in S. megalops from the seven sites revealed significantly elevated levels in individuals from Algoa and St. Francis Bay when compared to the other 5 sites. This was observed for all metals except manganese, which had a higher concentration at Cape Agulhas. After comparing the metal levels in muscle tissue (S. megalops) with data from the Department of Health it was concluded that the sharks caught at Algoa and St. Francis Bay were unsuited for human consumption. Liver and vertebral tissues from both S. megalops and M. mustelus were deemed unsuited for human consumption but further research should be conducted especially on vertebral metal concentrations. Comparing the metal concentration in different tissue types of S. megalops and M. mustelus significant difference were observed and attributed to differences in the maturity (thus age) of the two species. Significant differences in the metal concentration of males and females for both the species were observed. A higher number of differences were however observed in M. mustelus than in S. megalops, most probably due to the differences in maturity. Length and weight vs. metal concentration regressions in the males and females of both species revealed more negative than positive trends. This was found to be in accordance with other studies from around the world. Using the life stages of S. megalops, significant differences in the average concentration of metals were observed between the life stages, with embryos usually having the highest concentration. The metal concentration series determined for S. megalops were similar to that which have been reported in the literature in that iron and zinc occupied the highest concentration positions while cadmium, copper and manganese occupied the last three. The spleen usually had the highest metal concentration except for aluminium in skin and arsenic in muscle tissue. Pearson correlation revealed significant negative correlations between cadmium and the mentioned metals suggesting that cadmium replaces essential metals such as manganese, iron and copper as. Negative correlations between essential metals were observed most probably due to the competition for binding positions on ligands. Positive correlations were observed between metals in sp ecific tissue types and may be indicative of synergistic effects, e.g. aluminium may result in the accumulation of other metals such as cadmium. Positive and negative correlations between a specific metal and different tissue types have also been observed and it appears that there are movement of metals between tissue types. Positive correlation for essential metals between yolk and other tissue have been observed, although no correlations for non-essential metals were observed. This could potentially indicate a mechanism that prevents the accumulation of nonessential metals in embryos. Though no metallothioneins were isolated form the liver, the presence of cadmium in other proteins indicate that that cadmium is substituted for essential metals, and as this may result in deleterious physiological effects, further studies should be conducted. In conclusion it can be said that although some of the results were expected there are still quite a few unanswered question and further studies should be conducted.

The southeastern coast of South Africa is deemed unpolluted and with the exception of a few localized areas, is believed to be pristine. This study aimed to (1) assess the concentration of metals in the water and sharks (Squalus megalops, Mustelus mustelus) from different sites along the southeastern coast, (2) determine the effects that metals have on each other and investigate the accumulation and storage of metals in different shark tissues, (3) determine the effects of length and sex on metal concentration and the presence of metallothioneins in the liver. Metal concentrations in water from seven sites along the southeastern coast revealed concentrations below the detection limit of the extraction method, except for iron at St. Francis Bay which was below the recommended levels as supplied by DWAF. Met al concentrations in S. megalops from the seven sites revealed significantly elevated levels in individuals from Algoa and St. Francis Bay when compared to the other 5 sites. This was observed for all metals except manganese, which had a higher concentration at Cape Agulhas. After comparing the metal levels in muscle tissue (S. megalops) with data from the Department of Health it was concluded that the sharks caught at Algoa and St. Francis Bay were unsuited for human consumption. Liver and vertebral tissues from both S. megalops and M. mustelus were deemed unsuited for human consumption but further research should be conducted especially on vertebral metal concentrations. Comparing the metal concentration in different tissue types of S. megalops and M. mustelus significant difference were observed and attributed to differences in the maturity (thus age) of the two species. Significant differences in the metal concentration of males and females for both the species were observed. A higher number of differences were however observed in M. mustelus than in S. megalops, most probably due to the differences in maturity. Length and weight vs. metal concentration regressions in the males and females of both species revealed more negative than positive trends. This was found to be in accordance with other studies from around the world. Using the life stages of S. megalops, significant differences in the average concentration of metals were observed between the life stages, with embryos usually having the highest concentration. The metal concentration series determined for S. megalops were similar to that which have been reported in the literature in that iron and zinc occupied the highest concentration positions while cadmium, copper and manganese occupied the last three. The spleen usually had the highest metal concentration except for aluminium in skin and arsenic in muscle tissue. Pearson correlation revealed significant negative correlations between cadmium and the mentioned metals suggesting that cadmium replaces essential metals such as manganese, iron and copper as. Negative correlations between essential metals were observed most probably due to the competition for binding positions on ligands. Positive correlations were observed between metals in sp ecific tissue types and may be indicative of synergistic effects, e.g. aluminium may result in the accumulation of other metals such as cadmium. Positive and negative correlations between a specific metal and different tissue types have also been observed and it appears that there are movement of metals between tissue types. Positive correlation for essential metals between yolk and other tissue have been observed, although no correlations for non-essential metals were observed. This could potentially indicate a mechanism that prevents the accumulation of nonessential metals in embryos. Though no metallothioneins were isolated form the liver, the presence of cadmium in other proteins indicate that that cadmium is substituted for essential metals, and as this may result in deleterious physiological effects, further studies should be conducted. In conclusion it can be said that although some of the results were expected there are still quite a few unanswered question and further studies should be conducted.

M.Sc. / The increasing demand for water in South Africa arises from a rapid population growth and supporting industrial development. This demand has to be met from limited water resources that are shared by competing user groups and neighbouring states. Deteriorating water quality further limits the sustainable use of water through point and diffuse source pollution of macro constituents, metals and biocides. Metals in the environment can originate as point sources, including industrial and mining effluent, or diffuse sources from geological origin, agricultural activities, acid mine draining and leachate from dumps. Biocides increase crop yields, protect forests and control vectors of serious human disease, but are in the process also carried to water courses. Bioaccumulation studies have been identified as a method to monitor pollutants in the environment. The analyses of data from bioaccumulation studies share a common problem in having to deal with many independent variables, for example variations in time and space, environmental conditions and biological parameters. The objectives of this study were to; investigate the metal levels in water and metal and biocide bioaccumulation in fish tissues from the Berg, Luvuvhu, Olifants and Sabie Rivers; develop a procedure to predict areas and biocides at risk of causing diffuse source pollution and; evaluate the available data and procedures to propose more efficient and cost effective bioaccumulation programmes. The investigation evaluated current and potential contamination of the selected catchments and identified research needs. Fish tissues and water grab samples were collected in the Berg, Luvuvhu, Olifants and Sabie Rivers. The samples were tested for physical variables and analysed for macro constituents and metal levels according to standard analytical procedures. Fish were caught (gill and seine nets) and larger specimens were selected for tissue analyses. Species were identified and standard length, mass, gender, gonadotrophic development index and age indicators were collected. Standard procedures were used to analyse fish tissues for metal levels with the atomic absorption spectrophotometer (AAS) and biocide levels with gas chromatographic methods. A flow diagram of a statistical methodology was developed to incorporate the effects of all biological parameters in the analyses of bioaccumulation data. The procedures used to calculate human health risks are that of the Risk Assistant package reviewed by the United States Environmental Protection Agency (EPA). A Geographical Information System (GIS) model was developed, incorporating the factors affecting runoff of surface water and biocides, to predict areas and variables that could cause diffuse source pollution. The pH values of the rivers studied ranged from of 7.2 to 8.7 except for the Klipspruit River before its confluence with the Olifants River that had a pH of 4.8. Nutrient levels in the Berg River are within the South African guidelines. Nutrients in the Luvuvhu River mostly exceed the guideline values for ortho-phosphate, ammonia nitrogen, nitrate nitrogen and total phosphorus. In the Olifants River the values for ortho-phosphate, total phosphorus, ammonia nitrogen and nitrate and nitrite nitrogen often exceed the South African guideline values. Nutrients in the Sabie River have low concentrations with only ammonia nitrogen exceeding the guideline values. TDS values in the Olifants River is the highest, which is an indication of the high salt loads in the river. The concentrations of various metals in water from the Berg, Luvuvhu, Sabie and Olifants Rivers exceeded the proposed South African and international guidelines.