Abstract
Freshwater systems in South Africa are facing increasing pressures from pollution, including agricultural runoff, domestic sewage, industrial effluents, and various xenobiotics. The Bronkhorstspruit Dam, once considered one of the least impacted systems in the Olifants River catchment, has experienced noticeable ecological decline due to increasing industrial and recreational activities, as well as a severe invasion of water hyacinth (Eichhornia crassipes). This invasive proliferation has disrupted the local water treatment operations and prompted the use of control measures, including the recent glyphosate-based Kilo Max applications by the Bronkhorstspruit Catchment Management Forum (BCMF). Although Kilo Max is regarded as a low-risk herbicide for aquatic organisms, concerns from the BCMF have arisen regarding glyphosate persistence and bioaccumulation, as well as its potential impact on Bronkhorstspruit fish species, since comprehensive fish assessments have not yet been conducted.
Although glyphosate is effective for managing invasive plants, it has been reported to pose physiological and behavioural risks to non-target organisms, including freshwater fish species. Therefore, assessing ecologically important and sensitive indicator fish species following glyphosate spraying is crucial for understanding ecosystem integrity and potential risks to communities that rely on the dam. Given that the Bronkhorstspruit Dam is simultaneously influenced by multiple stressors, including metals reported in previous studies, the lack of histological research data on the health of both past and current fish in the system makes it currently impossible to attribute any observed effects to glyphosate.
Therefore, this study was conducted to provide a comprehensive histology-based assessment of Clarias gariepinus and Cyprinus carpio from the Bronkhorstspruit Dam. To achieve this aim, physical water quality parameters were measured in the inflowing rivers, the dam, and the outflowing river to characterise the environmental conditions of the study sites, then a total of
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40 fish were sampled in one fish sampling trip in December 2024 using gill nets, targeting Clarias gariepinus (n=20) and Cyprinus carpio (n=20). This was followed by standard necropsy procedures, alongside measurements of key blood parameters, including haematocrit and leukocrit, as well as somatic indices and condition factor. Qualitative and semi-quantitative histological evaluations were performed on seven target organs, which included gills, liver, kidney, spleen, heart, brain, and gonads, using standard Haematoxylin and Eosin techniques. Water and fish liver samples were collected for chemical analysis, with inorganic contaminants quantified using Inductively Coupled Plasma–Mass Spectroscopy (ICP-MS), and organic compounds in water and liver samples assessed via Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS). The same procedure was followed when sampling and processing the reference group (20 per target fish species from the UJ Research Aquarium).
The water quality results indicated that most parameters were within acceptable ranges, except for dissolved oxygen saturation at the inflow and outflow of the Bronkhorstspruit River (Site 1 = 67.7% and Site 5 = 61.4%), which were below the recommended 80–120% oxygen saturation. The inorganic chemical analysis of the water revealed that aluminium (Site 2: 0.122 mg/L), copper (Site 2: 0.011 mg/L), iron (Site 2: 0.337 mg/L), and mercury (Site 1: 0.001 mg/L) exceeded the guideline values. The organic chemical detected glyphosate only at Site 1 (0.24 μg/L), while AMPA was detected at Sites 1 (0.38 μg/L), 3 (0.08 μg/L), and 5 (0.06 μg/L). Inorganic chemical analysis detected the most elements in the reference C. gariepinus (31), followed by Bronkhorstspruit C. carpio (30), reference C. carpio (27) and lastly Bronkhorstspruit C. gariepinus (26). Of the 37 inorganic chemicals detected, only Al, Au, B, Ba, Ca, Cu, Fe, K, Mg, Mn, Na, P, Rb, Sr, and Zn were detected in all fish groups. Mercury was detected only in the Bronkhorstspruit C. gariepinus and C. carpio at mean values of 0.205 mg/kg and 0.255 mg/kg. No glyphosate or AMPA was detected in any of the fish groups. Qualitative histological assessment was conducted on both Bronkhorstspruit and reference C.
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gariepinus and C. carpio, with different alterations observed across all organs. The semi-quantitative assessment of the organ indices (Iorg) showed that the liver was the most affected organ, with index values (Iliver) of 7.9 in the Bronkhorstspruit C. gariepinus, 5.7 in the reference C. gariepinus, 4.7 in the Bronkhorstspruit C. carpio, and 4.6 in the reference C. carpio. The reaction pattern (IRP) results revealed that regressive changes (IRC) were the most common response, with index values of 11.9 for reference C, gariepinus, 11.6 for Bronkhorstspruit C. gariepinus, 9.4 for Bronkhorstspruit C. carpio and 9 for reference C. carpio. The mean fish indices (Ifish) showed that Bronkhorstspruit C. gariepinus was the most impacted (22) compared to the reference C. gariepinus (16.8), Bronkhorstspruit C. carpio (13.5), and reference C. carpio (11).
Based on these results, the Bronkhorstspruit Dam is environmentally impacted, with several metals (including Ti, Ni, Ga, As, Rb, Sr, Pd, Ag, Sn, Ba, and Au) found in its water samples; however, only P exceeded the guideline values, while in the liver samples, metals associated with histopathology were detected (Cd, Zn, Cu, Fe, Hg, and Pb), which was a concern. Histological assessments showed alterations in both Bronkhorstspruit C. gariepinus and C. carpio, suggesting a physiological response likely linked to the combined influence of multiple stressors present in the system. However, the semi-quantitative assessment showed that most tissues appeared relatively normal, with minimal changes (Class I), indicating that the fish are in generally acceptable condition. Furthermore, no direct link could be established between the observed histological changes and contaminants found in the Bronkhorstspruit Dam.