Biological Sciences

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Ecology of some mammals in relation to conservation management in Giant's Castle Game Reserve.
(1982) Rowe-Rowe, David Treloar.; Meester, Jurgens Anthonie Janse.
Abstract available in PDF.
A microbal study of water quality in the marine environment off Durban : 1964-1988.
(1989) Livingstone, Douglas James; Contrafatto, Giancarlo.
wastewater from the harbour mouth with the outgoing tides, while the discharge from a sewer on the Bluff into the surf-zone amounted to 20 x 103 m3 /day. In addition, there existed more than 90 beach pipes and storrnwater drains (not all of them legal) , about one third of which carried contaminative material on to the beaches and into the surf. Twenty-eight sampling stations were established between the Mgeni River and Isipingo and subjected to detailed bacteriological surveillance, prior to the construction of a pair of submarine outfalls to serve the region's disposal requirements . A microbial system of evaluating seawater quality was developed using Escherichia coli I, parasite ova, staphylococci, salmonellae (including Salmonella typhi) and the salinity as indicators. A comprehensive "before" picture was therefore created against which to measure future changes in the sea off Durban. In 1968/69 the pair of submarine pipelines was commissioned with their attendant treatment plants. The harbour effluent was diverted to the new complex , and pollution from the minor outfalls was progressively halted with their wastes similarly joined to the new works. The system of water quality gradation was applied to the surf-zone and out to sea to measure the efficacy of the new pipelines, providing an "after" picture . Throughout the subsequent engineering innovation of sludge disposa l via the outfalls (which proved successful), and during climatic extremes involving a severe drought (with stringent water consumption restrictions), cyclones and catastrophic floods, the classification system continued to function satisfactorily , covering 25 years in all: alterations in the water quality were shown to be invariably a consequence of changes effected upon the shore or meteorological events. The system has also proved useful in identifying and measuring the impact of contaminative foci in Cape waters and at Richards Bay. The relevant oceanography and current dynamics, the rationale for the selection of the indicators used and the methodology, along with more general aspects of marine pollution and associated risks are discussed. Finally, the feasibility is examined of curtailing the numbers of parameters measured and simplifying the classification system while retaining its usefulness and serviceability as an instrument for assessing the impact of domestic effluent on the marine environment off Durban.
Systematic revision of Southern African species in the genera Eptesicus, Neoromicia, Hypsugo and Pipistrellus (Chiroptera: Vespertilionidae).
(2005) Kearney, Teresa Catherine.; Contrafatto, Giancarlo.; Taylor, Peter John.
Abstract available in PDF file.
Studies on the biology, fungal contamination, and conservation of seeds of Welwitschia mirabilis Hook. fil.
(2006) Whitaker, Claire Rosalie.; Pammenter, Norman William.; Berjak, Patricia.
Abstract available in PDF file.
Grape fruit juice improves glycemic index and up-regulates expression of hepatic organic cation transporter protein (OCTI) in the rat
(2009-03) Owira, Peter Mark Oroma.; Ojewole, John Akanni Oluwole.
Abstract available in PDF file.
The financial and economic feasibility of biodigester use and biogas production for rural households.
(2011) Smith, Michael Trevor.; Blignaut, James N.; Goebel, Jessica Schroenn.
In South Africa, sustainable development is set in the context of two separate economies. The second of these economies consists of the rural population and is characterised by poverty and stagnant development. Sustainable development is an increasingly topical concept which highlights the need for development to proceed in a manner that does not deplete natural resources. In addition to narrowing the gaps between the various classes (layers) in an economy, the key ‘ingredients’ of sustainable economic development include “natural resource management, food, water, and energy access, provision and security” (Blignaut, 2009: cited in Blignaut and van der Elst, 2009: 14). A biodigester is a potential solution to some of the difficulties faced by remote rural populations. Biodigester systems are submerged tanks capable of producing a nutrient rich fertiliser and combustible gas when consistently fed with organic matter and water. A biodigester may be one simple answer to the key ingredient needs of sustainable development – reducing the depletion of natural resources, providing clean burning energy for cooking and fertiliser for growing food. The potential is clear for biodigesters to aid in the process of sustainable development. The question to be analysed is whether this technology would be financially and economically feasible for installation and use in rural households. This thesis focuses on a typically remote and rural community in KwaZulu-Natal, South Africa, in order to assess the potential feasibility of a biodigester system. The appraisal takes the form of a Cost Benefit Analysis (CBA) and aims to establish whether or not this technology is financially feasible for individual rural households and/or economically beneficial to society.
Water stress effects on growth, development, resource capture and resource use efficiency of two contrasting sugarcane genotypes.
(2014) Ngxaliwe, Sivuyile.; Singels, Abraham.; Eksteen, Alana.; Pammenter, Norman William.
Abstract available in PDF file.
The challenges and opportunities in conserving wide-ranging cross-border species : a case study of the Greater Mapungubwe Transfrontier Conservation Area elephant population.
(2015) Selier, Sarah-Anne Jeanetta.; Slotow, Robert Hugh.; Page, Bruce Richard.; Di Minin, Enrico.
Transfrontier conservation areas potentially play a key role in conserving biodiversity and promoting socioeconomic development. However, socio-political factors often affect their effectiveness in achieving biodiversity conservation and sustainable development objectives. Following a transdisciplinary approach, I assessed the challenges and opportunities in conserving and managing the African elephant (Loxodonta africana) population within the Greater Mapungubwe Transfrontier Conservation Area (GMTFCA) in Botswana, South Africa and Zimbabwe, southern Africa. The results showed that the current rate of offtake of bull elephant in the GMTFCA is unsustainable. At current rates of hunting, in fact, trophy bulls were predicted to disappear from the population in less than 10 years. Elephant densities were higher in South Africa and Botswana where the gross domestic product is higher. In addition, elephant densities were higher at sites where the proportion of agricultural land around them was the lowest and where vegetation productivity was the highest. Trophy hunting, as well as other localised human activities, also affected the distribution of elephant within sites, forcing them to trade-off between disturbance avoidance and the availability of food and water. While at the international level, a significant body of law and policy relevant to elephant conservation exists, I found that there was little cooperation among Botswana, South Africa and Zimbabwe, and a lack of implementation of these provisions on a national and trilateral level. Overall, this study confirmed that poverty was an important factor affecting elephant abundance at the country level, but highlighted that, at the site level, anthropogenic disturbance played a crucial role. A revision of the current hunting quotas within each country and the establishment of a single multi-jurisdictional (cross-border) management authority regulating the hunting of elephant is needed. Further, to reduce the impact of increasing human populations and agricultural expansion, the development of coordinated legislation and policies to improve land use planning, and the development of conservation corridors to link current protected areas, is needed. The issues regarding the management of this elephant population illustrate the significant challenges involved in achieving a comprehensive, consistent and effective implementation of a transboundary population approach. Southern African countries make an important contribution to elephant conservation and could soon become the last stronghold of elephant conservation in sub-Saharan Africa. Therefore, immediate actions are needed to reduce pressures from human activities in order to enhance the long-term persistence of the species.
Prevalence and genetic relatedness of Besnoitia besnoiti isolates from different geographical regions of South Africa.
(2015) Malatji, Mokgadi Pulane.; Mukaratirwa, Samson.; Chitanga, Simbarashe.
Bovine besnoitiosis is a protozoan disease caused by an apicomplexan parasite Besnoitia besnoiti and is reportedly re-emerging in Europe and occurring in many other countries including South Africa. The disease has long been neglected and has only recently started getting attention due to its increasing geographical distribution. This parasite causes significant economic losses due to reduced body condition, declined milk production, irreversible sterility in males and mortality. This study was conducted to determine the prevalence of Besnoitia besnoiti infection in cattle, as well as to establish the phylogenetic relationship among parasite isolates in different geographical regions of South Africa, where the disease was previously reported. A total of 688 cattle (688 blood and 376 skin samples) were randomly sampled from farms located in Limpopo, Gauteng, KwaZulu-Natal and Eastern Cape provinces of South Africa. Based on the analyses of DNA sequences of the nuclear ribosomal internal transcriber spacer 1 (ITS1), it was observed that 15.7% (108/688) of the sampled animals were positive, with 5.3% (20/376) and 14.4% (99/688) of the animals being positive on skin and blood samples, respectively. 2.9% (11/376) of the animals were positive on both blood and skin samples. The difference between in prevalence between the areas sampled were not significant, χ² = 0.263. The parasite was more prevalent in communal farms (30.8 %) and in exotic breeds (35.3 %) than local or mixed breeds, and the difference in prevalence between farming and breed type were not significant (χ² = 0.199, χ² = 0.227 respectively). Aligned sequences were analysed by Maximum parsimony, neighbour joining and maximum likelihood and phylogenetic analysis of the isolates was carried out. Results showed that, based on the ITS1 region, our isolates were closely related to the wildebeest strain which is currently used for the manufacture of the vaccine, forming a clade which is separate from the European strains. One of the isolates from Gravelotte, Limpopo province, was closely related to the European strains, forming a sister clade for the European strains from GenBank. This is the first report on molecular characterisation of the parasite in South Africa and will aid in disease surveillance and control in the studied areas.
A phylogeny of South African east coast intertidal rocky-shore Polychaete worms (Annelida) and the genetic structure and demographic history of an example, Marphysa corallina.
(2015) Kara, Jyothi Ashok.; Macdonald, Angus Hector Harold.
The Annelida is an evolutionarily ancient invertebrate taxon. Recent studies have found that the formerly described sister taxon of the Polychaeta, Clitellata, is a derived Polychaete group thus making Polychaeta a paraphyletic group. Polychaete worms represent one of the most diverse invertebrate groups and are well represented in a variety of environments such as temporary freshwater puddles, rocky intertidal shores, estuaries and the abyssal plain. Polychaetes are fundamentally important in their environments as many are regarded as ecosystem engineers. Phylogenetic relationships within the Polychaeta are poorly understood and some species level classifications are uncertain due to the large number of polychaete worms present. In Chapter two, the phylogenetic relationships within the commonly found polychaete families (Nereididae and Eunicidae) were analysed using the universal mitochondrial cytochrome oxidase subunit 1 (COI). Within Eunicidae, analyses supported a polyphyletic Marphysa and Eunice which is consistent with previous results as individuals from both genera are nested among one another. Within Nereididae, relationships between genera and species were poorly supported and complex. Genera did not form exclusive clades but instead grouped with one another. A large degree of homoplasy has been recorded for the family which could have attributed to the convoluted groupings. Thus it has been suggested that genera from both Eunicidae and Nereididae be revised. Marphysa corallina is a poorly studied Eunicid polychaete which has a tropical indo-west distribution. It was observed to be a common worm among others on the intertidal rocky shores of KwaZulu-Natal and the Eastern Cape. In Chapter three, the population genetic structure and demographic history of M. corallina was investigated using two genes: universal mitochondrial cytochrome oxidase subunit 1 (COI) and the nuclear intron spacer region (ITS1). Diagnostic taxonomic characters were used to identify and validate the specimens as Marphysa corallina. The COI marker revealed that populations were highly connected to one another and formed a large panmictic population whereas ITS1 showed shallow genetic structuring of populations. Family Eunicidae individuals are known to lack a long lived planktonic larval stage which could not have contributed to panmixia as demostrated by the COI marker. Demographic results indicated that populations had recently undergone sudden expansions which could have falsely resembled highly connected populations. Estimation of divergence times places the expansions in the mid to late Pleistocene. Populations had not reached migration-drift equilibrium thus contemporary population distributions of Marphysa corallina along the east coast of South Africa are largely shaped by past climatic events such as in the Pleistocene.
Socio-economic and environmental deteminants of malaria in four malaria endemic provinces of Zambia.
(2015) Shimaponda, Nzooma Munkwangu.; Mukaratirwa, Samson.; Gebreslasie, Michael Teweldemedhin.; Tembo-Mwase, Enala.
A large fraction of the global malaria burden occurs in sub-Saharan Africa and its endemicity depends on the interaction of environmental factors, vectors, parasites and the host. In Zambia, the negative effect of the break in interventions experienced in the late 2000s varied by regions. Therefore, it was necessary to determine the malaria determinants through the study of: statistical models that have been employed; knowledge of the community in malaria management and control; prevalence of malaria and presence of social and community-related factors influencing malaria control in selected communities; contribution of other social and environmental determinants of malaria from the household point of view; and also socio-economic and climatic determinants of malaria at provincial level, in Zambia. This work was achieved through a number of methods beginning with a systematic review of studies that have identified socio-economic and eco-environmental determinants of malaria through the use of statistical models in malaria burden determination and prediction in southern Africa. We also conducted a cross-sectional survey employing a simple random sampling technique to administer questionnaires to 584 household heads from selected communities, on the following components: knowledge, attitude and practices in malaria control; the role of social and community-related structures in malaria burden and control; and water sources and practices as well as housing structures in relation to self-reported malaria infections. Malaria testing was also performed using a rapid diagnostic test (RDT) in 756 individuals sampled from the 584 households. The household-level data was analysed in STATA and WinBUGS whereas the provincial-level malaria cases, government socio-economic and remotely-sensed climatic data were analysed in STATA, WinBUGS and also in R- integrated Nested Laplace (R-INLA) The focus of the studies conducted in southern Africa reviewed, has mainly been on malaria determinants related to intervention strategies and climatic factors. Additionally, the use of Bayesian statistical modelling was quite low (29.2%) in the studies reviewed. The community knowledge study showed that although knowledge levels in malaria were high they were not interrelated with attitudes and practices. In the malaria testing survey, a higher infection rate was seen in children and the highest RDT malaria prevalence was recorded in communities of Luapula province. Relating malaria burden with the role of community health workers (CHWs) in malaria control, malaria prevalence was inversely related with CHWs presence in Western Province. On the other hand, relating malaria burden with water practices and housing structures, “river” as a water source was the main predictor. The Bayesian hierarchical (or Generalised Linear mixed model) and R-INLA based models showed that region on one hand and region, time and precipitation on the other, were strong predictors of malaria incidence. More research in the area of statistical modeling as well as in other areas such as behaviour change, strengthening of existing CHW and exploring new avenues with regards to community social structures and ecological and climatic factors by locality is a great need.
Mapping the expression of key functional genes associated with murine corneal endothelial development.
(2015) Bengtson, Michel Leigh.; Sommer, Paula.
The cornea is a complex, layered, transparent structure that is responsible for transmitting and refracting light to ensure vision. Of all the layers, the corneal endothelium is physiologically the most important layer. Its principle function is to regulate deturgescence, a requirement for corneal transparency. Corneal deturgescence is regulated by key functional genes that include a network of tight junctions, water channels and ion pumps. In this study, we created an in vitro model of murine corneal endothelial development by immortalising endothelial cells at significant stages in corneal endothelial morphogenesis. Primary cultures of presumptive corneal endothelial cells were immortalised from wild-type mouse embryos at embryonic day (E) 14.5 and E15.5, and mature corneal endothelial cells were immortalised from wild-type pups at post-natal day (P) 13. These cell lines, together with already established cell lines at earlier stages in corneal endothelial development (E12.5 and E13.5), were used to map the expression patterns of three functional genes associated with corneal endothelial function, Zo1, Aqp1 and Slc4a4. Zo1 is a tight junction protein that facilitates the formation of the resilient monolayer that is characteristic of the corneal endothelium. Aqp1 is a water channel protein that mediates transcellular fluid transport while Slc4a4 is an ion pump that transports cellular bicarbonate ions across plasma membranes. Aqp1 and Slc4a4 are disease target genes which is indicative of their functional significance in corneal endothelial physiology. Quantitative real-time PCR (qPCR) and western blot analysis showed that Zo1 expression at both mRNA and protein level was progressively down-regulated from E12.5 to E15.5, after which it was up-regulated at P13 (p˂0.05 relative to E15.5 at mRNA level). Aqp1 gene expression was up-regulated at E13.5, with subsequent down-regulation at E14.5. Thereafter, Aqp1 gene expression was up-regulated at E15.5. Interestingly, Aqp1 gene expression was significantly down-regulated at P13 when the endothelium is completely matured (p˂0.05 relative to all other cell lines). Western blot analysis demonstrated Aqp1 expression throughout development in this model. Densitometric analysis showed that glycosylated Aqp1 protein expression levels decreased gradually from E12.5 to P13. Unglycosylated Aqp1 protein expression was only present at E12.5 and E13.5, and was up-regulated at E13.5. Slc4a4 was expressed at low levels during corneal endothelial development. However, it was highly expressed at P13 (p˂0.05 relative to all other cell lines). Protein expression for Slc4a4 could not be determined due to the lack of a commercially available antibody specific for murine Slc4a4. Confocal microscopy analysis demonstrated the protein localisation patterns for Zo1 and Aqp1 in monolayer and hanging drop culture, mimicking the 3D environment in which corneal endothelial development occurs. The results of this study provide valuable insight into the expression patterns of key functional genes associated with corneal endothelial development.
Spatio-temporal analysis of woody vegetation data at various height classes within a semi-arid savanna : insights on environmental drivers, distributions and dynamics.
(2015) Scholtz, Rheinhardt.; Kiker, Gregory Alan.; Scharler, Ursula Michaela.; Mwambi, Henry Godwell.; Smit, Izak P. J.
Savanna vegetation is regulated by bottom-up (e.g. soil and rainfall) and top-down (e.g. fire and herbivory) factors. Little is known about how these factors influence biodiversity at regional scales, particularly spatial patterns of woody plant cover, species richness, distribution and composition at various height classes. Vegetation studies are often restricted to hillslope or catchment-level scales due to time and/or logistical constraints, however there is a need to understand patterns of vegetation dynamics at regional scales as well. The main aim of this thesis was to better understand the factors (bottom-up and top-down) that influence vegetation structural dynamics in terms of species richness, distribution, composition and woody cover at regional scales. Using a rapid data collecting technique within n>1800 field sites across the entire Kruger National Park, South Africa, woody species were identified and categorized into one of three height classes; shrub (0.75m – 2.5m), brush (2.5m-5.5m) and tree (>5.5m). From this, several vegetation attributes were collated; such as species composition, richness and woody cover at each height class. A range of covariates related to disturbance (i.e. fire frequency and elephant density), landscape (e.g. aspect, altitude) and climate (temperature, rainfall) were used to identify which covariates were significantly associated with each attribute of interest. A primary finding of this research highlights the importance of bottom-up factors, such as geology and rainfall, for woody vegetation spatial distribution at regional scales. However, depending on the response variable (e.g. species richness, woody cover, species distribution), the various height classes were not influenced equally by each specific environmental factor. Overall, both climatic and non-climatic factors were highly associated with species distribution and richness, but top-down factors such as fire and herbivory were significantly associated with maintaining plant structural variation. This influenced species community composition but not species richness at each height class. Similarities in composition were related to degree of disturbance rather than the disturbance alone at specific height classes. Therefore, while species distribution and woody cover were spatially regulated by specific climatic and non-climatic factors, top-down disturbances were essential in preserving structural heterogeneity. Future vegetation monitoring campaigns in disturbance-driven systems should be cognizant of the importance of vegetation structure.
The effect of wastewater treatment works on foraging ecology, haematology, detoxification organs and reproduction in an urban adapter, the banana bat (Neoromicia nana).
(2015) Naidoo, Samantha.; Schoeman, Marthinus Cornelius.; Mackey, Robin L.; Vosloo, Dalene.
Natural land is rapidly becoming urbanized. Wastewater Treatment Works (WWTWs) are a ubiquitous component of this urban landscape. WWTWs may provide profitable foraging areas for insectivorous bats because of their association with a high abundance of pollution-tolerant chironomid midges (Diptera). However, bats that feed on these insects may also accumulate metal pollutants in their tissues, with acute or chronic effects on their health. There have been no studies to investigate whether African bats utilize these WWTWs as foraging grounds, and the potential physiological impacts from foraging at such sites. The aim of this study was to investigate the impact of WWTWs on foraging ecology and multiple tiers of physiology (haematology and genotoxicity, detoxification organs and reproduction) in an urban adapter, the banana bat (Neoromicia nana, family Vespertilionidae) in KwaZulu-Natal, South Africa. N. nana exhibited a significantly higher abundance and feeding activity at wastewaterpolluted sites than at unpolluted reference sites. Additionally, the most abundant insect order at wastewater-polluted sites and in the diet of resident bats was Diptera, compared to a diverse insect diet at unpolluted sites. Thus, WWTWs provide an optimal food resource to bats in the short-term. However, I found significantly higher levels of essential and non-essential metals at WWTW-polluted sites, and in the tissues of WWTW bats than at unpolluted sites. Further, I found sub-lethal haematological and genotoxic responses related to increased metals in WWTW bats. Specifically, N. nana at WWTWs had significantly lower antioxidant capacity and significantly higher levels of DNA damage and haematocrits than bats from unpolluted sites. An accumulation of DNA damage, especially from double-stranded breaks ultimately leads to tissue damage and disease. These longer-term effects of chronic pollutant exposure should be most evident in the organs involved in detoxification, the liver and kidneys. Indeed, I found evidence of disrupted balance of essential metals and mineral nutrients, histopathological tissue damage and whole organ effects in the liver and kidneys. Finally, I found reproductive system alterations in male N. nana at WWTWs. Although I did not find significant effects on the sex organs, testosterone hormone concentrations were significantly lower in male N. nana at WWTWs than in males from unpolluted sites. In addition, body condition indices for N. nana from the WWTWs were significantly lower than at unpolluted sites, suggesting lower quality male bats at WWTWs. Taken together, these results suggest the potential for serious long-term health risks, negative fitness implications and ultimately, population effects for these top predators within the urban landscape.
Efficacy of maslinic acid and chloroquine on the co-infection of Plasmodium berghei and Trichinella zimbabwensis in Sprague-Dawley rats.
(2015) Gcanga, Lorna.; Mukaratirwa, Samson.; Masola, Bubuya.
Many people living in Africa and elsewhere in the tropics suffer from combined malaria, soil-transmitted infections and tissue-dwelling nematodes. Most of these deaths are prevalent in sub-Saharan Africa (SSA) where the infections overlap, which often results in co-infection. Malaria and trichinellosis are one of the most important zoonotic diseases especially in sub-Saharan Africa caused by Plasmodium spp. and Trichinella spp., respectively. New drugs targeting malaria and trichinellosis have been examined with little success. The aim of our study was to determine and compare the efficacy of maslinic acid and chloroquine on the co-infection of muscle-dwelling larvae of Trichinella zimbabwensis and Plasmodium berghei in rats. Fifty-four Sprague-Dawley rats with an average weight of 150g and 200g for males and females respectively were infected with T. zimbabwensis and P. berghei. Infected rats were randomly assigned to nine groups which were subjected to treatments of maslinic acid and chloroquine and a combination of maslinic acid and chloroquine. Co-infected groups were infected with T. zimbabwensis on day 0, and then infected with P. berghei on day 30 post-infection (pi). Treatment was administered for 3 consecutive days on day 9 pi with P. berghei. Groups infected with P. berghei only were infected on day 0 and were treated on day 9 pi for 3 consecutive days. Groups infected with T. zimbabwensis only were infected on day 0 and treated on day 25 pi. Untreated control groups were a placebo (distilled water) on day 25 pi infected with T. zimbabwensis and from day 9 pi infected with P. berghei. In Trichinella-infected groups, the efficacy of each treatment measured by the rate of the reduction in muscle larvae was significant (P < 0.05) for both drugs compared to the untreated control group. In malaria-infected groups, the efficacy of each treatment, measured by the rate of reduction in parasitaemia, was significant (P < 0.05) for both drugs compared to the untreated control group. There was no apparent synergistic effect due to the combination of the two drugs in reducing the muscle larval burden and in reducing malaria parasitaemia. In all the treatment regimens, the reductions were significant when compared to the untreated control groups and not significant to each other (P > 0.05). From these results we can conclude that the efficacy of maslinic acid on the co-infection of T. zimbabwensis and P. berghei was comparable to that of chloroquine, making maslinic acid a promising drug to be used as an anthelmintic and anti-malaria against muscle larval stages of Trichinella spp. and malaria parasitaemia and no side effects were observed.
Technique establishment for in vitro selection of drought tolerant sugercane genotypes.
(2015) Mhlanga, Philisiwe Felicity.; Watt, Maria Paula Mousaco Deoliveira.; Snyman, Sandra Jane.
There is a need to have constant supply of sugarcane varieties adapted to different South African regions. However, the genetic improvement and selection of sugarcane cultivars with superior traits, e.g. drought tolerance, are difficult due to its complex polyploid and aneuploid genome. Biotechnology approaches are being investigated for the selection and/production of drought tolerant cultivars. Towards this end, the aim of this study was to establish: 1) the best source of meristematic explant for initiation and mass propagation of in vitro shoots; 2) in vitro conditions to screen and select for drought tolerance; and 3) physiological parameters as indicators of drought tolerance in vitro. Sugarcane stalks and shoots from single-budded setts of NCo376 were used. From the former, 1.3 cm-long meristems were isolated and used for shoot induction, shoot multiplication and rooting. The single-budded setts (approx. 50 mm) were first germinated in 20 ml sterile water or sterile moist paper, resulting in 100% and 60% sett contamination, respectively. With 1 mg lˉ¹ methylene blue (MB) there was 30% sett contamination, whilst 1 mg lˉ¹ MB in combination with 1 ml lˉ¹ Previcur® or 1 ml lˉ¹ BRAVO® resulted in 40% and 7% contamination, respectively. The uncontaminated germinated shoots (approx. 1 - 2 cm) were excised after 10 days in culture and used as the other source of meristems. Meristems from both sources were multiplied and rooted in vitro and their plantlet yield was 60 shoots/sugarcane stalk meristem and 10 shoots/meristem from in vitro-germinated sett. NCo376 and N41 varieties were used to determine the effect of mannitol (204, 326, 448 and 569 mM) on in vitro plantlet shoot and root re-growth. For both, increased mannitol in the media delayed shoot and root re-growth, with NCo376 being affected first. Stress was more significant on root than on shoot re-growth. For NCo376 plantlets, there were significant differences in root re-growth between 0 and 204 mM and the other tested treatments. For N41 plantlets, % root re-growth at day 10 on 569 mM mannitol was significantly higher than that at the other treatments. At 4 – 10 days, % shoot re-growth of NCo376 on 0 and 204 mM mannitol was greater than that at 326, 448 and 569 mM mannitol. Similar results were observed with N41 plantlets. The LD₅₀ and LD₉₀ for mannitol were 332 and 606 mM for NCo376, and 851 and 1493 mM for N41. There was no differences between the effects of polyethylene glycol-6000 (PEG-6000) and mannitol on root re-growth at the same osmotic potential. However, PEG-6000-containing cultures required to be aerated. As at 87 mM PEG-6000, NCo376 plantlets showed 50% root re-growth compared to 10% in non-aerated cultures, mannitol was used in subsequent investigations. Mannitol concentrations equivalent to LD₅₀ and LD₉₀ for NCo376 and N41 were used to screen N12, N36, N19 and N26 varieties. Based on the results obtained, the varieties were ranked on their tolerance to mannitol stress: N41 > N26 > N36 > N12 > N19 > NCo376. Leaf electrolyte leakage, leaf chlorophyll content measured with Soil Plant Analysis Development (SPAD) measurements, and histochemical detection of hydrogen peroxide (H₂O₂) (with nitroblue tetrazolium) and superoxide anion (O₂⁻˙) (with 3, 3’-diaminobenzidine) production were evaluated as indicators of stress using N41, N26, N19 and NCo376 on 332, 606 or 851 mM mannitol. N19 and NCo376 plantlets on 332 mM mannitol showed a higher % electrolyte leakage at day 5 (70%) than at day 10 (40 – 50%) of culture than N41 and N26 plantlets. A slight decrease in chlorophyll content was recorded at day 10 of culture in 332 and 851 mM mannitol, with no differences between NCo376 and N19, and N41 and N26. NCo376 and N19 accumulated more H₂O₂ than N41 and N26. O₂⁻˙ accumulation was also greater in NCo376 and N19 than in N41 and N26. All these parameters detected stress at lower levels of mannitol (332 and 606 mM), but not at 851 mM. It was concluded that mannitol stress in vitro (332 – 606 mM), in combination with the physiological assays allow for the discrimination of in vitro osmotic stress among sugarcane varieties. Further work is necessary before recommendations can be made regarding the use of the other stress biomarkers.
Lichen photobiology in relation to climate change : protection in peltigeralean lichens against excess ultraviolet (UV) radiation using induced melanins and the effects of UV on melanin synthesizing enzymes.
(2015) Matee, Lusanda Patrick Herbert.; Beckett, Richard Peter.
Depletion of the stratospheric ozone has allowed more solar ultraviolet (UV) radiation (100-400 nm) to reach the lower atmosphere. Despite the success of the continued implementation of Montreal protocol of 1987 and its amendments aimed at the protection of the ozone layer, it is possible that UV will remain elevated in some regions due to decreasing removal by clouds and aerosols. Strong interactions between ozone depletion and changes in climate induced by increasing greenhouse gases could result in increased UV radiation levels reaching the surface. Increased exposure to UV-B can cause significant damage to organisms including protein damage, inhibition of photosynthesis, formation of high energy reactive oxygen species (ROS), disruption of nitrogen fixation and induction of cell mutation and inhibition of growth. Lichens are composite organisms that arise from a symbiotic association between fungi and photoautotrophic algal partners, which may be microalgae and/or cyanobacteria. They have developed several protective strategies to protect themselves from excessive radiation. Lichens such as Cladonia rangiferina, Lobaria pulmonaria and Umbilicaria rossica are known to synthesize melanins in response to elevated levels of UV-B radiation. Synthesis of melanins is generally catalysed by enzymes that are involved in phenol coupling including P450 monooxygenases, ascorbate oxidases, peroxidases, laccases and tyrosinases (Nezbedová et al. 2001). The work presented in this thesis investigated the enzymes involved in melanin biosynthesis in lichens from suborder Peltigerineae. We present a detailed characterization of redox enzymes from Lobaria pulmonaria and Pseudocyphellaria aurata with a special focus on tyrosinases. Furthermore, an evaluation of the response of L. pulmonaria to UV radiation was done. The role of the redox enzymes tyrosinases, laccases and peroxidases in melanin synthesis is elucidated by exposing L. pulmonaria thalli to various combinations of UV and PAR. Laboratory experiments clearly indicated that L. pulmonaria and P. aurata have redox enzymes that are of similar nature to those of their free-living symbionts and other lichens. Tyrosinase from L. pulmonaria oxidized monophenolic compounds and was able to metabolize L-DOPA, thus showing characteristics of a typical fungal tyrosinase. L-DOPA can also be metabolized by peroxidases and laccase suggesting that peroxidases and laccases may participate in melanisation reactions. A field experiment showed that exposing shade-adapted L. pulmonaria to solar radiation in the field induces DOPA melanin synthesis. The synthesis of the brown cortical compounds occurred when lichen thalli were exposed to direct sunlight or placed under a wavelength neutral filter that marginally reduced overall light. Melanin synthesis was accompanied by an increase in laccase activity in lichens that were exposed to natural sunlight. By contrast, no changes in enzyme activity occurred in lichen thalli placed under the wavelength neutral filter. Placing lichen thalli under filters that removed both UV-A and UV-B prevent melanisation and was accompanied by an increase in tyrosinase activity. Filtering out UV-B had no effect on enzyme activity but slight browning occurred. Results showed that laccases could be involved in melanin biosynthesis under some conditions, but provided no evidence for a role of tyrosinases in melanisation. However, it could be that melanisation requires tyrosinase, but rates of melanisation are controlled by the levels of melanin precursors such as tyrosine. No differences in maximal PSII efficiency and chlorophyll contents occurred between the radiation treatments indicating that no deterioration of photosynthetically apparatus occurred and the photobionts remained relatively healthy even when exposed to UV. Taken together these results suggest that further research in needed into determining the levels of melanin precursors, the roles of lichen redox enzymes in melanin synthesis and whether a transcriptional activation of these redox enzymes by UV radiation in lichens exists or not. Furthermore, additional roles for tyrosinase need to be investigated, for example defence against pathogens.
Behavioural responses of rodents to the scent and taste of compounds associated with sugar and protein degradation : implications for the evolution of chemical signals in rodent-pollinated flowers.
(2015) Records, Darryn.; Johnson, Steven Dene.; Nicolson, Susan W.
Volatile compounds in nectar may influence the behavioural responses of animal flower visitors, and thus have fitness consequences for both animal and plant mutualists. Rodents may use certain volatiles associated with sugar fermentation or protein degradation as a cue to locate food. Plants pollinated by rodents may thus emit these volatiles to enhance their attractiveness to rodents. However the presence of certain compounds in nectar may also indicate reward degradation, reducing its attractiveness to potential pollinators. The effects of these compounds on small mammal flower visitors are largely unknown and the consequences of nectar degradation by microorganisms for small mammal flower visitors need investigation. The present study examines the responses of a known rodent pollinator, the Namaqualand rock mouse Micaelamys namaquensis, an occasional floral visitor, the four-striped field mouse Rhabdomys pumilio, and a closely-related congener, the mesic four-striped field mouse Rhabdomys dilectus towards four compounds - ethanol, ethyl acetate, acetic acid and dimethyl disulphide - that are associated with the degradation of sugars and proteins. The study aimed to: (i) Identify if fermentation and protein degradation volatiles act as behavioural cues for small mammals, and have the potential to assist in the finding of food resources; and (ii) to determine the responses of rodents to the taste of volatiles in nectar. In chapter 1, I investigated whether fermentation and protein degradation volatiles elicit a behavioural response in small mammals, using a traditional Y-maze choice apparatus. Rodent species differed in their responses to the four volatiles tested. Ethanol emerged as an attractant for all of the species, with the strongest response seen at the 0.3 % ethanol concentration, while only R. dilectus responded positively to dimethyl disulphide. Acetic acid and ethyl acetate were generally not attractive. In chapter 2, the palatability of fermentation and protein degradation compounds at varying concentrations in experimental nectars was tested. A paired choice test analysis was used to determine if animals altered their choice with the addition of volatiles to sucrose solutions (0.73 and 1.46 M). Rodents showed a dose-dependent response towards the volatile concentrations presented to them. Both M. namaquensis and R. pumilio preferred medium (0.3 %) concentrations of ethanol in high sugar concentration diets, but found this compound distasteful in lower sugar concentration diets. Acetic acid and ethyl acetate were generally not preferred by any species at either sugar concentration. Dimethyl disulphide was preferred by all species when present in greater sugar concentrations (1.46 M) and at low volatile concentrations (0.003 %). These results suggest that rodents do alter their choice of nectar in relation to the tested compounds. I conclude that certain compounds associated with sugar fermentation or protein degradation have the potential to act as behavioural cues in rodent pollination and that further studies to reveal the effect of nectar degradation by microorganisms are needed to help in understanding plant-pollinator interactions.
The effects of petroleum hydrocarbon contamination on selected intertidal macrophytes and meiofauna.
(2015) Naidoo, Krishnaveni.; Naidoo, Gonasageran.; Naidoo, Yougasphree.
The effects of bunker fuel oil on the growth of A. marina, B. gymnorrhiza and R. mucronata were investigated in glasshouse and field experiments. The effects of oil on community structure in micro-organisms were also investigated in microcosm glasshouse experiments. The differences in oil tolerance of the three mangroves were compared in propagule and sediment oiled treatments and growth monitored for 13 months under glasshouse conditions. In propagule oiled treatments, various portions of the propagule were coated with oil. In the sediment oiled treatments, 50ml oil were added to the sediment in each pot. In oiled treatments, plant height, number of leaves and chlorophyll content were significantly reduced in all species compared to the control. In A. marina and R. mucronata, oiling resulted in growth malformations such as abnormal phyllotaxy and deformity of leaves and stems. The effects of oil on root growth were investigated in rhizotrons for 245 and 409 days respectively. In oiled treatments, root growth rate, length and volume were significantly reduced in all species. In A. marina and B. gymnorrhiza oil increased root diameter. In another series of experiments, PAH accumulation in roots and leaves of the three species were determined in one year old seedlings subjected to oiling for 21 days. The concentrations of 15 PAHs in roots and leaves were determined by gas chromatography / mass spectrometry. The highest total concentration of PAHs was accumulated in oiled roots of A. marina (44,045.9μg/kg), followed by B. gymnorrhiza (10,280.4μg/kg) and R. mucronata (6,979.1μg/kg). In oiled treatments, the most common PAHs in roots of all species were fluorene and acenaphthene (two rings), phenanthrene and anthracene (three rings), pyrene and chrysene (four rings) and benzo[a]pyrene (five rings). In the leaves of all species in oiled treatments, the common PAHs were naphthalene and acenapthene (two rings) and phenanthrene (three rings). To test for living and dead root tip cells and to compare the effects of oil on cell ultrastructure in roots and leaves of the three species, one year old seedlings were subjected to a control and sediment oiled treatments for seven days. Control root tips, stained with fluorescein diacetate, exhibited green fluorescence in living cells of the meristematic and conducting tissue in all species. Oiled root tips, stained with propidium iodide, exhibited red fluorescence, indicating cell death or dead cells. Transmission electron micrographs revealed that oil damaged cell ultrastructure in root tips and leaves in all species. Anatomical changes induced by oil included, disorganization of cells in the root cap, epidermis and meristem. Oil also induced loss of cell contents and destruction of organelles in root tissue. Oil damaged chloroplasts and cell organelles in spongy mesophyll and palisade cells of leaves. To compare the effects of oil on the ability of the three species to tolerate salinity, healthy one year old seedlings were subjected to 10% and 50% seawater in control and sediment oiled treatments for 12 months. In the oiled treatments, 200ml oil were added to the soil in each pot. Oil significantly reduced growth in the 50% seawater treatment in all species. Results suggested that oil reduces salt tolerance in the three species. The effects of oil on salt secretion in A. marina were investigated by subjecting one year old seedlings to sediment oiling treatments at 0%, 10% and 50% seawater for three weeks. Sodium accumulated in the leaves of oiled seedlings at 10% and 50% seawater. The effects of oil on salt secretion in A. marina in the light and dark were compared in one year old seedlings subjected to oiling treatments for seven days. Sodium accumulated in the leaves of oiled seedlings in the light and dark within 11 hours. Oil reduced secretion rates of Na⁺, K⁺, Ca²⁺ and Mg²⁺ in all treatments. The effects of oil on species abundance, richness and community structure of soil micro-organisms were determined by subjecting microcosms to oiling treatments with or without fertiliser for four weeks. In the oiled treatments, 15ml oil and 5ml/L fertiliser were added to 200g soil. Fertiliser consisted of 4% N, 2% P and 5% K. Nematodes were extracted after the experimental period and identified to genus or species level. Oil significantly reduced species abundance and richness. Oil also eliminated sensitive species and altered the abundance of dominant species thereby altering the free living nematode community structure. Addition of fertiliser increased richness and dominant species in oiled treatments. The effects of oil coating on leaves and internodes on growth of the three mangroves were investigated in field experiments for 48 weeks. Oiling of the leaves resulted in leaf abscission and decreased leaf production in all species. The effects of sediment oiling (at a dose of 5Lmˉ²) on the three species were also investigated in a field study for 53 weeks. In A. marina, oil caused adventitious roots to develop on the stem, about 10-15 cm above the soil surface after 38 weeks of treatment. In oiled treatments, plant mortality occurred after 53 weeks in all three species. The ability of B. gymnorrhiza and R. mucronata to exclude PAHs from sensitive root tissues probably accounted for the higher oil tolerance than A. marina. The capacity of the species to adapt to residual oil contamination by increasing root diameter (A. marina and B. gymnorrhiza), producing adventitious roots (A. marina), increasing root/shoot ratio (R. mucronata) and abscising oiled leaves (all species) probably contributed to oil tolerance.
The leaf secretory apparatus of Hibiscus surattensis and Hibiscus sabdariffa (Malvaceae) : micromorphology, histo-phytochemistry and ultrastructure.
(2015) Raghu, Kashmira.; Naidoo, Yougasphree.; Nicholas, Ashley.
The research presented here forms the basis of the ethnobotanical and ethnopharmacological evaluation of Hibiscus surattensis, which is a widely distributed vegetable and medicinal shrub used by African and Indian traditional practitioners. Using light microscopy together with advanced electron microscopy techniques, the leaf topography and internal structure was examined. A closely related species, Hibiscus sabdariffa was also investigated for foliar and histophytochemical comparisons. Hibiscus sabdariffa is a widely consumed medicinal species with well-known biological activity and known chemical principles. Both species belong to the section Fucaria within the genus Hibiscus. Analyses of foliar secretory tissues showed that both H. surattensis and H. sabdariffa were characterised by capitate trichomes as well as mucilage-producing ducts and idioblasts. Head cells of capitate trichomes were between 5 and 7, while lignified stalk cells occurred in numbers of 2 or 3. A basal cell supporting the trichome was implanted in the epidermis comparative study between the two species showed some variation of chemical composition in trichome head cells. Polysaccharidic, alkaloids, phenolic and acidic lipid components were identified for both H. surattensis and H. sabdariffa whereas H. sabdariffa showed an intense staining of proteinaceous substances. Densities of capitate trichomes varied from emergent to mature developmental stages, with the highest number occurring on the emergent abaxial surface for both H. surattensis and H. sabdariffa. However, a distinct trend was observed for H. surattensis, where a decrease in capitate trichomes with progressive development was associated with the proliferation of mucilage idioblasts, particularly on the mature adaxial surface. Mucilage producing tissues are assumed to be associated with a protective role against dehydration in a number of plant families. Idioblasts observed in H. surattensis were visible leaf surface structures, embedded in the epidermal tissues. They contained considerable amounts of acidic polysaccharides and acidic lipids, and appeared to be implicated in reducing evaporative water loss in fully expanded leaves. Mucilage ducts were identified in vascular tissue within leaf veins. They occurred parallel with the conducting tissue and comprised of an epithelial layer of cells which seemed to be secreting a mucilage into an extracellular lumen. Crystal idioblasts were also identified in tissues of both H. surattensis and H. sabdariffa. The sequestration of calcium oxalate is assumed to be governed and regulated by specialised mesophyllous idioblasts that in turn appeared to be triggered by excess calcium within the plant body to differentiate into crystal forming cells. The supposed function of calcium oxalate crystals includes mechanical support and herbivory avoidance. TEM revealed the cellular processes which involved crystal idioblast development in H. surattensis. This appeared to demonstrate that degeneration of organelles which are assumed to occur when crystals reach maximal proportions. The non-glandular trichome component of each Hibiscus species was diverse in trichome type and might have accounted for differences in leaf texture and the leaf indumentum. The prickly texture of Hibiscus surattensis was attributed to rigid stellate and falcate trichomes as well as restrorse prickles, whereas the leaves of H. sabdariffa were glabrous with few falcate, bi- and trifurcate trichomes found mainly at the base or along the midvein. Preliminary phytochemical experiments which involved methanolic, chloroform and hexane extracts, yielded favourable results, which showed that leaves of H. surattensis and H. sabdariffa were chemically similar, in this regard, the author recommended further investigation into the phytochemical nature of H. surattensis.

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