Theses and Dissertations (Plant Production and Soil Science)

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Vegetative propagation of Pelargonium sidoides D.C. (Rabassam) using leaf-bud cuttings
Pholo, Mafamo Samuel (University of Pretoria, 2019-02)
Pelagonium sidoides (Rabassam) is an herbaceous plant of the Geraniaceae family used throughout history as a medicine for the people of South Africa which has and recently also been exported to other parts of the world. A number of studies have shown that the plant extracts have anti-microbial, anti-viral and immune boosting properties. Consequently, it has been harvested in the wild for these properties, exposing the species to possible extinction. This scenario can be addressed by artificially mass propagating and producing the species to satisfy demand. One method of propagating the plant is to use leaf-bud cuttings. Unfortunately, vegetative propagation of the species using leaf-bud cuttings is not documented. This study was therefore conducted to investigate the influence of rooting medium (sand and sand + coir), rooting hormone (0.1% indole-butyric acid (IBA), planting depth (1.5 and 3 cm) and season (summer, autumn, winter and spring) on rooting success of leaf-bud cuttings as well as impacts of these treatments on the survival and morphological characteristics of subsequent transplants. The experiments were conducted on a mist bed in a greenhouse located at the Experimental Farm of the University of Pretoria (25o45’S, 28o16’E). A 23 factorial experiment laid out in a randomized complete block design (RCBD) with 3 replicates was carried out. Root length, root number, planting depth, growth media and hormone application, were assessed by destructive sampling at 5 to 25 days after planting (DAP). Results of the study demonstrated that sand medium, compared to a mixture of sand + coir medium, resulted in higher rooting percentage and more roots. However, sand + coir medium produced longer roots on the cuttings. Planting depth of 3 cm resulted in higher rooting percentage and better establishment of cuttings as compared to a depth of 1.5 cm. Application of the rooting hormone 0.1% IBA improved the number of roots but not rooting percentage nor root length. Rooting of cuttings was improved when propagated in autumn (longer roots) and spring (higher number of roots) than in summer or winter. After one month transplanting of cuttings, significant differences (P < 0.05) were noted in survival percentage due to the media initially used. Cuttings initially propagated on sand showed a higher survival percentage (78.6%) and had a higher number of roots, which were longer, compared to cuttings initially propagated in a medium of sand + coir (61.2%). Cuttings initially propagated in sand and treated with hormone had significantly more roots after transplanting. Application of hormone also improved root and shoot length of transplants. Based on observations, cuttings taken from younger mother stock plants had a higher chance of survival than cuttings taken from mature stock plants. In general, leaf-bud cuttings of P. sidoides can be artificially propagated without rooting hormone but the success rate will be low. The ideal regime is to propagate cuttings treated with IBA in a sand medium.
Expression analysis of selected maize primary metabolism genes in response to Cercospora zeina inoculation in the glasshouse
Mlunjwa, Zimbili Ethel (University of Pretoria, 2018-09)
Maize is one of the most important grain crops in Africa especially in Southern and sub-Saharan Africa. This crop contributes not only to food security but also to the economy. However, there are factors that affect the development of maize including diseases, which pose as threats to food security. Grey leaf spot (GLS), a foliar disease of maize, is caused by Cercospora zeina, an economically important ascomycete worldwide. It causes over 65% yield losses, thereby being a great threat to food security. This disease was first reported in KwaZulu-Natal province, a GLS hot spot in South Africa in 1988. Strategies such as crop rotation, conventional tillage and fungicides are widely used to manage the progression of GLS. However, these strategies are labour-intensive and expensive for farmers. An alternative cost-effective method is the utilization of host resistance. To better understand the mechanism of host resistance, two primary metabolism genes, isocitrate lyase (ICL) and malate synthase (MS), involved in the glyoxylate pathway, were previously shown to be highly induced by the fungus in a field RNAseq study. This research was aimed at elucidating expression of these two primary metabolism genes. Glasshouse trials were conducted by inoculating the maize genotypes B73 (susceptible) and B73-QTL (resistant) with C. zeina. Genomic DNA was extracted and the fungal CTB7 gene region amplified to confirm that the infection was due to C. zeina. RNA was also extracted, and reverse transcribed to copy DNA (cDNA). The quality of RNA was confirmed by performing a reverse transcriptase PCR (RT-PCR) with banana actin primers as a positive control. Primer pairs flanking an intron in the ICL and MS genes were successfully designed to specifically amplify their respective cDNA. To further confirm these results, the amplified gene products were sequenced using Sanger sequencing platform and thereafter aligned to respective reference genes obtained from NCBI. Overall, ICL and MS genes were successfully amplified from the infected maize genotypes. The ICL amplification needs further optimisation as the bands were not intense to continue with expression analysis. The expression of the two MS transcripts increased at the different progression stages indicated that the glyoxylate pathway plays a role in fungal pathogenicity.
The role of soil hydrophobicity in the cause and maintenance of the mysterious Namibian fairy circles
Hurter, Jan Willem (University of Pretoria, 2018-07)
Circular barren patches, also known as fairy circles (FCs), occur throughout Namibia, devoid of vegetation and surrounded by a matrix of grasses. Several hypotheses regarding the origin and maintenance of FCs have been proposed, none widely accepted. Soil physical properties and chemical constituents were investigated to determine whether differences in properties are present and to what extent they differ. Results obtained from physical property analyses indicated greater hydrophobicity in soil collected from FCs than matrix soil as well as a higher infiltration rate. Extracts prepared from FCs and matrix soil, and from locations where Euphorbia damarana are decomposing (DP), were analysed by GC-MS and NMR to determine the cause of the increased infiltration rate and the FC soil’s hydrophobicity. To identify discriminative signals from GC-MS and NMR data, PCA-plots, OPLS-plots, and S-plots were created to aid compound identification. The PCA-plot created from GC-MS results displayed clear discrimination, segregating matrix (M) sample observations from DP and FC sample observations, simultaneously grouping DP and FC observations. This indicates greater similarity between DP and FC soil, than between DP and M or FC and M observations. Specific compounds were present in both FC and DP soil samples and not in matrix soil. NMR based metabolomics indicated similar concentrations of ester, phenol, alkene and aromatic functional groups within FC and DP soil, while differences between FC and matrix soil were greater. From these results, it is deduced that there’s a definite difference in physical properties and chemical constituents of FC and matrix soil. Causes of these differences can be concluded to be initiated by decomposing Euphorbiaceae species, as indicated by specific compounds such as 1-(4-acetamidoanilino)-3,7-dimethylbenzo[4,5]imidazo[1,2-a]pyridine-4-carbonitrile present in FC and DP soil samples, previously identified in a variety of plants’ non-polar latex. It is thus proposed that the formation of FCs in northern Namibia are indirectly caused by decomposing E. damarana, which alters the soil physical properties at the decomposition site, increasing hydrophobicity and infiltration rates in soil upon decomposition which It is thus proposed that the formation of FCs in northern Namibia are indirectly caused by decomposing E. damarana, which alters the soil physical properties at the decomposition site, increasing hydrophobicity and infiltration rates in soil upon decomposition which decreases the amount of plant available soil water content and leads to the formation of barren patches, devoid of any vegetation, in the same circular shape as the E. damarana plants which once grew at that location. This mechanism of formation is proposed as the most likely cause in other areas of Namibia, where succulent Euphorbia spp. are present.
The City of Tshwane Agri-parks initiative as a case study for the sustainable intensification of smallholder agriculture in South Africa
Hay, Richard Lee (University of Pretoria, 2023)
Over the next decade, sub-Saharan Africa (SSA) is predicted to experience the largest relative population increase of any subcontinent. The region is already greatly impoverished and the least food secure in the world, with 58.7% of the population classified as at least moderately food insecure. Smallholder farmers are the foundation of SSA’s agricultural sector, producing the majority of the region’s grains and legumes, and almost all of the tuberous and root crops. However, the Green Revolution that resulted in dramatic increases in yields across most of the globe is largely considered to have failed in SSA, and yield gaps in the region remain some of the largest in the world. Sustainable Intensification (SI) has been proposed as the most appropriate means to increase production of smallholder farmers in a way that is both environmentally sustainable and appropriate for the context and complexities of smallholder agriculture in SSA. Numerous examples of SI initiatives across SSA, such as complex cropping systems, small-scale irrigation schemes, and the use of improved germplasm, have demonstrated the potential for increasing yields, reducing the environmental impact of agriculture, and improving the livelihood of smallholder farmers. However, a number of socioeconomic factors still limit widespread adoption. One initiative that aims to overcome these challenges and intensify smallholder agriculture through state-funded development is the Agri-parks programme, which aims to provide farmers with financial and institutional support for a period of ten years, while developing linkages across the agricultural supply chain to ensure long-term economic viability. On paper, the Agri-parks initiative appears to be a model, scalable SI programme. However, five years since the launch of the Agri-parks programme, various challenges and delays as well as a disconnect between provincial and national governmental departments has resulted in significant delays in the programme’s timeline. The Agri-park in the City of Tshwane (CoT) was found to be limited in size and functionality, with much of the infrastructure originating from previous projects on the sites and in various states of disrepair. Nonetheless, the Agri-parks programme still holds potential to become a flagship SI programme if it is developed to the extent the original model describes. One of the major limiting factors of small-scale irrigation schemes in SSA is the relatively high costs of equipment, which are economically out of reach for most smallholder farmers. Examples from elsewhere in the world have shown that, given the right institutional and financial support, smallholder farmers can greatly benefit from investment into irrigation technologies. In recent years a number of tools, such as the Wetting Front Detector and the Chameleon sensor, have been developed specifically to help smallholder farmers better manage their irrigation water resources. These tools formed part of the research projects technology transfer through field trials at the Soshanguve and Rooiwal Farmer Production Support Units (FPSUs); however, uncontrolled circumstances disrupted the field trials at both sites but illustrated the daily struggles faced by the farmers at these sites. In order to replicate the conditions at the CoT Agri-park FPSUs in a more controlled environment, a trial to investigate the impacts of different mulching treatments and weeding practices on weed emergence and pressure was conducted. The trial revealed that mulching treatments had no significant effect on the total weed biomass (p > 0.05) and that weeding had a significant effect on weed population density (p < 0.001) and weed species richness (p < 0.001). However, consultation with the farmers of the Rooiwal FPSU revealed that mulching was not an appropriate strategy for them as they felt the mulch made it more difficult to weed the field. Weed pressure on the site was high, but farmers were hesitant to use chemical control measures. This illustrated the importance of providing comprehensive agricultural extension, to ensure farmers have access to knowledge of scientific best practices on which to base their management decisions. A second trial to investigate the impact of weeding frequency on yield and water productivity indicated that bi-weekly weeding can result in yields 4.2 times greater than yields obtained in unweeded fields (p < 0.001) and irrigation water productivities 4.1 times greater than irrigation water productivity obtained in unweeded fields. This reinforced the need for the use of basic agronomic practices, such as weeding, to increase production outputs and improve resource use efficiencies. The current limited size of the Agri-parks mean it is unlikely to be viable to appoint a dedicated extension officer to the CoT Agri-park, but social media has the potential to connect a number of Agri-parks to a single extension officer instead. In order to explore the viability of social media as a platform for hosting e-learning programmes, a 14 ‘chapter’ learning programme on the fundamentals of weed science was run through the ‘Ingesta: Farming for the Future’ Facebook page. Thirty-four individuals from six different countries, as well as eight of South Africa’s provinces, successfully completed the learning programme. Feedback from the learning programme was overwhelmingly positive, with 90% of the final participants finding the programme to be at least ‘easy to understand’ and 91% indicating the length of the chapters was ‘just long enough’. All of the final participants indicated that they had learnt information that they felt would better help them manage weeds in their fields, with 17 expressing additional positive comments. As a result, the pilot study was considered a success and social media proven to be a viable platform to provide access to agricultural science learning material for smallholder farmers in SSA. Although there is insufficient evidence to support that the CoT Agri-park can currently be considered an example of SI, the initiative does still have the potential to be if there is a coordinated approach to develop the Agri-park as the original model describes, and the incorporation of various tools and technologies will enhance support to smallholder farmers.
In-field climatic factors contributing to sunflower development and health across planting dates
Mapfumo, Phrasia (University of Pretoria, 2024-03-31)
This thesis explores the influence of climatic factors and planting dates on sunflower growth, yield, and disease dynamics. Chapter 1 reviews the literature on the optimal climatic conditions for sunflower development, highlighting the impact of deviations on morphological, floral, and yield traits. The study emphasizes the significance of understanding climatic influences on Sclerotinia head rot (SHR), a major disease affecting sunflower production in the Republic of South Africa (RSA). Although climatic factors influencing SHR are well-documented, the role of planting dates in SHR progression is less explored. Chapters 2 and 3 investigate these climatic factors, revealing that earlier planting dates generally lead to higher yields and that sunflowers demonstrate resilience under hotter, drier conditions. The impact of extreme weather events, such as the February 2021 heat wave, on sunflower growth and floral traits is examined, highlighting adaptive mechanisms in response to heat stress. Additionally, Chapter 4 introduces Bidens mottle virus (BiMoV) as a newly identified pathogen in RSA, with significant implications for sunflower yield and development. Finally, Chapter 5 analyzes SHR progression across planting dates, finding that consistent temperatures above 27°C during flowering limit disease establishment. These findings provide critical baseline data for optimizing planting strategies and managing disease risks in the context of climate variability and change.
Elucidating the role of microbial factors and water stress in causing corky crack blemishes on potato tubers in South Africa
Gush, Sasha-Lee (University of Pretoria, 2020-04)
Potato (Solanum tuberosum L.) is the most consumed and economically important vegetable crop in the world and is a staple crop in many communities. However, the potato crop is susceptible to devastation by various diseases that reduce the yield and quality of fresh produce. Blemishes, rots and deformation are some of the tuber symptoms caused by fungi, bacteria, viruses and nematodes. Abiotic factors such as water stress (dry conditions and thereafter high moisture conditions), has also been associated as causal agents of tuber blemishes. Potato tuber blemishes constitute a persistent quality problem in the production of potato throughout the world and can be a result of known causes (typical blemishes), which are manageable; or unknown causes (atypical blemishes), which are difficult to manage. Atypical corky crack blemishes are an emerging problem of potatoes in South Africa. There has been a large debate in the South African potato industry regarding the causal agents of corky cracks. This study investigated the role of microbial factors and water stress in causing corky crack blemishes in South Africa. In a pilot trial, R. solani AG2-2IIIB and various Streptomyces species were isolated from corky crack blemishes on potatoes in South Africa. However, none of these microorganisms, alone or in combination resulted in the development of corky cracks when tested in pot trials. Therefore, in an effort to confirm the causal agent(s) of corky crack blemishes, additional tubers with symptoms of either growth cracks or corky cracks were sampled from different potato growing regions of South Africa and further isolations of bacteria and fungi were made from the different symptoms. Water stress and the detection of Potato Virus Y (PVY) was also studied to describe the association with corky crack blemishes in South Africa. Binucleate Rhizoctonia (BNR) AG-A and BNR AG-R as well as Fusarium oxysporum were the most predominant fungal species isolated from the corky crack symptoms, while no fungal species were isolated from growth cracks. Streptomyces collinus, S. yaanensis, S. corchorussi, S. viridochromogenes and S. griseorubens were the most predominant bacterial species isolated from growth cracks and no Streptomyces species were isolated from corky cracks. The Streptomyces species identified are not known to be pathogenic to potatoes and are, therefore, not likely causal agents of the corky cracks. PVY was not amplified from corky crack tissue using molecular techniques and it was, therefore, assumed that PVY is not associated with corky cracks on the tested samples of potato tubers in South Africa. Greenhouse pathogenicity trials using the isolated microorganisms alone and in combination were done to confirm Koch’s postulates. Corky crack blemishes were not observed on progeny tubers in single inoculations for fungal or bacterial isolates; however, corky crack blemishes were observed on the progeny tubers inoculated with a combination of Rhizoctonia species. This suggests a synergistic interaction between the three Rhizoctonia species tested in this study. The findings of this study are crucial in the development of effective integrated strategies for the management of corky crack disease and to improve the tuber quality in the South African market and globally.
Strategies to bridge yield gaps in equatorial African upland rice (Oryza sativa L.) production systems
Alou, Isaac Newton (University of Pretoria, 2020-02)
The average rice (Oryza sativa L.) farmer in sub-Saharan Africa (SSA) harvests less than 1.5 t ha-1 of grain yield in uplands production systems, while the yield potential is substantially higher. There are large yield potential gaps due to nutrient (especially nitrogen [N]) and water stress (WS). The main study objectives were to determine the effects of WS imposed at different growth stages (GS) on upland rice performance, to evaluate and compare nitrogen and water use efficiencies, to calibrate and test a crop simulation model for predicting water uptake and yield under a wide range of agroecological conditions and to quantify rice yields, and yield gap for the equatorial climate in Uganda and propose adaptive management strategies for improving yields. Field experiments were conducted between 2013 and 2016 using two upland rice varieties commonly grown in SSA. Crop parameters were estimated from measured data for modelling purposes. The SWB-Sci model was parameterised, calibrated and tested using independent data from two seasons and secondary data from Ugandan research sites. Simulation studies were then performed for diverse rice growing areas along the equator (lying 0.10oS–3.28o N and 31.13o E–34.16o E) over the period 2008–2012. Grain yield measured under well-watered, adequately fertilised conditions for the medium-duration variety (Nerica 4) was 7.2 t ha-1, and 4.5 t ha-1 for the short-duration variety (Nerica 10). When water was withheld during tillering (Ti), anthesis and grainfilling for Nerica 4 it resulted in severe WS, but yield penalties were minimal (<25%), compared to a 75% yield loss with stress during panicle initiation. Considerable water savings (176–245 mm) are possible with WS during the non-sensitive GS. Increasing N level altered tiller development, reduced thermal time to key GS, increased water use by 17–33% and grain N uptake per unit water used of Nerica 10, compared to the zero-N treatment. Use efficiencies for input resources declined with N rates above 120 kg N ha-1. The calibrated SWB-Sci model generally predicted water uptake, growth and yield of both varieties under different treatment conditions well, with little error and bias, for both Hatfield and Ugandan research sites. The attainable (Yt)/potential yield (YP) ratio ranged from 0.04 to 0.59 between locations. If N limitations are alleviated, water-limited yield (Yw) /YP ratio values of Nerica 10 (0.37–0.98) were generally higher than for Nerica 4 (0.08–0.86) across agroecological zones (AEZs). Yield gaps of upland rice varieties were variable and specific to AEZs. Inter-seasonal differences were very apparent in the bimodal and transition rainfall zones. The gaps were small for the Eastern Savannah Moist (Yw/YP = 0.78 ± 0.03) and the Northern Moist Farming Systems (0.75 ± 0.03). Adaptive cropping tactics to increase yield and annual rice production for WS-prone zones were identified. The use of the model should be useful in future studies to identify specific agronomic practices to increase WUE. Information can be used to drive policy on upland rice, for instance government initiatives to intensify rice production.
Antimicrobial and antimalarial activity of Tabernaemontana elegans Stapf. (Apocynaceae)
Mkatshane, Zimasa (University of Pretoria, 2019-08)
Medicinal plants play a crucial role in the healthcare treatment of many infections and also serve as the basis for the synthesis of novel chemotherapeutic agents. Included in these plants is the Tabernaemontana genus (Apocynaceae) comprising of about 18 species in Africa of which two (Tabernaemontana elegans and T. ventricosa) are indigenous to South Africa. These plant species are used by indigenous people to treat various diseases including venereal diseases, malaria cancer, tuberculosis, stomachache and infertility. The aim of the study was to validate some of the ethnomedicinal uses of T. elegans as well as to isolate and identify the compounds responsible for their activity. In this study, different plant parts of T. elegans were collected based on their medicinal uses for venereal diseases and malaria by Vha-Venda people residing in Mutale Municipality, Limpopo Province. The plant parts were extracted sequentially using hexane, dichloromethane, methanol, water and decoctions were prepared separately. The recovered crude extracts were tested against Neisseria gonorrhoeae, Candida albicans, human immunodeficiency virus (HIV) and Plasmodium falciparum. Additionally, the plant extracts were subjected to an antiproliferative bioassay against mammalian skeletal myoblast cells. Based on the in vitro screening results, the plant extracts were then subjected to 1H-NMR-based metabolomics in an attempt to discriminate between selective and non-selective antiplasmodial plant extracts. The best antiplasmodial selective plant extract (stem bark) was further subjected to phytochemical analysis using conventional chromatographic and spectroscopic techniques. For the antigonoccocal assay, the positive control was ciprofloxacin with MIC value <0.05 mg/ml. The best antigonoccocal activity was observed from the methanol extract of roots with MIC value of 0.10 ± 0.04 mg/ml and methanol extract of the stem bark with MIC value of 0.20 ± 0.06 mg/ml. Root decoction also inhibited bacterial growth with MIC value of 0.80 ± 0.37 mg/ml. An activity of 6 mg/ml was observed in the water extract of the stem bark. This was the first study to report on significant antigonoccocal activity of T. elegans. Ciprofloxacin was also used as the positive control for the anticandidal assay and displayed an MIC value of <0.05 mg/ml. The extracts displayed an activity of >6 mg/ml for the anticandidal assay. The root extracts (water and dichloromethane) showed a moderate inhibition of 60.78 ± 0.16% and 50.24 ± 0.11% when tested against HIV 1 reverse transcriptase (RT) enzyme. The decoction and water extracts of leaves showed moderate inhibition activity of 58.32 ± 0.05% and 54.00 ± 0.12% and the stem bark also showed moderate inhibition activity in the decoction and water extracts with activity of 57.06 ± 0.03% and 51.29 ± 0.13%, respectively. This was the first study to report on the anti-HIV activity of T. elegans. The positive control (doxorubicin) displayed 95.83 ± 0.005% inhibition of the tested enzyme. Six plant extracts exhibited significant antiplasmodial activity ranging from 0.16 ± 0.01 – 4.62 ± 0.55 μg/ml with methanol extracts of stem bark (IC50 = 0.31 ± 0.01 μg/ml) and methanol root (IC50 = 0.16 ± 0.01 μg/ml) being the extracts with the best activity. The positive control, chloroquine, displayed an antiplasmodial activity of 0.004 ± 0.002 μg/ml. The plant extracts were subjected to 1H NMR-based metabolomics to discriminate between the selective and non-selective antiplasmodial plant extracts. An activity profile that could be used to predict activity of unknown samples was observed. Phytochemical analysis was conducted on the combined non-polar extracts (hexane and dichloromethane) of the stem bark which were the most active plant extracts. Four compounds were isolated and were subjected to further NMR analysis in an attempt to predict antiplasmodial activity and for the corroboration of their respective structures. Two of the compounds (2 and 3) were predicted to have significant antiplasmodial activity based on activity profile generated from metabolomics. The study supported some (malaria and gonorrhoea) of the ethnomedicinal uses of T. elegans by indigenous South African people and demonstrated that metabolomics could be used in predicting antiplasmodial activity of unknown samples. Further studies will focus on identification and in vitro screening of the four compounds as well as isolating more compounds.
Unraveling the potential of selected South African indigenous plants as biocontrol agents against aflatoxin-producing Aspergillus species
Chauke, Sipho Hayley (University of Pretoria, 2019-11)
Aspergillus flavus and A. parasiticus are saprophytic storage fungi, which primarily invade maize (Zea mays) and other cereal and legume crops. Contamination of stored grains with these fungal species reduces the quality of food and feed, which leads to yield loss. Moreover, A. flavus and A. parasiticus present a serious threat to human and animal health due to the production of aflatoxin B1 (AFB1), which is carcinogenic and is associated with stunting among children. Current control methods may be effective; however, due to the adverse residual effects associated with the use of synthetic chemical fungicides, alternate methods of pest control such as biodegradable agents are being researched. Plant extracts have been tested to be less toxic and environmentally friendlier compared to synthetic chemical fungicides. The goal of this study was to evaluate three plants, namely, Erythrophleum lasianthum, Heteropyxis natalensis and Warburgia salutaris for their antifungal activities and as seed treatments against known AFB1-producing A. flavus and A. parasiticus isolates in vitro. The extracts were screened using the broth microdilution method to obtain the minimum inhibitory concentration (MIC) values, of which the acetone, ethanol and water extracts of W. salutaris (0.117 mg/mL), E. lasianthum (0.234 mg/mL) and H. natalensis (0.469 mg/mL), respectively, showed noteworthy antifungal activity (≤ 1 mg/mL) against A. flavus. In contrast, the acetone extract of W. salutaris and the ethanol and water extracts of H. natalensis showed noteworthy antifungal activity (≤ 1 mg/mL) against A. parasiticus with MIC values of 0.117, 0.938 and 0.469 mg/mL, respectively. Following the antifungal screening, E. lasianthum (ethanol), H. natalensis (water) and W. salutaris (acetone) extracts were evaluated for their potential as AFB1 reducers at 15, 30 and 75 mg/mL extract concentrations. Using spectrophotometry, the extract which reduced AFB1 at 75 mg/mL the best was that made from the acetone extract of W. salutaris when compared to all the other tested extracts. Warburgia salutaris (75 mg/mL) was able to reduce AFB1 by 49.27 % in liquid media while AFB1 was least reduced by E. lasianthum (at 15 mg/ml), which was only able to reduce AFB1 production by 12.82 %. Furthermore, the selected five extracts, E. lasianthum (acetone), H. natalensis (acetone, ethanol and water) and W. salutaris (acetone) which showed good antifungal activity (against A. flavus and A. parasiticus) were screened for their potential toxicity on HeLa and HepG2 cell lines. All the extracts showed moderate to no cytotoxicity (> 50 μg/mL) on both cell lines with the exception of H. natalensis (water) and W. salutaris (acetone) extracts, which were toxic (≤ 50 μg/mL) to the HepG2 cell line, at the highest tested concentration (400 μg/mL). Bioassay-guided fractionation of the ethanolic extract of H. natalensis indicated that the liquid-liquid partition butanol fraction was the most active against A. flavus and A. parasiticus. The butanol liquid-liquid partitioned fraction was further subjected to isolation and identification using Column chromatography and GC-MS, revealing five major compounds as potential fungal growth inhibitors present in the butanol fraction of H. natalensis. Artificially inoculated maize seeds with A. flavus were treated with the three plant extracts, E. lasianthum (ethanol), H. natalensis (water) and W. salutaris (acetone), at 50 and 100 mg/mL and evaluated for their potential as seed treatments. The treated seeds were plated onto Potato Dextrose Agar (PDA) prior to storage, and three and six months after treatment. The extracts had no observable negative effects on seed germination. The extracts showed no significant fungal growth inhibition on treated maize seeds plated out prior to storage. However, after three months of storage, H. natalensis (water) and W. salutaris (acetone) at 100 mg/mL showed significant growth inhibition on maize treated seeds with fungal growth inhibition percentages of 65.00 and 70.50 %, respectively. After six months in storage, E. lasianthum (100 mg/mL) and H. natalensis (50 and 100 mg/mL) exhibited intermediate fungal growth inhibition of 48, 47 and 55 %, respectively. Therefore, these plants indicated the potential to be used as biological control agents against fungal contaminants of grains such as A. flavus in storage.
Pharmacokinetics and hepatoprotective effects of Lippia scaberrima Sond. and Aspalathus linearis (Burm.f.) R. Dahlgren
Kok, Anna-Mari (University of Pretoria, 2020)
In developing countries, the presence and impact created by tuberculosis (TB) is alarming. With the rise of resistant TB, initiatives such as adjunct host-directed therapy (AHDT) that aim to reduce treatment time, lower side-effects and costs involved, should be investigated. The Dutch East Indian Company mainly drank herbal medicinal teas in the Cape region during the settlement of the Cape. Many of the indigenous plants within the immediate area were used as substitutes for the much more expensive and imported tea. Lippia scaberrima Sond. (L. scaberrima) is an under investigated shrub which is endemic to South Africa. Although Lippia javanica may have been an obvious choice for testing in this regard, the fact that not much is known on L. scaberrima prompted the investigation into the biological properties of this specific species. Aspalathus linearis (Burm.f.) R. Dahlgren (A. linearis) a medicinal plant that has a variety of biological properties documented. Both these plants are consumed as teas and in this study an attempt was made to investigate their combined biological and therapeutic activity. The biological investigation in the present study included using a combination of ethanolic extracts of both L. scaberrima and A. linearis. L. scaberrima is known to be rich in volatile compounds and the essential oil was isolated and included in the study. The essential oil of L. scaberrima was incorporated as another sample to be tested for its biological activity, as this plant is known to be rich in volatile compounds. This study further included an investigation of the biological activity of verbascoside, a known compound, found in all endemic Lippia species. Lastly, the evaluation of synthesized gold nanoparticles from L. scaberrima were also included. Significant antimycobacterial activity was found for L. scaberrima against Mycobacterium tuberculosis H37Rv. Low to no cellular toxicity was observed against hepatocellular carcinoma cells (HepG2) at the tested concentrations. Hydro-steam distillation was effective in isolating the essential oils from the aerial parts of L. scaberrima, which showed effective hepatoprotective activity against acetaminophen-induced toxicity. The cytochrome P450 (CYP) inhibitory activity of L. scaberrima, combinations of L. scaberrima and A. linearis, and verbascoside has been reported in this study for the first time. Potent CYP inhibitory activity was found for L. scaberrima in combination with the green A. linearis. Verbascoside had no significant inhibitory activity against the CYP isoforms and therefore, it can be considered that it may have low risk for herb-drug interactions. This study included the cyclooxygenase-II (COX-II) inflammatory activity of L. scaberrima. All the samples, except L. scaberrima, that were investigated for their DPPH free radical scavenging potential were found to have significant activity. Moderate nitric oxide scavenging activity was found for verbascoside and green A. linearis in combination with L. scaberrima. Antioxidant and anti-inflammatory activity play a profound role in assisting with the adverse effects experienced during treatment for TB infection. This study provided a more comprehensive look into the specific biological properties of L. scaberrima relating to TB infection as well as its efficacy when combined with fermented and green A. linearis extracts. Synthesized nanoparticles of L. scaberrima provided the opportunity to lower the cytotoxicity towards HepG2 cells even more than was initially found. The investigated samples showed the potential to decrease drug-induced hepatotoxicity, one of the main adverse effects experienced. In addition, all the samples tested, showed potential as an adjuvant for host-directed therapies for TB. These initial results pave the way for further research into specialized avenues with endless possibilities for the discovery of new specialized treatment regimens that are tailored to be host specific and for specific stages of infection. Please note that all experiental studies and results reported on were on the ethanolic extracts of both L. scaberrima and A. linearis and not of the tea and/or the boiling water extract of L. scaberrima and A. linearis or otherwise stated.
Identification and control of Colletotrichum species associated with Eucalyptus seeds
Mangwende, Edgar (University of Pretoria, 2019-12)
The South African forestry industry largely relies on seed for its replanting programs following clear-felling. The high cost of Eucalyptus seed and planting targets makes it imperative to start with pathogen-free seeds to avoid inconsistences of emergence and development of seedlings. Together with 34 other fungal species, two Colletotrichum species in the Colletotrichum gloeosporioides species complex were found to be naturally associated with 14 commercial Eucalyptus seed lots. Eucalyptus nitens seed lot had the highest incidence of fungi (92.4%), whereas the lowest incidence occurred on E. dorrigoensis seed lot (28.9%). Seed germination of seed lots inoculated with seed-borne fungi was less than 62% and as low as 25%. Multiloci phylogenetic analyses of ITS, β-tubulin, actin, and glyceraldehyde-3-phosphate dehydrogenase gene regions identified Colletotrichum isolates PPRI 24314 as C. fructicola Prihastuti. and PPRI 24315 as C. kahawae Waller and Bridge. Biochemical tests based on utilisation of ammonium tartrate confirmed the latter isolate as C. kahawae subsp. cigarro. Colletotrichum fructicola and C. kahawae subsp. cigarro were naturally associated with seed lots of E. dunnii (3.5 and 0.5%, respectively), E. nitens (2.6 and 1.2%, respectively) and E. macarthurii (0.8% for only C. fructicola). Greenhouse pathogenicity tests showed that both C. fructicola and C. kahawae subsp. cigarro caused the highest incidence of anthracnose leaf spot on E. nitens (65 and 55%, respectively), with a severity of 67 and 63%, respectively. Sowing seeds inoculated with C. fructicola demonstrated the pathogen to be vertically transmitted into E. camaldulensis, E. dunnii and E. nitens seedlings (25.5, 38.3 and 64.0%, respectively), whereas C. kahawae subsp. cigarro inoculated seeds transmitted the pathogen in 27.3, 30.5 and 56.3% of the seedlings, respectively. Since both pathogens were seed-borne and seed-transmitted, non-chemical seed treatments that included Bacillus and Trichoderma, hot-water, microwave radiation and hydrogen peroxide were evaluated for their efficacy at sanitising seed lots of E. nitens and E. viminalis artificially inoculated with either C. fructicola or C. kahawae subsp. cigarro. Soaking Eucalyptus spp. seed lots in 15% H2O2 for 10 min effectively reduced incidences of Colletotrichum spp. but negatively reduced seed germination. Instead, soaking Eucalyptus spp. seeds in 10% H2O2 for 5 min and 10 min were optimal treatment/time combinations with an acceptable reduction of Colletotrichum spp. and significantly high seed germination percentages. Hot water optimum treatment/time parameters were 50 °C for 30 min against C. kahawae subsp. cigarro, and 55 °C for 15 min against C. fructicola. Exposure of moist Eucalyptus spp. seeds to microwave radiation at powers levels of 1 400 w for 30 s significantly improved seed germination similar to that of the Celest® XL treatment. However, sowing seeds treated with H2O2, hot water and microwave radiation failed to control anthracnose leaf spot developing on seedlings grown under greenhouse conditions. Seeds treated with a Bacillus biocontrol agent consistently reduced incidences of Colletotrichum spp. on Eucalyptus spp. and effectively suppressed appearance of anthracnose leaf spot on seedlings grown under greenhouse conditions. Due to the high efficacy of Celest® XL and Bacillus in-vitro and in-vivo, they can be recommended as sanitisers of commercial Eucalyptus seed lots against anthracnose leaf spot.
Cloning the Cercospora zeina Ecp2 effector gene for Agrobacterium-mediated transient transformation
Segal, Carol-Ann Crystal (University of Pretoria, 2019)
Maize (Zea mays) is a staple crop in Africa that is under severe threat to disease by pathogenic organisms. Grey leaf spot (GLS) caused by Cercospora zeina drastically limits the yield and quality of maize produced. Not enough is understood about how C. zeina causes GLS, but it is known that it is a maize-specific hemibiotrophic fungus. Proteins called effectors are essential for the virulence of pathogens such as Cladosporium fulvum. Extracellular protein 2 (Ecp2), an effector identified in some Dothideomycete fungi, has an unknown function but has been shown to play a role in the virulence of the fungi. The aim of this study was to clone C. zeina Ecp2 (CzEcp2) into a binary vector for agroinfiltration of Nicotiana spp. The C. zeina genome and RNA sequence data (in planta and in vitro) were searched for a candidate Ecp2 gene. The complete CzEcp2 sequence (with the fungal signal peptide) and the mature sequence (lacking the fungal signal peptide) were cloned into the pTRAkc-ERH binary vector. pTRAkc-ERHCzEcp2 EB (fungal signal peptide) and pTRAkc-ERHCzEcp2 NB (LPH signal peptide) were respectively transformed into Agrobacterium tumefaciens GV3101 (pSOUP+pMP90). Phytophthora infestans INF1 was used as the positive control for HR expression. Untransformed Agrobacterium and the pTRAkc-ERH empty vector were used as negative controls. Nicotiana benthamiana, Nicotiana tabacum cv. Petit Havana and Nicotiana tabacum cv. LA Burley were then transiently agroinfiltrated. The plants were monitored for a hypersensitive response (HR) for 10 days. CzEcp2 expression did not result in HR for the three Nicotiana spp., but chlorosis was observed. INF1 caused a HR in all three Nicotiana spp. and the negative controls did not cause any changes. The lack of HR where CzEcp2 was expressed, may be due to lack of CzECP2 transport and recognition, a delayed HR or that the T-DNA was not adequately transferred into the host cells.
The microbiome of Agaricus bisporus (Lange) Imbach and the prevalence of foodborne pathogens and mycoparasitic fungal pathogens in white button mushroom production systems
Rossouw, W. (Werner) (University of Pretoria, 2019-12)
Food safety related disease outbreaks and challenges could cause serious crop and market related losses in any industry. This is also true for the commercial mushroom sector in South Africa. Disease outbreaks adversely affect economic viability of the industry, on both the production and consumer margins. In order to secure the economic feasibility of the industry this study focussed on production practices and diseases that contribute towards losses. Included in this research are investigations into method development for foodborne and mushroom antagonistic organisms in production systems. An important research objective was to understand the dynamics of the microbiological significance in each major aspect of the mushroom production chain to determine the viability of early detection systems and ultimately prevention of disease outbreaks. Different molecular techniques were optimized and employed to confirm prevalence and persistence of the various microorganisms identified to be of importance. Successfully developed and implemented aspects of the work performed in this study will be directed to the South African mushroom industry for further development of a holistic mushroom quality and safety management programme. Not only will it provide a better understanding of the mushroom microbial ecology, prevalence of foodborne pathogens and accurate identification of mushroom pathogens, but also contribute to improved product quality and safety for consumers. These aspects have a level of novelty that has the potential to offer the South African industry with solutions to improve disease management and overall safety standards.
The host response of maize towards Exserohilum turcicum and its toxin, monocerin
Kotze, Richard Gavin (University of Pretoria, 2020)
Northern leaf blight (NLB) is a devastating foliar disease of maize (Zea mays L.) throughout the maize growing regions of the world. The causal agent of NLB is the hemibiotrophic fungal pathogen, Exserohilum turcicum. Exserohilum turcicum produces a secondary metabolite, monocerin, which is phytotoxic and could aid the fungus in causing NLB in maize. This study was conceptualised to assess the infection strategy of E. turcicum in maize, as well as the host response of maize towards E. turcicum and its toxin monocerin. The infection strategy of E. turcicum was evaluated through the use of high resolution light (LM), scanning (SEM) and transmission electron microscopy (TEM) to obtain a better understanding of the hemibiotrophic lifestyle of the fungus. During the biotrophic phase of fungal infection, the disease was characterised by chlorotic spots whereas cigar shaped lesions formed during the necrotrophic phase. Infection structures as well as conidiophores were observed for the first time through SEM. At 9 days post inoculation (9 dpi) the fungus was observed in the epidermal cells, visible in the xylem at 11 dpi, at 14 dpi the xylem was almost completely blocked, and at 18 dpi conidiophores formed through the stomata, and the fungus completed its life cycle. The results of this study provide updated insight into the infection strategy of the fungus in maize as well as supporting previous findings that E. turcicum is a hemibiotrophic pathogen. Pathogenesis-related (PR) proteins are one of the many defence mechanisms plants use to protect themselves against fungal infection. Reverse transcription-quantitative PCR was applied to evaluate whether PR protein genes were upregulated in maize in response to E. turcicum infection and the presence of the E. turcicum toxin, monocerin. Expression of selected PR protein genes (PR-1, PR-2, PR-3, PR10) associated with fungal infection was induced in response to the fungus but only during the necrotrophic phase of the fungal growth. Monocerin did induce the gene expression of PR proteins but at a low level when compared to the fungus. PR-10 (ribonuclease-like) was the only PR protein gene which was induced at a higher level by monocerin as compared to the fungus. The phytotoxic effects of monocerin on the maize leaf cell ultrastructure were studied using LM and TEM. The cytoplasm as well as the vacuole and chloroplast were most affected by the phytotoxic nature of monocerin. The chloroplast was the most sensitive to the toxin due to disruption of the double-membrane, stroma and thylakoid membranes. As monocerin treatment caused an over accumulation of starch granules in the chloroplast, the gene expression of enzymes (gwd, pwd, amy3) involved in degradation of starch granules in the chloroplast was assessed following fungal infection and monocerin treatment. Expression of the all the starch degradation enzymes genes was inhibited during fungal infection but only amy3 was inhibited by monocerin treatment. Response of the maize host to E. turcicum infection and monocerin infiltration provided new understanding in the host-pathogen interaction which could be exploited in developing new control strategies against NLB in maize.
Molecular genetics of resistance to Spongospora subterranea f. sp. subterranea infection in potato (Solanum tuberosum L.)
Lekota, Moleboheng (University of Pretoria, 2020-01)
Infection of potato by Spongospora subterranea (Wallr.) Lagerh. f. sp. subterranea Tomlinson (Sss) causes tuber lesions, which directly affect the quality and marketability of the tubers, as well as root infections, which are associated with tuber yield and weight reductions. The pathogen is also a vector of potato mop-top virus (PMTV), which is considered economically important in some potato growing countries. Management of Sss is difficult because of the ineffectiveness of the current control options and it therefore requires integration of different control measures, of which host resistance represents the most economical, long-term approach. However, the information on the mechanisms of potato resistance to Sss infection is limited. In this study, the molecular and biochemical mechanisms of different potato cultivars that vary in their susceptibility to Sss infection were assessed. Potato cultivars commonly grown in South Africa were assessed for their differential responses to Sss root and tuber infection. A greenhouse pot trial was conducted to evaluate the susceptibility of 10 cultivars to Sss infection. Visual observation was performed to determine the severity of root galling and powdery scab infection, while root microscopic examination was carried out to assess root zoosporangia infection. Quantitative polymerase chain reaction (qPCR) was used to confirm and measure the quantity of Sss DNA in the inoculated and not-inoculated potato roots and tubers. No potato cultivar was found to be resistant to Sss root or tuber infection in this study, as all cultivars had zoosporangia and root galls on or in the roots, and powdery scab lesions on tubers, with the exception of cultivar Innovator, which did not show any powdery scab symptoms on progeny tubers. However, a range of susceptibility of potato cultivars to root infection, root gall formation and powdery scab, from tolerant to susceptible was observed in the study. There was no association between development of root diseases and powdery scab. A negative correlation was observed between powdery scab and number of root zoosporangia, with a weak positive correlation observed between powdery scab and number of root galls. Nevertheless, three cultivars; Fianna, Lanorma and Mondial were found to be moderately tolerant to all three diseases. These results were confirmed with qPCR, in which the amounts of Sss DNA confirmed the presence of Sss infection in roots and tubers of all 10 cultivars evaluated. The molecular mechanisms underlying quantitative potato resistance to powdery scab; discovery of defense-related genes was undertaken by ribonucleic acid sequencing (RNA-seq) in a susceptible and tolerant potato cultivar in response to powdery scab infection. Validation of the RNA-seq results and gene expression profile analysis of the two cultivars were carried out using quantitative real-time reverse transcriptase PCR (RT-qPCR) on a subset of genes induced at different tuber developmental stages. Differentially expressed genes related to plant defense responses were identified in both cultivars. Expression of nine genes involved in defense was up-regulated more in the tolerant cultivar than in the moderately susceptible cultivar upon infection by Sss. The involvement of defense-related metabolites in potato root tolerance to Sss root infection was investigated. Metabolic profiling of the chemicals in root exudates and root extracts of potato cultivars differing in susceptibility to Sss root infection, using a non-targeted metabolomics approach, was carried out. Ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS) analysis was used to compare metabolic profiles of tolerant cultivars and susceptible cultivars inoculated or not-inoculated with Sss, to identify and quantify the resistance-related (RR) metabolites involved. Significant differences between the moderately tolerant (Fianna, Innovator, Lanorma, Mondial and Valor) and the moderately susceptible/susceptible (Avalanche, BP1, Ronaldo Sifra and Up-to-date) cultivar groups were identified in the levels of several metabolites, including amino acids, organic acids, alkaloids, phenolics and sugars. These secondary metabolite classes play important roles in plant defense. Monitoring Sss development in potato roots and tubers can enable efficient assessment of Sss infection in potatoes, resulting in proper, timely and improved control strategies for the diseases. Moreover, more information on the chemical composition of roots and root exudates as well as identification of defense-related genes is important for marker-assisted selection of potato germplasm in breeding programs for developing novel potato cultivars with broad and durable resistance to Sss.
Water and nutrient use efficiencies of potato-based (Solanum tuberosum L.) rotation systems in North West Province, South Africa
Mukiibi, Alex (University of Pretoria, 2019-11)
Water and nutrient management has a direct impact on yield, input-use efficiencies of crops, and is crucial to the ecological sustainability of production. Moreover, water-use efficiency (WUE) and nutrient-use efficiency (NUE) are common indicators of ecological sustainability in crop production. Measuring WUEs and NUEs of potato-based (Solanum tuberosum L.) rotation systems, rather than those of potato alone, provides a true image of water and nutrient use as well as the potential environmental pollution associated with potato production. Little is known, however, on the WUE and NUE of potato-based rotation systems within South Africa. This study was conducted to quantify WUE, NUE, drainage and nutrient leaching of potato fields, as well as to evaluate nutrient carry-over effects to the subsequent crop in rotation. Six irrigated potato fields on commercial farms in the North West province were selected for monitoring. Five of the six potato fields were monitored during the 2017/18 summer season, and one field during the 2018/19 season. After the potato season, follow-up crops of paprika (Capsicum annuum L.), groundnuts (Arachis hypogaea L.) and onions (Allium cepa L.) were monitored during the 2018/19 season in three fields. Surveys and field measurements were conducted regarding fertilizer type and application rate for both potato and follow-up crops. Flow meters and pressure transducers were used to monitor irrigation amounts. Drainage lysimeters were installed in four of the six fields to monitor drainage and nutrient leaching. Soil and plant samples were taken for nutrient analysis. Final yields of potato and the follow-up crops were determined at crop harvest. Calculated WUE was based on total water inputs from rainfall and irrigation (WUER+I), potential WUE (PWUE) was based on simulated tuber yield and evapotranspiration (ET), and irrigation WUE (IWUE) was based on observed irrigation. Nutrient-use efficiency was expressed as partial factor productivity (PFP), nutrient utilization efficiency (NUtE) and nutrient harvest index (NHI). Results revealed that average nutrient rates of 300 kg ha-1 N, 220 kg ha-1 P, 386 kg ha-1 K, 580 kg ha-1 Ca, 252 kg ha-1 Mg and 99 kg ha-1 S were applied to potato fields. Irrigation water applied to potato fields varied greatly and ranged from 590 – 1011 mm (average 866 mm) per season. Fresh tuber yields ranged from 60 – 93 t ha-1 (average 83 t ha-1). Potato WUER+I, PWUE and IWUE ranged between 53 – 124, 96 – 151 and 59 – 129 kg mm-1, respectively. Drainage of 488 mm was measured in one potato field and leaching of 29 kg ha-1 N, 20 kg ha-1 K, 484 kg ha-1 Ca, 179 kg ha-1 Mg and 129 kg ha-1 S was recorded. Partial factor productivity of N, P and K was quite similar for all potato fields, with average values of 288, 379 and 229 kg fresh tuber kg-1 of applied nutrient, respectively. A great variation in PFP among fields, however, was observed for Ca, Mg and S. Nutrient utilization efficiency and NHI of potato showed small field differences. Follow-up crop total input rates ranged between 140 – 328 kg ha-1 N, 108 – 284 kg ha-1 P, 171 – 406 kg ha-1 K, 1141 – 1232 kg ha-1 Ca, 405 – 944 kg ha-1 Mg and 165 – 216 kg ha-1 S. In follow-up crops, total water inputs ranged between 805 – 1526 mm (average of 1155 mm) per season. Paprika dry fruit yield of 5.5 t ha-1, groundnut dry grain yield of 3.4 t ha-1 and onion fresh bulb yield of 75 t ha-1 were obtained. The WUER+I for paprika, groundnuts, and onion were 3.6, 3.0 and 11.9 kg dry matter yield mm-1, respectively. In conclusion, the growers achieved high potato yields, although substantial variability occurred between fields. Relatively high water and nutrient input rates were applied to potato and follow-up crops. However, these input rates were not necessarily proportional to crop yields achieved by the growers, which contributed to WUE and NUE variability among fields. The results revealed that the growers have room to improve WUEs and NUEs in potato and the follow-up crops, which can be achieved by using irrigation scheduling tools and adjust nutrient rates based on soil and irrigation water nutrient status.
Developing integrated silvicultural management systems for woodlands of northwestern Zimbabwe
Chichinye, Angella (University of Pretoria, 2019-11)
Silvicultural management systems based on natural disturbance-recovery processes are fundamental so as to integrate multiple-use practices in the natural tree resources inorder to address local needs and global concerns. The study aimed at developing a new understanding of the Baikiaea-Guibourtia-Pterocarpus woodland recovery dynamics when exposed to different disturbances such as single tree selection, harvesting of poles and firewood and cultivation. Four specific objectives were studied to examine different parts of this overall study: Disturbance impacts on the composition and diversity of the Baikiaea-Guibourtia-Pterocarpus woodlands of North-western Zimbabwe; Ecological drivers of floristic and structural composition of Baikiaea-Guibourtia-Pterocarpus woodland communities; Disturbance impacts on regeneration of key ecological and/or economic species in the Baikiaea-Guibourtia-Pterocarpus woodlands; and Age and growth rate determination using growth rings of selected Baikiaea-Guibourtia-Pterocarpus species. Variation in floristic composition of recovering Baikiaea-Guibourtia-Pterocarpus woodlands was studied in different development stages (from early re-growth to mature woodland). TWINSPAN (TWo-way INdicator SPecies ANalysis) was used for classification of plots from different utilization systems and different development stages into communities. One-Way ANOVA in SPSS Version 21 was used to test for differences in species diversity for the different communities. Detrended Correspondence Analysis (DCA) in the CANOCO ordination programme was used to determine the extent of variation amongst the identified communities. Correspondence analysis in SPSS Version 21 Statistical Package was performed to analyse grain status for nine tree communities. Stem diameter class profiles were analysed in STATISTICA version 7 to determine the ratio of regeneration to mature trees of individual canopy tree species across tree communities. . The research revealed that even if the sites provide a potential for woodland of a specific type to develop, plant community formation is mainly influenced by existing utilization systems. The intense disturbance factors of clearing for crop cultivation and to a lesser extent harvesting for poles and firewood are necessary disturbances for the regeneration and growth of the Baikiaea-Guibourtia-Pterocarpus woodlands. The grain status (relative similarity between the composition of canopy species in the regeneration and in the canopy of the same stand) and stem diameter distributions in the different communities suggested that the species require some more intense disturbances, such as with cultivation and harvesting for poles and firewood, to promote regeneration and growth to higher size classes. The study also revealed that disturbances that cause the opening of canopy and more light penetration on the forest floor are required to facilitate sprouting and growth of suppressed shoots in these woodlands. The study concludes that clearing for cultivation and harvesting for poles and firewood are important components to which the woodland ecosystem is adapted and important for the recovery of the Baikiaea-Guibourtia-Pterocarpus woodlands. It is recommended that woodland utilization and management should integrate cultivation and harvesting for poles and firewood into forest management. Also, a comprehensive program is necessary to monitor the levels of cultivation and the impacts on woodland recovery. Cutting cycles should be based on growth rate of the selected species. Timber species harvesting should go side by side with these disturbance factors so as to open up the canopy to allow maximum sunlight for the regeneration stock.
Biocontrol potential of bacteria isolated from Carpobrotus spp. as seed treatments against Fusarium pathogens of maize
Moshabesha, Mantsele Thuso Sylvia (University of Pretoria, 2019-12)
Fusarium spp. cause significant losses in yield and quality in the maize (Zea mays L.) industry. Fusarium pathogens affect the development of the plant tissue and the quality of the grain. The mycotoxins produced by Fusarium spp. are also a threat to the health of animals and humans. This, therefore, has a negative impact on food safety and security of the countries which depend on maize as a staple food crop, such as the Republic of South Africa (RSA). Large yields of maize are necessary in RSA since maize is also an export crop, boosting the gross economic value total of field crops. The demand for sustainable maize production strategies necessitates intervention against Fusarium pathogens. For years, Fusarium pathogens have been controlled through the use of chemical fungicides whose progressive use is hindered by public concerns about the environmental and health safety of accumulated fungicide residues, and development of resistance against fungicides. Possibly safer, more effective and environmentally friendlier alternative control methods like biological control (biocontrol) continue to be evaluated. The application of naturally occuring organisms and their secreted metabolites to reduce disease incidence and severity may be a long-term means of control. The ability of these biocontrol agents (BCAs) to withstand harsh conditions while protecting the maize seed or plant is what is desired for the development of a potential biocontrol product for seed treatment. Bacillus spp. show potential as BCAs as they employ different modes of pathogen antagonism, stimulate plant growth and form stable endospores to help them survive harsh conditions. Bacteria (Bac A and Bac B) isolated from seeds of a mesemb, Carpobrotus acinaciformis (L.) L. Bolus were investigated for their antagonistic potential against Fusarium graminearum Schwabe, Fusarium verticillioides (Saccardo) Nirenberg, and Fusarium solani (Martius) Appel & Wollenweber emend. Snyder & Hansen. In vitro dual culture and two-plate inverted culture assays showed that Bac A and Bac B were able to inhibit all three selected Fusarium spp. by the production of antifungal diffusible and volatile metabolites. Further metabolite characterization was done for Bac A as Bac A showed better inhibition of the Fusarium spp. in vivo, and the antifungal diffusible metabolites produced by Bac A were characterized as terpenoids. The efficacy of Bac A in vivo seed treatment against F. graminearum and F. verticillioides on maize was evaluated on Bac A-treated seeds, which were placed on potato dextrose agar (PDA). The results thereof revealed that 1.5 × 109 CFU was a more efficient inhibitory concentration gainst the Fusarium spp. As Bac A was not effective for control of F. solani in vivo, phytotron investigations of the in vivo potential of Bac A as a BCA for seed treatment was evaluated against F. graminearum and F. verticillioides. Results showed that Bac A enhanced plant shoot height in Trial 1, and dry shoot mass and dry root mass of maize seedlings in the presence F. verticillioides in Trial 2. Bac A also increased shoot height of maize seedlings in the presence of F. graminearum in Trial 1. In addition, Bac A showed no adverse effects on the germination and emergence of maize. Bac A reduced shoot and root rot symptoms caused by F. verticillioides in both Trials 1 & 2. Only in Trial 1 were shoot and root rot severity caused by F. graminearum reduced by Bac A. The shelf-life of Bac A formulation for seed treatment over a storage period of three months was evaluated. Bac A formulation for seed treatment was consequently determined to retain viability and bioactivity of Bac A for the entire three months’ period of storage. Bac A proved to survive with its antifungal activity viable at 4 °C and at 25 °C. However, the antagonistic potential of Bac A after three months of storage was found to better at 4 °C storage conditions. Bac A had no negative impact of the germination of maize seeds after one month, two months, and three months of storage and at both temperatures. Bac A and Bac B showed significant biocontrol potential in that they were antagonistic against the selected Fusarium pathogens associated with maize in vitro. In addition, Bac A showed further protective ability in vivo against root and stem diseases caused by F. graminearum and F. verticillioides, as well as enhanced growth of maize shoot height and dry mass of seedlings. With simple Bac A suspension formulation, Bac A retained a better shelf-life at 4 °C for three months. Bac A has shown potential as a biofungicide and as a potential biofertilizer, which are both beneficial and cost effective in crop protection.
Classification of tree species based on their accumulation and exclusion of heavy metals, and the influence of heavy metals on species abundance on Copperbelt tailings dams in Zambia
Matakala, Nalukui (University of Pretoria, 2019-09)
Classification of tree species into excluders or accumulators is vital for ecological restoration of mine tailings dams via phytostabilization. The study dealt with classification of indigenous tree species found on tailings dams and also how heavy metal concentration influence the abundance and distribution of tree species on tailings Storage facilities in the Copperbelt Province, Zambia. Among the studied species a number of them were classified into excluders and accumulators of the 12 metals. For example, Rhus longipes, Syzigium guineense, Senegalia polyacantha, Ficus craterostoma, Bauhinia thonningii, Albizia adianthifolia, Combretum molle, Peltophorum africanum and Ficus sycomorus, Albizia antunesiana, Albizia versicolor, Azanza garckeana, Bauhinia petersiana, Brysorcapus orientalis, Combretum molle, Combretum microphyllum, Dichrostachys cinerea, Dodonaea viscosa, Ficus capensis, Lannea discolor, Phyllanthus guineensis, Senna singueana, Terminalia mollis, Terminalia stenostachya, Combretum apiculatum and Vachelia sieberiana were classified as Cu excluders while Dombeya rotundifolia, Albizia amara and Parinari curatellifolia, Combretum zeyheri, Annona senegalensis and Ozoroa insignis were classified as Cu accumulators. All the species were classified as Mn accumulators while Albizia adianthifolia, Albizia antunesiana, Combretum apiculatum and Ficus capensis were classified as Zn excluders and the other 28 species classified as Zn accumulators. The dominance of species in terms of importance values varied from (a species with the lowest IVI value) to (Species with the highest IVI value). Species with high IVI values dominated the tailings dams than those with low IVI values. Rhus longipes, Syzygium guineense, Senegalia polyacantha, Ficus craterostoma, Bauhinia thonningii, Albizia adianthifolia and Combretum molle were identified as the most dominant species on Copperbelt tailings dams. Low species richness (composition) and abundance was observed in high metal concentration. Most species showed high affinity to heavy metals in low concentration while a few species (Rhus longipes, Albizia versicolor, Albizia amara Dombeya rotundifolia, Albizia antunesiana, Phyllanthus guineensis and Dichrostachys cinerea) showed high affinity to metals in high concentration. Furthermore, the study identified Ni, Cu, S, Cr, Al, Co, Mn, B, Mo and Cd as metals influencing species richness and abundance on the Copperbelt tailings dams.
Studies of the growth and mineral nutrition of cassava (Manihot esculenta Crantz) grown as an industrial crop
Thabethe, Zethu Sifundo Isiphile (University of Pretoria, 2019-12)
Cassava (Manihot esculenta Crantz) is a substantial contributor to the food and industrial crops sectors in most African, Asian and Latin American countries. However, cassava production in Africa has been threatened by the shortage of improved cultivars and a decrease in soil fertility linked with continuous cultivation, while inadequately applying macronutrients; nitrogen (N), phosphorus (P) and potassium (K). The objectives of this study were to: (i) Assess the growth and yield performance of two local cassava cultivars (P4/10 and MSAF-1) in Mbombela, Mpumalanga, South Africa (SA), (ii) Examine the effects of different combinations of NPK on cassava growth and yield, and (iii) Evaluate the utility of SPAD-meter for assessing N status in cassava plants. The P4/10 and MSAF-1 cultivars growth and yield parameters were evaluated in a field experiment. MSAF-2 cultivar was planted in a greenhouse to optimize N and K applied at 0, 100, 150 and 200 kg ha-1 and P at 0, 15, 20 and 30 kg ha-1. MSAF-2 was also planted in a greenhouse to assess N status of the plants fertilized at N rates of 0, 100, 150 and 200 kg ha-1. SPAD-meter readings collected from five different leaf positions (Lf1 – Lf5) located at the top of the main stem. The results showed that MSAF-1 growth and yield parameters were significantly higher (p < 0.05) than those of P4/10 over time. The differences between the parameters were attributed to genotypic variation since the cultivars were grown under the same conditions. The three-way interaction (N × P × K) had a significant effect (p < 0.05) on the growth and yield over time. Both growth and yield parameters increased with increase in NPK application rates. Growth parameters were promoted by the different NPK combinations, but the best combination for cassava output was 200 kg ha-1 N, 30 kg ha-1 P and 150 kg ha-1 K. SPAD readings were found to be significantly influenced (p < 0.05) by N application rates and leaf position over time. The N application rates and leaf N concentration levels at 98 days after planting (DAT) were positively correlated with the lower leaves (Lf4 and Lf5) together with Lf3, but negatively correlated with uppermost leaves (Lf1 and Lf2). However, the lower leaves readings were almost similar but higher than Lf3 and therefore their average was used to assess nitrogen sufficiency index (NSI). The highest mean NSI values were obtained when 200 kg ha-1 N was applied (NSI = 1.04) compared to 0.99 and 0.88 when 100 and 0 kg ha-1 N was applied.

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