Electronic Theses and Dissertations (Masters)
Permanent URI for this collectionhttps://hdl.handle.net/10539/38018
Browse
Search Results
Item Metagenome sequencing of the lichen species Flavopunctelia flaventior and Parmotrema tinctorum from Gauteng, South Africa(University of the Witwatersrand, Johannesburg, 2024-06) Katane, Malebogo Dimpho; Botes, Angela; De Maayer, PieterLichens are defined as a mutualistic association between fungi (mycobiont) and an algal and/or cyanobacterial photobiont. Increasing evidence suggests that lichens comprise more diverse microorganisms than initially thought, where lichens represent an interaction between archaea, bacteria, filamentous fungi, green algae, yeasts, and viruses. Not many comprehensive studies have been done of South African lichen species. The present study employed metagenome sequencing to investigate the lichen microbiomes of Flavopunctelia flaventior and Parmotrema tinctorum sampled from Bryanston, Gauteng province, South Africa. Furthermore, the roles played by the members of the lichen microbiome within symbioses were also studied by functionally annotating the assembled metagenomes of the two lichen species. This study sets the groundwork for future studies on South African lichen species. In Chapter 1, an extensive literature review on lichens, their ecology, taxonomy and biology is discussed. Furthermore, it delves into the existence and shape of the microbiome beyond the mycobiont and the photobiont. Additionally, possible roles that the lichen microbiome may play in sustaining the lichen symbiosis is also discussed. In Chapter 2, the metagenomes of two lichen species were sequenced, the quality of the reads were assessed, and taxonomic classification was performed to elucidate the composition of microorganisms associated with each lichen species. Both microbiomes were dominated by bacteria, with limited fungi, viruses, and archaea. The majority of the identified phyla and genera were found to be common between the two lichen species. Similarities in the core microbiome was accounted for by the fact that F. flaventior and P. tinctorum were sampled from the same location and they are both members of the Parmeliaceae family. In Chapter 3, the metagenomic reads were assembled and functionally annotated using various bioinformatics tools. We demonstrate that the members of the lichen microbiome are involved in the cycling of nutrients such as carbon and nitrogen. We also found differences in carbon fixation pathways, which were attributed to the accessory microbiome. Finally, a summary highlights key results and recommendations on future work that could be undertaken to further provide insight into biological pathways essential to sustain the lichen symbiosis.Item Exploring the Structure, Function and Stability of Glutathione Transferases Engineered from Intra- and Inter-class Consensus Sequences: How Forgiving is Nature?(University of the Witwatersrand, Johannesburg, 2024-10) Mulenga, Thabelo; Achilonu, Ikechukwu; Sayed, YasienProtein folding is an enigmatic biochemical process that is foundational to the structural and functional requirements of a cell. The problem of protein folding, in a nutshell, concerns itself with the rate of protein folding as well as the conversion of amino acids from a linear sequence to a fully folded structure. This problem is partly answered by the existence of folding pathways. The folding funnel was conceptualised as a depiction of folding pathways, and it is a framework that illustrates that native proteins naturally favour the lowest energy state, encountering kinetic and thermodynamic barriers as they fold. Consensus protein design, based on this understanding, aims to: (1) enhance stability and (2) navigate the pitfalls of folding by modifying the folding funnel of a protein. This approach can also shed light on the significance of evolutionarily conserved residues. In this study, consensus protein mutants were generated for the Alpha and Mu glutathione transferases (GSTs) classes. The consensus proteins were then benchmarked against the parental proteins that were chosen (hGSTA1-1 and hGSTM1-1). The Alpha consensus mutant had 11 consensus mutations, including a notable M50L mutation, which affects the dynamic behaviour of helices α2 and α9, while the Mu consensus mutant had 13 unique mutations. Protein production and purification showed that the Mu consensus mutant had larger and purer yields. Data from far-UV circular dichroism studies and root-mean-squared-fluctuation (RMSF) from molecular dynamics (MD) simulations showed that the secondary structural components of the Alpha and Mu proteins remained largely the same, although the Alpha consensus mutant displayed a far lower molar residue ellipticity reading than its wildtype counterpart, indicating the disruption of secondary structural elements, likely caused by the M50L mutation. The ANS binding results showed that the M50L mutation in the Alpha consensus protein caused an increase in exposure of the surface area of the H-site, while the Mu consensus protein had a decrease in the solvent accessibility of its H-site. Thermal shift assay results indicated the consensus proteins had increased thermal stability. Enzyme kinetics results showed that the functionality of the proteins was severely diminished in the consensus mutants, particularly the Alpha consensus mutant. MD simulation results showed that there was an overall increase in the rigidity and compactness of the consensus mutant proteins, further affirming the improvement of thermal stability, while signalling the loss in functionality. The results produced herein have the potential to facilitate the proliferation of engineered GSTs for biotechnological applications that require proteins with an increased half-life and greater stability.Item Diversity and Abundance of Arthropods on Conventional Sugarcane under Field Conditions in South Africa(University of the Witwatersrand, Johannesburg, 2024-09) Smith, Roshay; Malinga, Lawrence; Bouwer, GustavInsect diversity and abundance are often the base for formulating strategies that involve the appropriate application of pest control methods, considering the ecosystem services provided by insects. Therefore, the aim of this study was to provide recent baseline data on the diversity and abundance of insects in conventional sugarcane based on two sugarcane fields in KwaZulu-Natal. Three sampling methods, namely pitfall, sticky and water pan traps, were used to sample insects in rain-fed and irrigated sugarcane in Gingindlovu and Pongola from March to October 2022. This study collected 12 493 insects belonging to 14 insect orders and 88 families in rain-fed sugarcane and 22 309 insects belonging to 14 orders and 94 families in irrigated sugarcane. Significant differences in the diversity indices were found between the sampling methods and the sampling periods. This study provides recent baseline data on the diversity and abundance of insects in sugarcane.Item Isolation and characterisation of entomopathogenic fungi(University of the Witwatersrand, Johannesburg, 2024) Kwinda, Fhatani; Lephoto, Tiisetso E.The purpose of the study was to isolate and identify fungal isolates in soil samples, followed by virulence characterisation to study their effectiveness in controlling insect pests using Tenebrio molitor as our model organism. Lastly, a combination study was conducted to evaluate the effectiveness of the joint use of two entomopathogenic microorganisms. For isolation, T. molitor was used as bait then the isolated fungal isolates were identified using molecular and morphological characterisation. Morphological characterisation included macroscopic (fungal cultures) and microscopic (conidia shape and size) analysis while molecular characterisations included extraction of DNA, amplification of the internal transcribed spacer region and sequence alignment. Once identification was done, virulence was assessed through in-vitro virulence parameter like vegetative growth and in-vivo assessment where bioassays were done against T. molitor. Lastly, entomopathogenic fungi were combined with Cruznema sp. NTM-2021 in a soil assay. From the study, two of the five isolates were identified as entomopathogenic fungi, Metarhizium anisopliae ARSEF 7487. M. anisopliae had the slowest vegetative growth but was the highest in virulence. When used for a single application in a soil environment it reaches 97.8% mortality and its combination with Cruznema sp. NTM-2021 resulted in a 57.8% mortality and an additive interaction. In conclusion, M. anisopliae used alone was effective in its control of T. molitorItem The genome sequence of the Yellow-billed Duck (Anas undulata)(University of the Witwatersrand, Johannesburg, 2024-01) Ngxamani, Namhla; De Maayer, Pieter; Mollett, JeanNo abstract given