Masters Degrees (Biological Sciences)

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    Spatio-temporal patterns of hover fly (Diptera: Syrphidae) diversity across three habitat types in KwaZulu-Natal, South Africa.
    (2022) Mva, Luhlumelo.; Van der Niet, Timotheus.; Midgley, John.; Jordaens, Kurt.
    Hover flies (Diptera: Syrphidae) are poorly studied in southern Africa and as a result, little is known about their abundance throughout the year. Hover fly abundance is generally expected to vary according to vegetation types, due to differences in the type and diversity of floral resources, but this has not yet been investigated in South Africa. This study aims to investigate temporal and spatial patterns of hover flies across three different, but adjacent habitats (forest, grassland, and plantation) in KwaZulu-Natal (South Africa). This was done in two ways, first, by analysing occurrence data from the Global Biodiversity Information Facility’s (GBIF) database across months of the year. Second, by quantifying fly presence and abundance using Malaise traps set up in three different habitats and supplemented with hand-netting data. The Malaise traps were set up from October 2020 - September 2021 and were generally serviced weekly. Hover flies were sorted and identified using available keys at the KwaZulu-Natal Museum. The KZN GBIF database contained 11 tribes that represented three distinct phenological patterns. The Karkloof GBIF dataset contained seven tribes that represented two distinct phenological patterns. The field collection dataset contained eight tribes that represented two distinct monthly patterns, although these slightly differed from the patterns found in the GBIF data. Most records were from January, September and December whereas few records were from May, June and July. Minimum-minimum, average-minimum, average maximum and average temperature showed a positive correlation with hover fly abundance. In terms of spatial patterns, grassland had the highest number of adult hover fly individuals (154), followed by forest (106) and plantation (20). There was an overall effect of habitat type on both the mean number of species and the number of individuals per trap per week. In addition, Shannon’s diversity showed variation among the three habitat types whereas Simpsons Evenness index showed no variation. The results show that hover fly populations remain active throughout the year, but with distinct fluctuations in their abundance. They also show that heterogeneous indigenous habitats such as grassland and forest represent high abundance, diversity, evenness and richness of hover flies and this may be due to the diversity in available microhabitats, compared to the plantation, which is a modified habitat that lacks microhabitat diversity. Nonetheless, further studies should be undertaken in different regions of South Africa to better understand the monthly patterns as well as the effect of habitat type on hover fly diversity.
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    The importance of sunbirds as pollinators of the southern African plants Alberta magna and Streptocarpus dunnii.
    (2023) Jansen, Cally Julia.; Johnson, Steven Dean.; Cozien, Ruth Jenny.
    Bird pollination is important for plant reproduction and is found in about 65 flowering plant families. I studied putative bird pollination systems of two southern African plant species, Alberta magna (Rubiaceae), which grows above cliffs and Streptocarpus dunnii (Gesneriaceae), which occurs among boulders in open grassland. A special emphasis of this study was to test the utility of close-focusing motion-activated camera traps for documenting bird pollination of plant species that are rare and occur in habitats that present challenges for conventional bird pollination studies. The aims of this project were to 1) quantify floral traits for comparison with other bird-pollinated species; 2) establish the breeding systems, including reliance on pollinators for reproduction; 3) identify floral visitors and measure their contribution to reproductive success; and 4) determine if seed production is pollen limited. Floral traits of S. dunnii, including large volumes of dilute nectar and long tube lengths, also aligned with an ornithophilous pollination syndrome. In naturally-pollinated plants, overall fecundity was high, with 80% of flowers developing fruits with large numbers of seeds. Malachite sunbirds (Nectarinia famosa) were the only legitimate visitor to flowers of S. dunnii. Greater double-collared sunbirds (Cinnyris afer), which have shorter bills than malachite sunbirds, occasionally rob flowers of nectar. Streptocarpus dunnii was found to be fully self-compatible but does not self-autonomously. Selective exclusion experiments showed that the species is almost entirely reliant on birds for seed production. The species did not experience pollen limitation, indicating that sunbirds are effective pollinators. Measured floral traits of A. magna, including large volumes of dilute nectar and long tube lengths, were consistent with an ornithophilous pollination syndrome. Motion triggered cameras and observations showed that southern double-collared sunbirds (Cinnyris chalybeus) and olive sunbirds (Cyanomitra olivacea) were the most common visitors to A. magna, while amethyst sunbirds (Chalcomitra amethystina) were occasional visitors. Selective exclusion experiments showed that bird visitors are important for reproductive success which was found not to be pollen-limited. The low levels of fecundity in A. magna indicate that there may be other potential factors that limit seed production in this species and the degree of self-compatibility in this species was not firmly established. I conclude that both species are specialized for pollination by sunbirds and that close-focusing camera traps are effective for documenting bird pollination systems of rare plant species which occur in habitats that are challenging for conventional human observations.
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    Assemblages of surface-active arthropods in pristine and disturbed savannah.
    (2022) Mavasa, Risuna Wain.; Tsvuura, Zivanai.; Yekwayo, Inam.; Mwabvu, Tarombera.
    Savannahs are structurally complex ecosystems rich in biodiversity, and the conversion of savannahs into human-modified landscapes poses a significant risk to organisms with limited dispersal abilities. Surface-active arthropods have limited dispersal abilities and are sensitive to changes in their environment; as a result, they are restricted to specific habitats with the resources that they need. Surface-active arthropods are essential in ecosystem processes, such as playing a direct or indirect role in water infiltration into the soil and its storage, decomposition of plant and animal matter, nutrient cycling, humification and biological pest control. Many landscapes of natural vegetation in South Africa have been transformed into croplands, residential and commercial areas. In Mpumalanga province, croplands (including fruit plantations) dominate the landscape where savannah vegetation occurred previously. This study investigated the influences of disturbance (transformed savannah vs pristine savannah) and season on the diversity of four groups of surface-active arthropods (ants, beetles, millipedes and spiders) in the savannah biome in Mpumalanga province. The objectives of the study were to determine 1) whether flightless arthropod assemblages differ between disturbed and pristine savannah, and 2) whether functional guilds of flightless arthropod assemblages differ between summer and winter. Field sampling using pitfall traps was done to collect the surface-active arthropods in pristine and disturbed savannah during summer and winter. All samples were sorted into morphospecies and identified into family or genus where possible and then assigned into functional guilds. Functional guilds were allocated based on the feeding habits of the surface-active arthropods, resulting in three distinct functional guilds (detritivores, herbivores and carnivores) being identified. Surface-active arthropods with diverse feeding habits were lumped to form a fourth functional guild, the “diverse functional guild”. I found significant differences in the species composition of ants, beetles and spiders between disturbed and pristine savannahs. The species richness and abundance of ants was significantly greater in the disturbed than pristine savannah, while beetles and spiders had similar species richness and abundance between the two habitat types. The number of unique species of the surface-active arthropods in the disturbed savannah were two times more than those in the pristine savannah. The species richness and composition of all functional guilds of the surface-active arthropods differed between summer and winter, with greater species richness in summer than in winter. Herbivorous and carnivorous arthropods had significantly higher abundance in summer than in winter, while the abundance of detritivores and the “diverse functional guild” was similar between the two seasons. This study shows that different vegetation types support different compositions of surface-active arthropods and that there may be a positive relationship between vegetation structure and the assemblages of surface-active arthropods in the savannah. Furthermore, the study highlights a potentially positive relationship between the assemblages of herbivorous and carnivorous arthropods. I recommend that it may be essential to consider both disturbed and pristine habitat types in the conservation of surface-active arthropods, especially in South Africa where most of the land is outside of formal conservation areas. This assertion is supported by the greater number of unique species of surface-active arthropods found in the disturbed than in the pristine savannah. My study provides relevant baseline information because little is known about the assemblages of surface-active arthropods in savannah landscapes dominated by croplands. Furthermore, my study gives some insight on how the assemblages of surface-active arthropods may respond to disturbance in the savannah.
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    Effects of high light intensity and desiccation stress on moss species.
    (2021) Mbatha, Sikhethile.; Beckett, Richard Peter.
    Bryophytes are desiccant tolerant non-vascular plants, capable of growing and surviving in extreme conditions. They are divided into three groups: liverworts, hornworts, and mosses. Most mosses grow in shady and moist environments, although some formpart of arid soil crusts where they protect soil from erosion. The moss flora of the Afromontane vegetation around Pietermaritzburg is dominated by two acrocarpous mosses, Atrichum androgynum, and Dicranella subsubulata. A. androgynum tends to grow in wetter, more shaded habitats and is a rather delicate species, while D. subsubulata grow in open, drier habitats and is more robust. Rarely, the species grow together, for example at the transition of a shaded indigenous woodland to plantations. We hypothesized that the more robust species has higher stress tolerance and has largely constitutive stress tolerance mechanisms. By contrast, we hypothesized that the more delicate species is less tolerant and may have inducible tolerance mechanisms. In the present study, desiccation tolerance and tolerance to high light stress were investigated in A. androgynum and D. subsubulata. Results confirmed that D. subsubulata was more tolerant of high light stress than A. androgynum. Exposure to moderate light intensities did not increase tolerance to subsequent high light stress in either species. Similarly, D. subsubulata was more desiccation tolerant than A. androgynum. Not consistent with our original hypothesis, mild desiccation, and treatment ABA-induced tolerance to desiccation in both species. Furthermore, detailed studies of the antioxidant enzyme peroxidase showed that enzyme activity was induced during slow drying in both D. subsubulata than A. androgynum. It appears that inducible tolerance mechanisms are present in both species. The work presented here represents a contribution to the autecology of two important mosses in the KwaZulu-Natal midlands Afromontane vegetation.
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    The curious case of the Satyrium neglectum complex: a taxonomic, ecological, and phylogenetic study.
    (2021) Rule, Matthew James.; Van der Niet, Timotheus.; Johnson, Steven Dene.
    Classical herbarium taxonomy uses a combination of morphological features from pressed specimens and distribution information as data in species delimitation. Whilst morphology is critical to species delimitation, additional information on phylogenetic relationships and fieldbased studies of ecology can inform taxonomic decisions. Satyrium neglectum Schltr. is a highly variable taxon currently comprising two subspecies, ssp. neglectum (Schltr.) A.V. Hall and ssp. woodii (Schltr.) A.V. Hall both of which are widely distributed in subtropical southern and eastern Africa. Variation in morphology and differences in apparent pollination systems, both between the subspecies, and within ssp. neglectum suggests the presence of two taxa within the current concept of the latter. This information, as well the exclusive reliance on herbariumbased taxonomy in the most recent species revision, formed the background to this re-evaluation of the systematics of the complex. A combination of detailed morphometrics and molecular phylogenetics demonstrated that the current concept of S. neglectum ssp. neglectum is inaccurate. A principal components analysis using traits measured from herbarium records from across the range, and ethanol-preserved flowers from South Africa, confirmed the presence of two morphologically distinct clusters within ssp. neglectum. Representatives from these two forms are separated along an elevation gradient. Bayesian inference using a combination of plastid and nuclear DNA sequence data revealed incongruent topologies. The nuclear topology is not consistent with current taxonomic boundaries, and revealed a separation between the high-elevation form and the low-elevation form of S. neglectum ssp. neglectum as well as ssp. woodii. The plastid data set also confirmed the separation of the high-altitude form of ssp. neglectum, but otherwise the topology reflects geography, as southern African accessions are separate from eastern African accessions. Floral scent was investigated in combination with published and novel pollinator observations in the Satyrium neglectum species complex. Floral scent composition and emission rates are mostly representative of associated pollination syndromes in previously published pollination work, with the exception of the high-elevation form. Published observations of ssp. woodii indicate that the species is pollinated by amethyst and the greater-double-coloured sunbirds, and the scent composition and emission rate is largely reflective of this. In addition, published observations showed a system of butterfly pollination in the low-elevation form of ssp. neglectum and long-proboscid fly pollination in the high-elevation form. However, new night time observations revealed additional pollination by settling moths in the high elevation form. These observations make sense in the context of the scent profile, which is dominated by compounds typically associated with moth pollination, such as phenylethyl alcohol and eugenol. In addition, two settling moths showed electroantennographic responses to the two dominant compounds in the scent bouquet of the high-elevation form. The compounds that dominate the scent of the butterfly-pollinated low elevation form are hexan-1-ol, octan-1-ol and benzyl alcohol, while previously published work on ssp. woodii showed that it is virtually unscented, and has emission rates that are markedly lower than other taxa pollinated by insects. Based on the morphological, phylogenetic and scent chemistry analyses, a case was made for the high-altitude form to be considered a separate taxonomic entity. This is formally described under the new name Satyrium basutorum. The analyses suggest that ssp. neglectum and ssp. woodii should be retained as subspecies, but a further investigation is required to identify the status of the eastern African synonyms Satyrium sceptrum and S. neglectum var. brevicalcar.
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    Varying soil nutrition in grassland and savanna ecosystems affect plant-microbe symbiosis, nitrogen nutrition and growth of Pisum sativum L.
    (2019) Zungu, Ntuthuko Sifiso.; Magadlela, Anathi.; Khoza, Thandeka Ntokozo.; Mathabe, Patricia Mmatshetlha Kgomotso.
    Abstract available in pdf.
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    Foliar secretory cavities of Vepris lanceolata (Lam.) G. Don (Rutaceae): micromorphology and chemical composition of the secretion.
    (2017) Nxumalo, Nozipho Ntombikayise.; Naidoo, Yougasphree.; Naidoo, Gonasageran.
    Secretory structures such as ducts, trichomes and cavities consist of cells that are primary sites synthesizing essential oils and other phytochemical compounds with medicinal properties. Little is known about the micromorphology of secretory structures and the composition of the chemical constituents. There was no information documented on the micromorphology of secretory structures of Vepris lanceolata (Lam.) G. Don (Rutaceae family). The aim of this research was to investigate the micromorphological characteristics of foliar secretory cavities, the chemical composition of the secretion, and the antibacterial activity of leaf extracts of Vepris lanceolata. Scanning electron microscope (SEM) images by chemical fixation and freeze drying revealed no external secretory structures on the surfaces of leaves. SEM images by freeze-fracture showed secretory cavities present in the leaf blade. The cavities were embedded amongst palisade and spongy parenchyma cells, next to the vascular bundle. Cavities were made up of the lumen surrounded by varying layers of epithelial cells, depending on the secretory phase of the cavity. Semi-thin and ultra-thin sections showed that foliar cavities develop schizo-lysigenously, i.e. cavities develop by both separation and degradation of epithelial cells. Transmission electron microscope (TEM) sections showed that during the secretory stage, secretory cells contained oil droplets, vacuoles and vesicles indicating active secretion. Histochemical assays of fresh leaves showed the localization of phytochemical compounds. Cavities turned orange red when stained with Sudan III indicating the presence of lipids and pink with NADI reagent to show essential oils. Cavities also stained positive for polysaccharides, sugars, phenolic compounds, proteins and alkaloids. Phytochemical screening showed the presence of alkaloids, glycosides, carbohydrates, proteins, tannins, phenolic compounds, flavonoids, fixed oils and fats. Preliminary thin layer chromatography (TLC) showed separation of bands indicating groups of active compounds in leaf extracts. Crude (ethanolic and methanolic) and water extracts of leaves showed antibacterial activity against gram positive bacteria Staphylococcus aureus (ATCC 25923) and methicillin-resistant Staphylococcus aureus (ATCC BAA-1683); and five strains of gram-negative bacteria: Escherichia coli (ATCC 25922), Escherichia coli (carbapenem-resistant) (ATCC BAA 2340), Klebsiella pneumonia (ATCC 314588), Pseudomonas aeruginosa (ATCC 27853), as well as Salmonella typhimurium (ATCC14026) according to the disc diffusion method. Leaf extracts have tannins, alkaloids, flavonoids, essential oil and flavonoids responsible for the antimicrobial activity of the plant.
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    Histo-phytochemical evaluation and characterisation of the foliar structures of Tagetes minuta L. (Asteraceae).
    (2018) Rikisahedew, Jesamine Jöneva.; Dewir, Yaser Hassan.; Naidoo, Yougasphree.
    Plants have been used as ethnomedicine for millennia. In recent years, there has been an upward surge of interest in the use of plants as medicine due to the interest in drugs with fewer side effects as well as the fight against antibiotic resistance. This study is based on Tagetes minuta, an aromatic essential herb that is cultivated for its high percentage essential oils which have been used in the treatment of various ailments. In addition, T. minuta contains a myriad of secondary metabolites that serve in numerous industrial and clinical applications. The aim of this study was to characterise the foliar structures responsible for the production, storage, and exudation of these useful compounds, as well as to examine the chemical constituents of the crude organic solvents derived from the leaves of T. minuta. The potential for green synthesis of silver nanoparticles from the crude methanolic extract and its potential as an antibacterial was also determined. Stereomicroscopy and scanning electron microscopy revealed the presence of uniseriate non-glandular trichomes on the foliar surfaces, as well as large pellucid secretory cavities. Histochemical analyses on the non-glandular trichomes showed that they are capable of storing various bioactive compounds, which is a novel discovery for this species. The development of the subdermal secretory cavities show that the cells undergo autolysis in order to form a schizolysigenous cavity in mature leaves, which was revealed using light microscopy. The ultrastructure of the secretory epithelium within the secretory cavity was analysed using transmission electron microscopy, which displayed the changes of the plastids to contain lipid molecules as well as an increase in vesicles indicating the presence of essential oils. Phytochemical analysis on the crude organic solvents derived from the leaves of T. minuta revealed the presence of alkaloids, sterols, saponins, terpenoids, phenols, and lipids. Gas-chromatography mass-spectrometry was carried out to reveal that the constituents with the highest percentage were 9-octadecen-1-ol (4.51 %), β-sitosterol (6.07 %), olean-12-en-3-one (7.47 %), and 3-methyl-1-butanol (14.77 %), all of which cause bacterial growth inhibition, as well as showing acaricidal activity, and anticancer properties in studies focussed on clinical applications. Silver nanoparticles were successfully synthesised from the methanolic leaf extract, which was confirmed using UV-visible spectroscopy and energy dispersive x-ray analysis. UV–visible spectrum of synthesised silver nanoparticles showed maximum peak at 442 nm, and transmission electron microscopy revealed the silver nanoparticles to be spherical in shape, ranging from 10 to 50 nm in diameter. Preliminary antimicrobial activity was determined using the agar well diffusion method, which showed growth inhibition against E. coli, S. aureus, methicillin-resistant S. aureus, B. subtilis and P. aeruginosa. This study has shown that T. minuta contains numerous bioactive compounds that have pharmacological and medicinal uses, as well as characterising the non-glandular trichomes present on the adaxial and abaxial leaves for the first time. The synthesis of silver nanoparticles from the methanolic extract of T. minuta in this study is novel, and shows promise for cheaper, more effective, and less risky nanotechnological applications.
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    Taxonomic revision of the Memecylon natalense species-complex in southern Africa.
    (2018) Mona, Imercia Gracious.; Ramdhani, Syd.
    iv ABSTRACT Two Memecylon species (M. natalense and M. bachmannii) were previously recognized in southern Africa, and three species of M. section Buxifolia as occurring in Mozambique (M. natalense, M. torrei and M. insulare). However, a recent nrDNA phylogeny revealed that M. natalense as previously circumscribed is not a monophyletic group and includes some geographically outlying populations warranting recognition as distinct evolutionary lineages. In this dissertation, I present the results of a comprehensive morphological study and revision of the ‘M. natalense species-complex’ including the following taxa: M. natalense sensu stricto (endemic to South Africa in KwaZulu-Natal, Eastern Cape, Mpumalanga and Limpopo), M. incisilobum (southern Mozambique), M. nubigenum (southern Malawi), M. rovumense (southeastern Tanzania), M. aenigmaticum (northern Mozambique), M. sp. nov. 1 (northeastern Kwazulu-Natal, South Africa), M. sp. nov. 2 (Eastern Cape province, South Africa), and M. sp. nov. 3 (Limpopo province, South Africa). Although M. bachmannii (Eastern Cape and KwaZulu-Natal) is morphologically distinct, it was also included in the present study because of recent nrDNA work indicating a close relationship with M. natalense. Species boundaries were investigated using phenetic and cladistic methods based on morphology. Phenetic analysis using the Unweighted Pair Group Method with Arithmetic Mean (UPGMA) revealed that M. bachmannii clustered independently, M. sp. nov. 1 clustered close to M. incisilobum, M. aenigmaticum clustered close to M. rovumense, and M. nubigenum clustered close to a subcluster consisting of M. natalense, M. sp. nov. 2 and M. sp. nov. 3. The morphological cladistic results were congruent with a previous molecular analysis in that M. sect. Buxifolia was monophyletic. Memecylon rovumense + M. aenigmaticum and M. incisilobum + M. sp. nov. 1 appeared in separate clades. Memecylon bachmannii and M. nubigenum were each monophyletic, while M. natalense, M. sp. nov. 2 and M. sp. nov. 3 appeared in an unresolved polytomy. Overall, the cladistic results were in agreement with the UPGMA analysis but were less resolved. Vegetative and fruit morphology have proven to be of great diagnostic value in delimiting the members of the M. natalense species-complex. Four new species have been named and described, while a further three new species are proposed. Dichotomous keys are provided, and the conservation status of each species is assessed according to the criteria of the IUCN. A lectotype should be designated for M. natalense. With these additions, the flora of Mozambique now has seven species of Memecylon (including six species in M. sect. Buxifolia). In South Africa, the number of recognized Memecylon species has increased from two to five.
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    DNA barcoding of KwaZulu-Natal afromontane forest parmelia (parmeliaceae) species: a molecular approach to accurate specimen identification and sensitivity to climate change.
    (2018) Ndhlovu, Nqobile Truelove.; Willows-Munro, Sandi.; Beckett, Richard Peter.
    Accurate species identification is challenging, especially in groups with subtle taxonomically diagnostic characters such as lichens. Molecular-based techniques have shown to be a valuable tool for accurate specimen identification in fungi, in particular the use of DNA barcoding has become popular. Specifically, the internal transcriber spacer (ITS) region has been shown to successfully discriminate a broad range of fungal species. In this study, the utility of the ITS DNA barcode for use as a species diagnostic tool in the cosmopolitan lichen-forming fungus, Parmelia (Parmeliaceae) was investigated. Sixty-eight ITS sequences were generated from specimens collected from five sites around the province of KwaZulu-Natal and analysed. Phylogenetic analysis indicated that unlike European Parmelia species that form strong monophyletic clades, what appeared to be morphologically very similar Parmelia species in KwaZulu-Natal are paraphyletic or polyphyletic. No barcode gap was detected between the intra and interspecific distances. This suggests that the taxonomy of Parmelia lichens in South Africa needs to be thoroughly revised. The molecular data presented in this study provides evidence of previously hidden species-level diversity in Parmelia and as such contributes to the knowledge and understanding of the biodiversity of lichenized fungi in South Africa. The thermotolerance of Parmelia collected from different sites along an altitudinal gradient around Kwa-Zulu Natal was invesigated Chlorophyll fluorescence was used to assess the performance of lichen photobionts following stress, while ion leakage that of the mycobiont. For heat tolerance, results suggested that tolerance was correlated with the climatic conditions in which the lichens grow. Material from the coastal site of Hawaan were more heat tolerant than that from the three Midlands sites. Counter to our expectations, the coastal collections were more cold tolerant than those from the other sites. However, the genus clearly contains genetic variation with respect to stress tolerance, suggesting that it may have the potential to adapt to climate change.
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    Chemokines and haematological profile in Sprague-Dawley rats infected with trichinella Zimbabwensis and Plasmodium berghei ANKA.
    (2017) Mdleleni, Yanga.; Mukaratirwa, Samson.; Murambiwa, Pretty.
    Trichinellosis is an important re-emerging parasitic zoonosis caused by nematode species of the genus Trichinella and Plasmodium falciparum malaria is among the leading causes of mortality and morbidity in sub-Saharan Africa (SSA). Co-infection of the two diseases in rural communities is likely to be a common phenomenon although no reports have been made up to date. There is paucity of information on the consequence of co-infection regarding the clinical outcome of these diseases, especially malaria. Helminths, such as tissue-dwelling Trichinella spp larvae induce Th2 immune responses, while malaria induce Th1 immune responses as a survival strategy. On the other hand, chemokines are chemotactic cytokines produced by the host macrophages in order to elicit a protective immune response. Immuno-pathogenesis during co-infection remain obscure. It is against this background that this study aimed to determine host chemokine and haematological responses in male Sprague-Dawley rats during co-infection with Trichinella zimbabwensis and Plasmodium berghei. A 42 day follow up study was carried out, where 168 male Sprague-Dawley rats (90-150g) were randomly divided into four separate groups, control (n=42); malaria infected (n=42); Trichinella infected (n=42) and co-infected group (n=42). On day 0, male Sprague-Dawley rats were orally infected with 3 T. zimbabwensis muscle larvae per gram body weight. Followed at day 28 post-Trichinella infection with malaria induction using 105 P. berghei parasitized RBCs for the mono-infected group. While 42 male Sprague-Dawley rats were co-infected with T. zimbabwensis and P. berghei on day 0 and P. berghei infection occurred on day 28pi. Experimental animals were sacrificed on day 0,7,14,21,28,35 and 42 pi. Where whole blood and sera were collected. Plasmodium berghei percentage parastaemia, T. zimbabwensis adult worm count and muscle larvae load, haematological parameters and serum levels of IP-10, RANTES and EOTAXIN were determined. Co-infection with P. berghei and T. zimbabwensis showed elevated P. berghei percentage parastaemia, as well as upregulation of RANTES and EOTAXIN as a Th2 immune response, while IP-10 was downregulated as an effective immune response to parasitic infections. Mono-infection with P. berghei caused an upregulation of IP-10 as a Th1 immune response.
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    Screening, selection and clonal propagation of Amaranthus dubius genotypes with different calcium and iron content.
    (2018) Dladla, Phindile.; Shaik, Shakira.; Watt, Maria Paula Mousaco Deoliveira.
    Decontamination of Amaranthus dubius field-derived nodal explants was achieved in a 10 min soak with 1% (v/v) NaOCl and 2 drops of Tween 20® followed by three rinses in sterile distilled water, immersion in an antibiotic solution (¼ strength Murashige and Skoog basal salt medium, 50 μg l-1 rifampicin, 100 μg l-1 streptomycin/penicillin), at 1500 rpm for 5 h. Of the tested plant growth regulators, MS media supplemented with 2 mg l-1 benzylaminopurine (BAP) + 0.5 mg l-1 indole-3-acetic acid (IAA), and 0.1 mg l-1 IAA, respectively, gave the best in vitro responses of 4 shoots/nodal explant and 100% rooting. Plantlets were acclimatised over 21 days (d) in soil (S) and 1soil:1vermiculite (v/v) (1S:1V) substrates; a significant increase in the number of leaves occurred up to 21 d (6.8 to 16.2 in S and 6.3 to 13.1 in (v/v) 1S:1V). At 21 d in (v/v) 1S:1V, there were more leaves than in S, in contrast longer plant height and root length were observed in S than in (v/v) 1S:1V. The post-acclimatisation yield was 2 plants/nodal explant. The variation in calcium (Ca) and iron (Fe) content within a population of greenhouse-germinated A. dubius seedlings was then evaluated, and specific genotypes were selected to investigate the effects of micropropagation, acclimatisation in S and (v/v) 1S:1V and physiological age (time) on their growth and Ca and Fe accumulation. After 60 d, using inductively coupled plasma-optical emission spectrometry, the content of leaf Ca ranged from 246.3 to 765.3 mg 100 g-1 dry mass (DM) and the Fe from 5.3 to 26.7 mg 100 g-1 DM. Based on the significant differences of these levels amongst the parent genotypes seven were selected and were ‘ranked’ as G47 > G45 > G11 > G41 = G8 > G39 > G15 for Ca and as G47 = G45 > G39 = G41 > G8 > G15 > G11 for Fe. Nodal explants of the selected parent genotypes were subjected to the established micropropagation protocol (using S and (v/v) 1S:1V during acclimatisation). The post– acclimatisation yield was 2 to 4 plants/nodal explant. Over the 21 d of acclimatisation in the two substrates, there were clear genotypic effects on all the tested growth parameters in S. There were significant increases in S–grown plants in the number of leaves of G39 (7.0 to 10.6) and G47 (7.0 to 13.3), the plant height of G11 (5.6 to 12.3 cm) and the root lengths of G8 (6.6 to 17.3 cm) and G41 (10.0 to 16.6 cm). When grown in (v/v) 1S:1V, the plant height significantly increased from d 0 to 21 for G8 (7.0 to 12.3 cm) and G47 (6.3 to 10.6 cm). With regards to the effect of substrate, only the clones of G8 preferred nutrient-poor soil to produce more leaves (8) than when grown in S at d 21 (5). After transferring the clones to the greenhouse for 90 d, no significant differences in the root:shoot dry masses amongst the clones were observed on each substrate, and the substrate had no effect on the root:shoot dry mass for each genotype. After acclimatisation and transfer of the clones into the greenhouse it was observed that both physiological age (time, i.e. 15, 30, 60, 80 and 90 d) and substrate influenced their accumulation of Ca and Fe. Over time, in both substrates, the Ca content increased while Fe content decreased. Significant interactions were found between the genotype and substrate for both Ca and Fe, and between physiological age (time) and genotype for Fe only. In S, the clones of all the parent genotypes matched the Ca content of their parents at 15 d while for Fe that of five of the seven selected genotype clones were similar in Fe content to their parents at 60 d. Clones of four of the seven selected parent genotypes accumulated higher Ca and Fe levels when grown in (v/v) 1S:1V than in S at certain time intervals. In S, the Ca ‘rankings’ of all the clones did not match their respective parent genotypes at any time interval while in the case of Fe, the clones of two genotypes in S (G47 and G11) and one genotype in (v/v) 1S:1V (G47) matched their respective parents between 60 to 90 d of growth in the greenhouse. In conclusion, nodal explants of the selected A. dubius genotypes with varying Ca and Fe contents, were clonally propagated in vitro using BAP and IAA and the yield after acclimatisation was 2 to 4 plants/nodal explant. The physiological age (time) and substrate affected the number of leaves of the cloned genotypes, whilst in the case of Ca and Fe, these levels were influenced by micropropagation, physiological age and substrate type. Phenotypic plasticity can be further evaluated by exposing the clones of the selected genotypes to varying water, salinity and heat stresses. Additionally, investigations to understand the clones’ ability to accumulate Ca and Fe would be valuable, in this regard, quantifying inhibitory factors and exploring the effects of substrate properties such as pH and porosity are suggested.
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    Molecular identification of hookworm isolates from stray dogs, humans and selected wildlife from KwaZulu-Natal and Mpumalanga provinces of South Africa.
    (2019) Ngcamphalala, Philile Ignecious.; Mukaratirwa, Samson.; Lamb, Jennifer Margaret.
    Hookworms are nematodes that cause infections to the host via skin penetration of the third stage larvae and they are widely distributed in the tropical and subtropical regions. They parasitize a wide range of host species including humans and companion animals such as dogs and cats. An estimated 740 million people are infected worldwide, with sub-Saharan Africa having the highest documented prevalence. With the exception of the study conducted by Lamb et al. (2012), Ancylostoma species identification in Southern Africa was solely based on egg morphology and morphological characteristics of adult worms. This study therefore aimed at using molecular techniques to identify the hookworm species and their prevalence in stray dogs, school-going children from KwaZulu-Natal province, and selected wild canids and felids from Mpumalanga province of South Africa. A total of 356 faecal samples were collected and screened for the presence of hookworm eggs using coproscopy and coproculture, which yielded prevalence of 23.04% and 21.67%, respectively. Larvae derived from coproculture of a total of 55 samples were subjected to molecular analysis. DNA was isolated and subjected to PCR amplification, PCR-RFLP and sequencing of the nuclear ribosomal internal transcribed spacer (ITS1) and 5.8S rRNA region. PCR-RFLP showed an overall prevalence of 72.7% (40/55) for A. caninum, 12.7% (7/55) for a mixture of A. caninum, A. ceylanicum, and A. braziliense, 7.27% (4/55) for A. caninum and A. ceylanicum mixed infection, and 7. 27% (4/55) for A. caninum and unidentifiable species. These results are consistent with other studies which show that A. caninum is the most dominant hookworm species worldwide even though A. braziliense is regarded as a more important zoonotic species. Phylogenetic analyses of alignments based on the DNA sequences were also used to identify isolates which were sequenced. However, results of phylogenetic analysis were not consistent with results from PCR-RFLP analysis as none of the sequences matched with A. ceylanicum. Sequencing also showed a 0.68% prevalence for both A. caninum and A. braziliense (mixed infections) in dogs. Of great importance was the revelation that A. caninum can now cause a patent infection in South African (Ingwavuma area) school-aged children as all human isolates matched with A. caninum with a prevalence of 6%. For wildlife, a prevalence of 10% was recorded. However, due to the small sample size, these results cannot be regarded as a true reflection of the prevalence in wildlife. Thus there is a need for future studies that will increase sample sizes, broaden ranges as well as look into finding better detection methods, primers and/or restriction enzymes that are specific to hookworm species isolated in South Africa.
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    An investigation of the prerequisite conditions which enable invasion of moist grasslands by Dalbergia obovata.
    (2019) Stewart, Terry.; Scogings, Peter Frank.; Baijnath, Himansu.
    Encroachment by Dalbergia obovata into moist coastal grasslands within eThekwini Municipality was identified as a potential threat to the conservation of this threatened habitat type. An investigation was undertaken to confirmation whether or not D. obovata was in fact encroaching into the city's grasslands and to identify the probable underlying drivers and local factors that contributed to encroachment. The investigation comprised four areas of focus. The first was an assessment of the management practices and records of D. obovata encroachment extracted from the available reserve management records of seventeen nature reserves and conservation areas. The summarised data from the records was then processed using a Chi-square test and Principal Components Analysis. The second phase focussed on the application of field surveys to record the distribution patterns of D. obovata within four selected management areas. The results were then captured in a GIS map against which the results of the Chi-square and PCA tests were compared. In phase three a simple theoretical ballistics model was developed from experimental data to predict the anomochoric distribution of D. obovata diaspores from the forest into adjacent grasslands and the critical positioning of parent plants in the host trees in relation to the edge of the canopy. The final focal area involved the mapping of individual plants within stands of D. obovata and comparison against sixteen years of historical orthophoto records in order to determine the historic patterns and rates at which D. obovata became established and propagated within the grasslands. During this part of the investigation the vegetative propagation of D. obovata by clonal propagation from lateral branches and roots was identified and which provided an explanation of how D. obovata was able to form large dense stands with no evidence of canopy contact mortality. The final conclusions were that D. obovata parent plants had to be positioned on the edge of the forest canopy for diaspores to successfully escape the canopy, that successful establishment of D. obovata in grasslands from any particular parent plant was infrequent and that D. obovata did not become established in the presence of large mixed feeder herbivores. However once established in the grasslands, the ability to spread via clonal propagation enabled D. obovata to both outcompete grass species for available resources and be resilient to conventional bush encroachment management practices which relied on controlled burning programs.
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    Small mammal responses to Scarp Forest Restoration in the Maputoland-Pondoland-Albany Hotspot, South Africa.
    (2017) Lazarus, Angelique Tiara.; Schoeman, Marthinus Cornelius.; Vosloo, Dalene.;
    Restoration ecology is a relatively new field. Although a range of attributes have been used to assess restoration success, they have not been standardised across studies. Recently, three main ecological attributes have been identified as key measures to standardise the assessment of restoration success: species diversity, vegetation structure and ecological processes. However few studies have combined more than two of these ecological attributes when assessing restoration success. The aim of this study was to apply these three ecological attributes to determine whether Scarp Forest restoration has been successful from the perspective of small mammals at the Buffelsdraai Landfill Site, Durban, South Africa. I assessed the response of small mammals to Scarp Forest restoration at 2, 4 and 6 year post-restoration periods. I surveyed small mammals every three months for one year in three restoration treatments (2010, 2012 and 2014 restored), as well as in surrounding sugarcane and riverine forest sites. At each site I measured the vegetation structure and small mammal diversity. Additionally, I conducted stable isotope analysis on vegetation and invertebrate samples to compile a baseline database of potential prey, and compared these data with the stable isotope composition of hair and tissue samples collected from rodents and shrews to analyse the trophic structure of the small mammal assemblages. In support for the prediction that vegetation structure should increase in complexity at restored sites, tree species richness, density and height were higher at the 2010 restored than more recently restored sites; and grass height and percentage cover were highest at 2012 restored sites. Except, forb and grass species richness were higher at newly restored sites. Second, rodent abundance was higher at the 2010 restored sites than the 2012 and 2014 restored sites and sugarcane sites. However, shrew abundance and species richness were not significantly different among the study sites. Third, carbon and nitrogen isotopic composition of rodent hairs suggest that these species utilised resources associated with the 2010 restored sites rather than those associated with recently restored sites, sugarcane sites and forests. Further, the stable isotope ratios of carbon and nitrogen in Mastomys natalensis’ tissues showed that these rodents predominantly utilised resources associated with the 2010 restored sites irrespective of the tissue that was analysed. Conversely, carbon and nitrogen isotopic composition of shrew hairs suggest that these species foraged at the sites where they were captured. Taken together, my results suggest that at Buffelsdraai, the restoration efforts have ensured progressive succession in the scarp forest after 10 years, at least from the perspective of most small mammals.
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    Secretory structures of croton gratissimus Burch. Var. gratissimus (Euphorbiaceae): Micromorphology and Histophytochemistry.
    (2018) Naidoo, Danesha.; Naidoo, Yougasphree.; Naidoo, Gonasageran.
    Croton gratissimus Burch. variety (var.) gratissimus (Euphorbiaceae) has a widespread distribution in tropical Africa and is frequently used in African traditional medicine to treat various ailments. In South Africa, dried leaves of C. gratissimus are smoked to treat influenza, colds and fever. Due to its extensive use in traditional medicine, research on the phytochemical composition of C. gratissimus has been documented. According to literature, these phytochemicals are possibly secreted or accumulated in secretory structures. However, little or no research is available on the structures involved in the production and/or accumulation of phytochemicals in C. gratissimus. Therefore, this study aimed to describe the micromorphology of trichomes and laticifers from the leaves and stems of C. gratissimus as well as to identify the possible site of synthesis of phytochemicals. Furthermore, the chemical composition and antibacterial properties of phytochemicals in the leaves and stems were also determined. In addition, the antibacterial activity of biosynthesised silver nanoparticles (AgNPs) from leaf and stem crude extracts was also investigated. Microscopic investigations revealed the presence of lepidote and glandular trichomes, and non-articulated unbranched laticifers on/in the leaves and stems of C. gratissimus. The lepidote trichomes formed a dense indumentum over the abaxial surface of leaves throughout all developmental stages, canopying the underlying glandular trichomes. Laticifers were present in the leaves and stems and were predominantly associated with the vascular tissue in both organs. All structures stained positive for alkaloids, phenolic compounds and lipids with histochemical tests. Phytochemical analyses of the leaves and stems revealed alkaloids, amino acids, phenolic compounds, flavonoids, carbohydrates, terpenoids, saponins and fixed oils and fats in both leaf and stem extracts. The methanolic leaf and stem extracts demonstrated weak to strong activities against various bacteria strains, which are attributed to the several bioactive compounds identified from Gas Chromatography-Mass Spectrometry (GC-MS) analyses. In addition, AgNPs were successfully biosynthesised from the methanolic leaf and stem extracts. Particles synthesised from both extracts were spherical in shape, but their size distribution differed between organs. Antibacterial assays demonstrated a stronger activity of particles from leaf extracts compared to those from stems. These findings corroborate the use of C. gratissimus in traditional medicine and indicate that various structures are involved in the production of bioactive compounds which contribute to the medicinal properties of this plant. Furthermore, the antibacterial activities exhibited by the extracts and AgNPs suggest that C. gratissimus is a potential source of antibacterial agents.
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    Medicinal properties and micromorphology of Rauvolfia caffra Sond.
    (2017) Govender, Valentina.; Naidoo, Yougasphree.; Naidoo, Gonasageran.
    Medicinal plants are effective treatments for various ailments and conditions due to the fact that they are easily accessible, cost effective, trusted and have little to no side effects. Plants produce bioactive or secondary compounds that serve as a defence mechanism to deter pests, attract pollinators and continue to assist in the survival of the species. These compounds or phytochemicals are useful to humans in the form of natural medicine, Rauvolfia caffra Sond. (Apocynaceae) is no exception. The traditional use of the bark is to alleviate skin ailments. A decoction of the bark is consumed for the treatment of abdominal discomfort, abscesses, pneumonia and fever. The research problem is that R. caffra has not been fully documented and the traditional uses cannot be supported scientifically. Furthermore, micromorphological characteristics remain to be described. This is a key component since R. caffra synthesizes latex and transports it through laticifers. The micromorphological characteristics were described by stereomicroscopy, scanning electron microscopy, light microscopy and histochemical tests on the leaves and petioles. This detected the presence of laticifers. To investigate the medicinal value, preliminary phytochemistry and antibacterial screening was performed on crude extracts of the stems and leaves. Methanol, chloroform and hexane were used as solvents of extraction and the classes of compounds detected were alkaloids, glycosides, sterols, flavones and flavonones. Thin layer chromatography provided a visualization of the classes of compounds present. The methanolic stem extract was found to inhibit seven strains of bacteria including E. coli and methicillin-resistant Staphylococcus aureus. The findings do not discredit the traditional utilization of this plant. Additionally, silver nitrate was combined with the crude methanol and water extracts of the stem and leaves to determine if silver nanoparticles (AgNPs) can be formed using a protocol that is safe, not toxic to the environment and simple to carry out. Three different mixing ratios (1:1, 1:2 and 1:4) were used to discover the optimum conditions for synthesis and the extracts screened for their antibacterial activity. The AgNPs synthesized ranged from15.84nm to 34.99nm in diameter which falls within a range that is preferred in nanoscience. The water stem 1:4 AgNPs was found to inhibit two different strains of bacteria, viz. methicillin-resistant Staphylococcus aureus and Klebsiella pneumonia. Rauvolfia caffra does have the potential to be used in drug formulation and in nanotechnology to treat prevalent health problems in South Africa.
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    Selection and micropropagation of solanum nigrum genotypes with varying calcium and iron content.
    (2018) Goordiyal, Kimerra.; Shaik, Shakira.; Watt, Maria Paula Mousaco Deoliveira.
    A direct organogenesis protocol was established for Solanum nigrum using leaf explants from seedling plants. The post acclimatisation yield of the seedling-derived leaf explants was 25 plants/explant. It included decontaminating the leaves with 1 % (v/v) sodium hypochlorite and Tween 20® (10 min), shoot multiplication on medium containing 3 mg l-1 benzylaminopurine (BAP) for 4 weeks, elongation on medium containing 0.1 mg l-1 BAP for a week, rooting on hormone-free Murashige and Skoog medium for 3 weeks and acclimatisation in pots (1 soil : 2 vermiculite [1S : 2V]) in a growth room for 2 weeks. A population of fifty 6-week old seedlings were screened using Inductively Coupled Plasma-Optical Emission Spectrometry. They varied in leaf calcium (Ca) (331.05-916.30 mg 100 g-1 dry mass [DM]) and iron (Fe) (0.64-14.95 mg 100 g-1 DM) contents. Based on these results, genotypes for high Ca (G5 and G20), high Fe (G6 and G15), low Ca (G43 and G45) and low Fe (G35 and G50) were selected for further investigation. These were micropropagated using the established protocol to determine whether their clones maintained similar levels of Ca and/or Fe to those of their parents when grown in soil. Micropropagation influenced the Ca and Fe levels of the clones of the selected genotypes, i.e. the 6-week old clones of six (i.e. G5, G20 and G45 for Ca; and G6, G15 and G50 for Fe) out of the eight selected genotypes had either significantly higher (G45 and G50) or lower (G5, G6, G15 and G20) levels of Ca and/or Fe than their 6-week old parents when grown in soil. There were also genotypic differences regarding the in vitro and ex vitro growth responses (i.e. percentage of explants with shoots, number of shoots/explant and post acclimatisation yield) and leaf Ca and Fe levels of the clones of the selected genotypes. The Ca and Fe contents of the clones of most of the selected genotypes were not affected by substrate type, suggesting that both soil and 1S : 2V were adequate to grow the S. nigrum genotypes for the benefit of high Ca and Fe. Four of the selected genotypes (viz. G15, G20, G35 and G43) were then chosen to investigate the effect of physiological age on their leaf Ca and Fe contents when grown in soil and 1S : 2V. Advancing age only affected the Fe levels of the G35 clones in soil and G15 clones in both the tested substrates. The significant differences in the Ca (G20>G43) and Fe (G15>G35) contents of the 6-week old parents were compared with those of their clones at 4, 6 and 8 weeks ex vitro in soil. A similarity (i.e. G20>G43) in the ‘ranking’, was only found at 4 weeks ex vitro for Ca. Initial attempts to establish a minimal growth protocol for storing germplasm of the eight selected genotypes showed that in vitro shoots could be kept on medium containing ¼ MS + 5 g l-1 sucrose for 8 weeks. After this period, the in vitro shoots of the eight selected genotypes that were placed onto shoot multiplication medium produced 7-13 shoots/explant. It can be concluded that micropropagation and genotype influenced both the Ca and Fe levels of the selected genotypes, while physiological age only influenced their Fe content. As the present study was a smaller component of a larger research program, further investigations need to be carried out on the selected S. nigrum genotypes prior to distribution to community gardens. Future work should include evaluating the effects of various environmental conditions (i.e. growth in a greenhouse, shadehouse, glasshouse and/or in the field), different light intensities, watering regimes, fertiliser treatments and soil pH on the growth, and leaf Ca and Fe levels of the clones. Regarding minimal growth storage, future work should include investigating whether the germplasm of the eight selected genotypes can be stored under the same conditions for longer than 8 weeks.
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    Roles of ROS scavenging enzymes and ABA in desiccation tolerance in ferns.
    (2018) Mkhize, Kwanele Goodman Wandile.; Beckett, Richard Peter.
    The fern species Loxogramme abyssinica and Crepidomanes inopinatum occur widely in KwaZulu Natal Afromontane forests. Although both species are suspected to be desiccation tolerant (DT), little is known about their mechanisms of tolerance. The present study was undertaken to characterize DT in both ferns, specifically the roles of the reactive oxygen species (ROS) scavenging enzymes superoxide dismutase (SOD) and peroxidase (POX), sugar afccumulation, and abscisic acid. Broadly speaking, DT mechanisms are either constitutive (always present) or induced in response to stress. C. inopinatum dries rapidly because it is a “filmy” fern, lacking a cuticle; in contrast L. abyssinica possesses a cuticle, and therefore dries slowly. It was predicted that the fast-drying C. inopinatum would rely mainly on constitutive mechanisms, while the slow-drying L. abyssinica would depend on inducible mechanisms. Plant were collected from the field, transported to the laboratory and then subjected to desiccation. Two methods of desiccation were used. The first one was a relatively mild, slow and short term (48 h) desiccation over calcium acetate, and the second was a harsher, rapid and long term (1 week) desiccation over silica gel. Measurement of chlorophyll fluorescence parameters showed that both species displayed rapid recovery during rehydration after slow or fast desiccation, confirming that both species are genuinely poikilohydric. POX activity remained constant in both species during slow desiccation and subsequent rehydration, suggesting that ROS scavenging by POX is a constitutive DT mechanism. However, the absolute POX activity of C. inopinatum was much higher than in L. abyssinica. Rapid long term desiccation reduced POX activity in both species, but the activity recovered during rehydration. In both species, slow desiccation increased SOD activity, and activity declined to original values during rehydration. Slow and fast drying increased the concentrations of soluble sugars in both species, and concentrations rapidly declined to initial values during rehydration. ABA pretreatment had little effect on DT in either species, although tolerance was slightly increased in L. abyssinica. Results of this study suggested that both species depend largely on inducible DT mechanisms. Counter to the original hypothesis, inducible mechanisms occur even in filmy ferns that desiccate rapidly. The results from this project will contribute to our understanding of how ferns can survive in stressful environments in South Africa, and potentially could help improve abiotic stress tolerance in crop plants.