M.Sc. (Aquatic Health) / South Africa’s freshwater ecosystems are under threat due to anthropogenic activities. Recent reports have shown that the Hartbeespoort- (HBPD), Klipvoor- (KD) and Bospoort Dam (BD) impoundments situated in the North-West Province are polluted as a result of mining, industrial and agricultural activities. These impoundments are classified as being in a hypertrophic state. The health status of fish in these impoundments is of concern, and needs to be investigated. For this purpose, Clarias gariepinus and Cyprinus carpio were collected for a histology-based fish health assessment. These two species are present in the selected dams and are some of the preferred fish species for human consumption by the local people. The aim of the study was to determine the health status of these fish species in HPBD, KD and BD using a semi-quantitative histopathological assessment on selected target organs and compare the results to fish from a reference site, the Marico-Bosveld Dam (MBD). Gill nets were used for the collection of specimens. Fish were macroscopically examined, weighed and measured for the calculation of the condition factor. Blood samples were collected for haematocrit, leukocrit and total protein determination. Once sacrificed, a necropsy-based health assessment (HAI) was performed to determine any abnormalities within the visceral cavity. The selected target organs were excised and weighed for the calculation of selected biometric indices. Collected tissue samples were processed using standard histological techniques and the prepared slides were stained using Haematoxylin and Eosin. Macroscopically, a number of fish from polluted sites exhibited liver with fatty change and focal discoloration as well as severe parasites within the visceral cavity. As a result, higher HAI values were recorded in polluted sites compared to the reference site. The haematocrit values varied from normal to below and above the normal range. Leukocrit values were within the normal range except for C. gariepinus from HBPD, which was above the normal range. Total protein values were within the normal range for both species for all sites. The condition factor and hepatosomatic index (HSI) values were within the accepted range except for the HSI of C. gariepinus from MBD. The mean organ index for all study sites fell within class 1 with the exception of the liver index from HBPD, KD and BD which was in class 2. Class 1 (index value <10) is indicative of slight histological alterations while class 2 (index value 10-25) translates to moderate histological alterations. Overall, a higher prevalence of histopathological alterations was noted among fish from polluted sites compared to fish from the reference site. Hence, on the basis of the macroscopic and histopathological results, the fish from the polluted sites were more affected in terms of these parameters compared to fish from the reference site. The results therefore provide valuable information for future bio-monitoring studies in the HBPD, KD and BD impoundments.

M.Sc. (Zoology) / The Vaal River system is constantly subjected to pollution from various industrial, mining and agricultural activities. Consequently, pressure is continually exerted on the health of its fish communities and it was suspected that this may be a contributing factor to the repeated fish kills that have occurred in various sections of the river over the past few years. The aim of this study was to assess the health status of three fish species that were affected by the most recent mass fish mortality in the Vaal River. This was done using a combination of techniques including a histology-based health assessment (HHA) that involved a standard fish necropsy, the evaluation of blood parameters as well as a microscopic assessment of the liver, kidney, gills, and gonads. The histological results were semi-quantitatively assessed. Selected chemical substances were also tested for in water, sediment and the skeletal muscle tissue of the fish. Clarias gariepinus, Labeo capensis and Labeo umbratus specimens were collected from the Vaal River, approximately 2480m below the Vaal Dam (Latitude 26°51'17.47"S; Longitude 28° 7'16.58"E) following the latest occurrence of a fish kill in November 2011. Fish were collected using gill nets. Water was sampled in triplicate at three locations, and in situ measurements were done for selected physico-chemical parameters. Sediment samples were also collected and subjected to a sequential extraction technique and quantification of the heavy metals was done using Inductively Coupled Plasma-Mass Spectroscopy (ICP-MS). Metals in skeletal muscle tissue and water were quantified using the same technique. An analysis of variance (ANOVA), followed by the appropriate post-hoc test was performed to determine differences between means of variables i.e. morphometric, histological, substance concentration and blood parameters...

Ph.D. / The current study evaluated a bio-monitoring technique developed in the USA by Adams, Brown and Goede, 1993. This project was sponsored by the Department of Water Affairs and Forestry (DWAF), to enable testing of the Health Assessment Index (HAI) under South African conditions. Testing took place in the Olifants River system, one of the most polluted river systems . in South Africa. Initially two river points were tested using Oreochromis mossambicus (Robinson, 1996), Clarias gariepinus (Marx, 1996) and Labeo rosae (Luus-Powell, 1997). The current study re-tested the HAI at the same two sample sites, namely Mamba and Balule in the Kruger National Park, using 0. mossambicus and C. gariepinus respectively. Two additional sites were tested in the upper catchment area, namely Loskop Dam and Bronkhorstspruit Dam. The current study further enabled the comparison of HAI results collected during drought and flood conditions. Results obtained after deployment of the HAI were corroborated using chemical analysis of water, sediment and biota. Water and sediment analysis was carried out by the Institute for Water Quality Studies using standard techniques. Bio-accumulation of aluminium, copper, iron, lead, manganese, nickel, strontium and zinc was assessed in the gills, liver, skin and muscle tissue of sample fish using standard Atomic Absorption Spectrometry techniques. Modifications made to the original HAI involved the inclusion of variable ranking in the assessment of fish parasites, with endo- and ectoparasites evaluated separately. Testing of this parasite hypothesis lead to the development of a Parasite Index component to the HAI. Assessment of water, sediment and fish tissue determined that the Olifants River system is indeed exposed to macro and heavy metal pollutants, which negatively affect aquatic health. Constituents posing the greatest threat are chlorides, fluorides, phosphates, total dissolved solids, copper and iron concentrations. Testing the HAI and parasite hypothesis using C. gariepinus, provided the most meaningful results. During testing of the parasite hypothesis both endo- and ectoparasite numbers conformed to the suggested idea that higher endoparasite numbers will occur at highly impacted areas, whereby ectoparasite numbers will be low. This was particularly evident in the lower catchment area, whereby comparisons between drought and flood conditions were carried out. Subsequent decreases in water quality directly after the flood were noted using water and sediment analysis. This observation reflects the results gathered using the HAT and during testing of the parasite hypothesis at all four sample sites. During statistical analysis of the HAI, using logistic regression analysis, parasite numbers, more specifically endoparasite numbers, were the most indicative of fish health. Environmental stressors (flood conditions) result in immunological responses observed in fish, and are reflected statistically using the HAI as changes in WBC %. It is suggested that endoparasites and WBC % provide the best overall assessment of fish condition. These variables should thus not be eliminated, in order to streamline the HAI evaluation procedures. Testing of this bio-monitoring technique under South African conditions provided meaningful results. This indicates that the HAI can be used to assess water quality, with existing water monitoring programmes further benefiting from its incorporation.

PhD / An important component of fish health is an optimally functioning reproductive system. The Luvuvhu River Catchment in the Limpopo Province, South Africa, is a tropical, high-risk malaria area where 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT), an endocrine disrupting chemical (EDC), has been used annually since 1945 as a malaria vector control. DDT is known to affect testes morphology and motility of fish sperm. As such, testicular histology and sperm motility (kinematic) parameters were studied as reproductive indicators of the reproductive capacity for two wild, indigenous fish species (Oreochromis mossambicus and Clarias gariepinus) from the currently DDT sprayed area. Three field studies were carried out over two years (2007 – 2008), including two high flow (HF) periods and one low flow (LF) period [HF 1 (March 07), LF (October 07), HF 2 (February 08)]. Both species were sampled from three sites on the Luvuvhu River for testicular histology and computer assisted sperm analysis (CASA), during all three field studies. The sites included a reference site outside the DDT sprayed area, Albasini Dam (AD), and two exposed sites within the DDT sprayed area, Xikundu Weir (XW) and Nandoni Dam (ND). CASA, based on open-source software, was used for the first time in South Africa to assess sperm kinematic parameters of indigenous fish species in field conditions. These included percent motile sperm (% MOT), curvilinear velocity (VCL μm s-1), velocity of an average path (VAP μm s-1), straight line velocity (VSL μm s-1), linearity (LIN %), progression (PROG μm), and average efficiency (AVE. EFF.). Water and sediment samples were collected during all field studies from the three sites for metal and EDC analysis. Controlled laboratory studies were also carried out on the sperm of both species, externally sourced from aquaculture farms equipped to breed and raise fish in toxicant free water. The laboratory studies involved in vitro exposure of spermatozoa to two different, but environmentally relevant, concentrations of both DDT (DDT 1: 0.27 μg L-1; DDT 2: 0.5 μg L-1) and 1,1-dihloro-2,2-bis(p-chlorophenyl)ethylene (DDE) (DDE 1: 0.11 μg L-1; DDE 2: 1.0 μg L-1) with the aim to provide data to support the possible outcomes found in the field studies using CASA. Furthermore, peroxidation of sperm lipids was assayed by production of malondialdehyde (MDA) after in vitro exposure of spermatozoa to DDT and DDE. DDT and its metabolites were found in varying concentrations in the water from all three sites (0.1 μg L-1 – 1.2 μg L-1). Levels of dieldrin (3.5 μg L-1) and lindane (9.4 μg L-1) residues were also found at XW in HF 2. The histological results revealed alterations to testis tissue of both species at all three sites. The testes were assessed through the identification of alterations and an organ index was calculated: Testes Index (IT). The index is indicative of the histological response in the respective tissue type. O. mossambicus at XW had the highest mean IT value during LF (7.45 ± 5.73) and for all field studies combined (5.47 ± 4.63), primarily due to the occurrence of testicular oocytes (intersex), where the frequency of prevalence was 72.73% and 58.82% respectively. These results were statistically higher than the laboratory control (C) group. The CASA results showed statistical differences primarily for O. mossambicus, where motility parameters were lower at XW when compared to AD. Laboratory exposures found a decrease in sperm motility (% MOT) between the control (C) group and the DDT 1, DDE 1 and DDE 2 exposed groups for C. gariepinus. No significant differences were seen for lipid peroxidation (MDA). On the other hand, no significant differences were seen in CASA parameters between the control and exposed laboratory groups for O. mossambicus, but there was an increase in MDA production from the control to the DDT 1 exposure group.

Durch die Nutzung des fischfutterbedingten Nährstoffeintrags für die kombinierte Produktion von Fischen und Pflanzen können Aquaponiksysteme eine nachhaltige Erweiterung von Kreislaufanlagen der Aquakultur (RAS) darstellen. Herkömmliche Aquakulturfutter zielen jedoch auf Fischproduktion mit geringer Umweltbelastung ab und sind somit nicht für die Aquaponik optimiert. Daher weist RAS-Wasser häufig Mängel im Profil gelöster anorganischer Pflanzennährstoffe auf. So war es Ziel dieser Arbeit, die Auswirkungen unterschiedlicher Proteinquellen auf die Nährstoffdynamik in RAS durch Fütterungsversuche mit Afrikanischem Raubwels und Nil-Tilapia zu untersuchen, bei denen Wachstum, gelöste anorganische Nährstoffkonzentrationen im RAS-Wasser und die Ausscheidung von Nährstoffen über den Kot verfolgt wurden. Der Fokus lag auf nachhaltigen alternativen Proteinquellen zu marinem Fischmehl und terrestrischen Pflanzenproteinen: Larvenmehl der Schwarzen Soldatenfliege (BSFM), Welsschlachtabfallmehl (CM), Geflügelschlachtabfallmehl (PM) und Geflügelblutmehl (PBM). Experimentalfutter, die phosphorreiches PM und CM enthielten, förderten erhöhte Ausscheidung von löslichem reaktivem Phosphor, erzeugten die besten gelösten N:P-Verhältnisse im RAS-Wasser verglichen mit einer Hydroponik-Nährlösung und ermöglichten in Kombination mit PBM besseres Wachstum beim Wels als ein vergleichbares kommerzielles Futter. In Futtern basierend auf einer einzigen Proteinquelle führte PM bei Wels und insbesondere bei Tilapia zu ähnlichem Wachstum verglichen mit marinem Fischmehl, wohingegen BSFM und PBM bei beiden Arten Wachstumsleistung beeinträchtigte. Meta-Analysen aller Versuche legen nahe, dass höherer Phosphor-, Kalium- und Magnesiumgehalt im Futter erhöhte Ausscheidung dieser Elemente in gelöster Form bedingt, was sie zu Zielnährstoffen in Aquaponikfuttern macht, während die Optimierung des Protein zu Energie-Verhältnisses im Futter die gelösten N:P- und N:K-Verhältnisse im RAS-Wasser verbessern kann. / By using the nutrient input from fish feeds for the combined production of fish and plants, aquaponic systems can be a sustainable extension of recirculating aquaculture systems (RAS). However, conventional aquaculture feeds are optimized for fish production and reduced environmental impact rather than aquaponics. Hence, RAS water is often characterized by deficiencies regarding its dissolved inorganic plant nutrient profile. Therefore, this thesis aimed to explore the effect of purposeful dietary protein choice on nutrient dynamics in RAS through four systematic feeding trials involving African catfish and Nile tilapia in which growth performance, dissolved inorganic nutrient concentrations in RAS water and solid fecal nutrient excretion were tracked. Focus was on sustainable alternative protein sources to marine fish meal and terrestrial plant proteins: black soldier fly larvae meal (BSFM), catfish by-product meal (CM), poultry by-product meal (PM) and poultry blood meal (PBM). Experimental diets including phosphorus-rich PM and CM supported increased excretion of soluble reactive phosphorus, produced the most favorable dissolved N:P ratios in RAS water when compared to a renowned hydroponic nutrient solution, and, combined with PBM, enabled better growth performance in African catfish than a comparable commercial diet. In single protein source diets, PM produced similar growth performance in African catfish and particularly Nile tilapia versus marine fish meal, whereas BSFM and PBM impaired growth performance in both species. Meta-analyses covering all trials suggest that higher dietary phosphorus, potassium and magnesium content leads to their increased excretion in dissolved form, making them target nutrients for aquaponic feed formulation, while the optimization of the dietary protein to energy ratio can further improve dissolved N:P and N:K ratios in RAS water.

Nomenclature: Sensitivity to anesthetic clove oil different species of adult fish Sensitivity of breeding fishes of various species (Siberian sturgeon, Brook trout, Black carp- young breeding fishes) for anesthetic clove oil was probed by the tests performed at temperature in the period of stripping (apart from Brook trout and Black carp). The tests were carried out with 17 fish species- with cyprinids (Ide- golden form, Common dace, European Chub, Sneep, Barbel, Grass carp, Black carp, bighead carp), salmonids (Brown trout, Rainbow trout, Brook trout), thymallinae (Grayling), pikes (Northern pike), perches (Pike-perch), sturgeons (Siberian sturgeon, Beluga) and airbreathing catfishes (North African catfish). Using concentration of 0,03ml.l-1 (Brown trout, Rainbow trout, Brook trout, Grayling, Pike, Pike perch, Ide, Dace, Chub, Barbel and Sneep), 0,04ml.l-1 (Grass carp, Black carp, Bighead carp), 0,05ml.l-1 (North African catfish) and 0,07ml.l-1 (Siberian sturgeon, Beluga) the IIb. Phase has been achieved, which is sufficient for manipulation with fishes during stripping. The anesthesia subsided after time period of 10-17 minutes (12 species) at the majority of fishes and at others after interval of 20-26 minutes (5 species). The results confirmed concentrations recommended for salmodis and Grayling (0,025-0,03ml.l-1), sturgeons (0,07ml.l-1), and North African catfish (0,05ml.l-1). At the majority of cyprinids (apart from Grass carp, Black carp and Bighead carp), the concentration used was lower than recommended (0,04-0,05ml.l-1)

Ph.D. (Zoology) / Histomorphology as a tool to assess the health status of five economically important fish species in the Okavango Delta Panhandle, Botswana the Okavango Delta, and an inland freshwater wetland of national and international importance was designated by the Ramsar Convention in 1996. This site is of global importance because of a wide diversity of fish species found in this freshwater ecosystem. The Okavango Delta is also one of the last 'near pristine' sites on the African continent and therefore serves as an ideal reference site to describe the normal histomorphology of selected organs in sentinel fish species which could be used as reference for future toxicological studies. The aim of this study was to assess the health status of five economically important fish species namely, Clarias gariepinus (Burchell, 1822), Clarias ngamensis (Castelnau, 1861), Serranochromis robustus jallae (Gunther, 1864), Serranochromis angusticeps (Boulenger, 1907) and Hydrocynus vittatus (Castelnau, 1861) sampled from the Okavango Delta Panhandle, by describing the normal histomorphology of the liver, gills and gonads of the sentinel species. The sentinel fish species are of significant importance because they form an integral part of the small-scale commercial, subsistence and recreational fisheries in this ecosystem. In the past few decades, fish-health research has focused almost exclusively on histopathological alterations resulting from pollution caused by exposure to chemicals, parasitesand pathogens. Limited attention has been devoted to the normal histomorphology and health status of fish species in various aquatic ecosystems including the Okavango Delta Panhandle. Also, histopathological evaluations noted only the presence of abnormalities without quantitative assessment of the extent of the abnormalities within each tissue section. The absence of baseline normal histomorphology and numerical data however, makes it difficult to establish cause-and-affect relationships between pathology and contamination patterns in various fish species. It is imperative therefore, that the normal histomorphology of organs in fish species occurring in this region should be known so that histopathological conditions maybe accurately diagnosed…