Jan G. Myburgh

Responses of phytoplankton upon exposure to a mixture of acid mine drainage and high levels of nutrient pollution in Lake Loskop, South Africa

The relationships between water quality and the phytoplankton community within Lake Loskop were studied during the late summer and autumn of 2008 to evaluate the impacts of acid mine drainage and high nutrient concentrations. The higher concentrations of metal ions and sulphate had adverse effects on certain phytoplankton species in the inflowing riverine zone of Lake Loskop, in comparison to the reference site in the lacustrine zone of the lake, which was dominated by the larger and slower growing late summer species of Coelastrum reticulum Nägeli, Straurastrum anatinum Meyen ex Ralfs and Ceratium hirundinella Müller. The high nutrient concentrations (nitrogen: 17 mg l(-1) and orthophosphate: 0.7 mg l(-1)) during the mid-summer peak of the rainy season were associated with the development of a bloom of the cyanobacterium Microcystis. Water quality data associated with the development of the Microcystis bloom suggest that the aquatic system of Lake Loskop has now entered an alternate, hypertrophic regime. This change overshadowed the adverse effects of high concentrations of heavy metal ions and low pH. Throughout this study, the reference site in the lacustrine zone of Lake Loskop had lower concentrations of metal ions and sulphate, and higher pH values. The response of phytoplankton bioassays on integrated water samples from the different sampling sites did provide potential answers to the reasons for the absence of the algal group Chlorophyceae in the phytoplankton community structure in the riverine zone of the lake.

Identification of toxigenic Microcystis strains after incidents of wild animal mortalities in the Kruger National Park, South Africa

The eutrophic process potentially caused by a high urine and faecal load resulting from an unusually high hippopotamus (Hippopotamus amphibious) density in the Nhlanganzwane Dam, Kruger National Park, South Africa, triggered a chain of events characterised by an increase in the growth of primary producers (Microcystis aeruginosa). This increase in M. aeruginosa biomass was followed by bio-intoxication incidents in wild animals. In this study, we determine if a M. aeruginosa bloom with a total microcystin level of 23,718microgl(-1) have been responsible for mortalities of megaherbivores in the Nhlanganzwane Dam. We further use microcystin molecular markers derived from the mcy gene cluster to identify potentially toxigenic environmental Microcystis strains in the dam during the occurrence of animal intoxications. The estimated total microcystin-LR daily intake by an adult male white rhinoceros (Ceratotherium simum) from cyanobacterial-contaminated water of the dam during the toxic event was an order of magnitude higher (754.29microgkg(-1)bw) in comparison with the lowest observed adverse effecting level (LOAEL) value measured for pigs in a previous study by other authors. In this study the presence of toxic cyanobacterial strains was confirmed with the use of molecular markers that detected the presence of the mcy gene cluster responsible for the production of toxin by M. aeruginosa.

Bioaccumulation of aluminium and iron in the food chain of Lake Loskop, South Africa.

Concentrations of total aluminium (Al) and iron (Fe) were determined in Lake Loskop over a period of four months in 2009 in samples of phytobenthos, phytoplankton, macroinvertebrates, amphibians and fish. The highest concentrations of Al and Fe were measured in the filamentous algae Spirogyra fluviatilis (Hillse) and Spirogyra adanata (Kütz), (Al=18,997.5mgkg(-1) dry weight and Fe=22,054.2mgkg(-1) dry weight) in the riverine zone of the lake with a near-neutral water average pH of 7.3. However, a negative correlation exists between the Al and Fe concentrations measured in the filamentous algae in comparison with the corresponding concentrations of these elements in the water column of the riverine zone. The Al concentrations in the macroinvertebrate families collected ranged from 140.6 to 385.7mgkg(-1) dry weight, with the highest values measured for Al and Fe in the family Gomphidae (385.7 and 1710.0mgkg(-1) dry weight, respectively) in comparison to other macroinvertebrate families sampled. Al and Fe concentrations (2580 and 10,697mgkg(-1) dry weight) in the stomach contents of adult Oreochromis mossambicus fishes were much higher in comparison with adult Micropterus salmoides fishes (98.5 and 439.6mgkg(-1) dry weight), respectively. In all cases of dissected fish species either white or yellow body fat was observed, thus in none of the samples both type of body fats occurred simultaneously. The concentrations of total Al and Fe in the different organs of O. mossambicus were along a mean sequence of intestine>yellow body fat>brain>gills>liver>heart>white body fat, while the mean sequence of total Al and Fe in M. salmoides was: intestine>gills>liver>heart>brain>white body fat. From the levels of Al detected in the yellow body fat of the studied fish species O. mossambicus, we suggest that this phenomenon may be related to the feeding habits of this species. Furthermore, the intake of certain species of phytobenthos by O. mossambicus could have played a role in the bioaccumulation of Al in the food chain and the possible development of pansteatitis in predators at higher trophic levels.

The effect of body size on the rate of decomposition in a temperate region of South Africa

Forensic anthropologists rely on the state of decomposition of a body to estimate the post-mortem-interval (PMI) which provides information about the natural events and environmental forces that could have affected the remains after death. Various factors are known to influence the rate of decomposition, among them temperature, rainfall and exposure of the body. However, conflicting reports appear in the literature on the effect of body size on the rate of decay. The aim of this project was to compare decomposition rates of large pigs (Sus scrofa; 60-90 kg), with that of small pigs (<35 kg), to assess the influence of body size on decomposition rates. For the decomposition rates of small pigs, 15 piglets were assessed three times per week over a period of three months during spring and early summer. Data collection was conducted until complete skeletonization occurred. Stages of decomposition were scored according to separate categories for each anatomical region, and the point values for each region were added to determine the total body score (TBS), which represents the overall stage of decomposition for each pig. For the large pigs, data of 15 pigs were used. Scatter plots illustrating the relationships between TBS and PMI as well as TBS and accumulated degree days (ADD) were used to assess the pattern of decomposition and to compare decomposition rates between small and large pigs. Results indicated that rapid decomposition occurs during the early stages of decomposition for both samples. Large pigs showed a plateau phase in the course of advanced stages of decomposition, during which decomposition was minimal. A similar, but much shorter plateau was reached by small pigs of >20 kg at a PMI of 20-25 days, after which decomposition commenced swiftly. This was in contrast to the small pigs of <20 kg, which showed no plateau phase and their decomposition rates were swift throughout the duration of the study. Overall, small pigs decomposed 2.82 times faster than large pigs, indicating that body size does have an effect on the rate of decomposition.

The post-occipital spinal venous sinus of the Nile crocodile Crocodylus niloticus: its anatomy and use for blood sample collection and intravenous infusions.

The post-occipital sinus of the spinal vein is often used for the collection of blood samples from crocodilians. Although this sampling method has been reported for several crocodilian species, the technique and associated anatomy has not been described in detail in any crocodilian, including the Nile crocodile (Crocodylus niloticus). The anatomy of the cranial neck region was investigated macroscopically, microscopically, radiographically and by means of computed tomography. Latex was injected into the spinal vein and spinal venous sinus of crocodiles to visualise the regional vasculature. The spinal vein ran within the vertebral canal, dorsal to and closely associated with the spinal cord and changed into a venous sinus cranially in the post-occipital region. For blood collection, the spinal venous sinus was accessed through the interarcuate space between the atlas and axis (C1 and C2) by inserting a needle angled just off the perpendicular in the midline through the craniodorsal cervical skin, just cranial to the cranial borders of the first cervical osteoderms. The most convenient method of blood collection was with a syringe and hypodermic needle. In addition, the suitability of the spinal venous sinus for intravenous injections and infusions in live crocodiles was evaluated. The internal diameter of the commercial human epidural catheters used during these investigations was relatively small, resulting in very slow infusion rates. Care should be taken not to puncture the spinal cord or to lacerate the blood vessel wall using this route for blood collection or intravenous infusions.

Ecosystem change and the Olifants River crocodile mass mortality events

Nile crocodile (Crocodilus niloticus) mass mortality events in the Olifants River between the Letaba River confluence in South Africa and Lake Massingir in Mozambique have been attributed to pansteatitis: a disease that affects fat depots of the animals. The disease is also found in sharptooth catfish (Clarias gariepinus) in the same area, and the cause of the disease is attributed to pollution. Although the Olifants River Valley is polluted, the impact of interventions such as dam construction on biodiversity receives little attention. We show that the onset of the pansteatitis epidemic in crocodiles and sharptooth catfish at the Olifants/Letaba confluence coincided with back-flooding of Lake Massingir that changed the Olifants River from a rock and sand substrate river to a clay substrate lake. Isotopic analysis shows that sharptooth catfish shifted from a predominantly vegetarian to a piscivorous diet that is highly correlated with pansteatitis prevalence, and crocodiles and tiger fish (Hydrocynus vittatus) show coincident trophic level increases. The evidence suggests that the ecosystem change altered the structure of the lotic foodweb and that an exotic or extralimital fish has invaded the confluence and is the vector of the pansteatitis epidemic. The invasive fish species is yet to be identified. The pansteatitis epidemic is an unintended ecological consequence of damming this river.

A comparison of in vivo and in vitro assays to assess the toxicity of algal blooms

The toxicity of purified microcystin-LR (MC-LR) and algal material collected during the winter and summer seasons (2005/2006) from the Hartebeespoort dam, South Africa, was investigated using the enzyme-linked immunosorbent assay (ELISA), mouse bioassay, catfish primary hepatocytes (in vitro assay) and protein phosphatase inhibition (PPi) assays. Microcystis aeruginosa, known producer of microcystins, was the dominant cyanobacteria present in the water samples. Exceptionally high cell numbers per millilitre were observed, especially with the summer samples (approximately 1.442 x 10(8)cells/ml), indicating a severe algal bloom in the dam. The toxin concentration as detected by ELISA and PPi assay in the winter and summer extracts was at least 1000 times more than the provisional guideline value (1 microg/l) set by the World Health Organization (WHO) for MC-LR in drinking water. Hepatotoxic effects and death of mice were observed after dosing with the summer extracts, while no hepatotoxic effects were observed with winter extracts. The EC(50) values obtained after exposure of the catfish primary hepatocytes for 72h to MC-LR, winter and summer extracts was about 0.091, 0.053 and 0.014 mg/l, respectively. Similar toxicity results were obtained when the mouse bioassay and primary hepatocytes were used.

Technique for the collection of clear urine from the Nile crocodile (Crocodylus niloticus).

Urine samples can be a very useful diagnostic tool for the evaluation of animal health. In this article, a simple technique to collect urine from the Nile crocodile (Crocodylus niloticus) was described, based on a similar unpublished technique developed for the American alligator (Alligator mississippiensis) using a canine urinary catheter. With this technique, it was possible to collect relatively clean urine samples from Nile crocodiles of different sizes using canine urinary catheters or small diameter stomach tubes. Based on the gross anatomical features of the cloaca of the Nile crocodile, it was confirmed that urine accumulates in a chamber consisting of the urodeum and coprodeum. Faecal material is stored temporarily in the very short rectum, which is separated from the urinary chamber by the rectocoprodeal sphincter.

Linking climate change and progressive eutrophication to incidents of clustered animal mortalities in different geographical regions of South Africa

Cyanobacterial blooms have become an increasing problem in South African freshwater bodies. Since certain species of cyanobacteria are well-known for biosynthesis of potent hepatic and neurotoxins, such blooms can pose a significant threat to the health of animals and humans. The massive proliferation of these organisms in rivers and lakes is largely due to progressive eutrophication. However, a warming trend in the Southern hemisphere, indicated by a threefold increase in the minimum temperature compared to maximum temperature between 1950 and 1990, is likely the cause of the increasing occurrence of toxic cyanobacterial bloom forming species, previously hampered by low water temperatures in different geographical regions of South Africa.

Biotransformation and Oxidative Stress Responses in Captive Nile Crocodile (Crocodylus niloticus) Exposed to Organic Contaminants from the Natural Environment in South Africa

In the present study, the biotransformation and oxidative stress responses in relation to chemical burden in the liver of male and female Nile crocodiles—Crocodylus niloticus—from a commercial crocodile farm passively exposed to various anthropogenic aquatic pollutants was investigated. In general, the data showed that male crocodiles consistently produced higher biotransformation and oxidative stress responses compared to females. Relationships between these responses and concentrations of aliphatic hydrocarbons and polycyclic aromatic hydrocarbons (PAHs) were also observed. Specifically, the catalytic assays for EROD and BROD (not PROD and MROD) showed sex-differences between male and female crocodiles and paralleled immunochemically determined CYP1A and CYP3A protein levels; the relatively similar levels of PAHs in both sexes suggest an estrogen-mediated reduction of this pathway in females. The antioxidant system exhibited higher levels in male crocodiles with slight or significant higher values for catalase (CAT), glutathione reductase (GR), glutathione peroxidases-H2O2 (GPx-H2O2), glutathione peroxidases-Cu (GPx-Cu), total antioxidant capacity towards peroxyl radicals (TOSC-ROO) and hydroxyl radicals (TOSC-HO), total glutathione (GSH) and malondialdehyde (MDA). On the other hand, the activities of acyl-CoA oxidase (AOX) and glutathione S-transferases (GST) were significantly higher in females. Principal component analysis (PCA) produced significant groupings that revealed correlative relationships (both positive and negative) between biotransformation/oxidative stress variables and liver PAHs and aliphatic hydrocarbon burden. The overall results suggest that these captive pre-slaughter crocodiles exhibited adverse exposure responses to anthropogenic aquatic contaminants with potentially relevant effects on key cellular pathways, and these responses may be established as relevant species biomarkers of exposure and effects in this endangered species.