Thesis (Ph. D.)--University of Queensland, 2002. / Includes bibliographical references.

Bananas (Musa sp.) serve as a staple diet and source of income for millions of people worldwide. The crop, however, is vulnerable to several important diseases such as Fusarium wilt, caused by the soilborne fungus Fusarium oxysporum f.sp. cubense (Foc) . During the mid-20th century, thousands of acres of export banana plantations were lost in Central America due to Fusarium wilt. The epidemic was brought under control only by replacing the highly susceptible Gros Michel banana with Cavendish cultivars. Cavendish bananas, however, are susceptible to Foe race 4, a highly virulent variant of the pathogen that also causes disease to banana varieties susceptible to Foc races 1 and 2. Only this time, no resistant replacement varieties acceptable to the commercial market exist, while cultural and/or chemical control strategies proved to be ineffective for disease control. The only sustainable solution, thus, would be the improvement of existing banana varieties that are acceptable to consumers for resistance to Fusarium wilt. An understanding of how plants defend themselves against pathogens is an imperative first step towards the development of disease-resistant plants. Two broad defence mechanisms against pathogen attack exist in plants: Constitutive defence mechanisms that are pathogen non-specific and induced plant defence responses following recognition of specific pathogen features. A primary objective of this study was to elucidate induced banana defence responses. Induced defence responses may include the induction of regulator and antifungal proteins, the production of active-oxygen species or products from the phenylpropanoid pathway. A few studies have attempted to elucidate the genetic factors involved in the banana defence response but to date no clear answer has arisen. Forward and reverse genetics encompass approached by which plant defence responses can be studied; a particular technique of interest which can be classified under both forward and reverse genetics is cDNA-Amplified Fragment Length Polymorphism (AFLP) In a field trial conducted in an Foc-infested field in Kiepersol, South Africa, cultivars Rose and Calcutta IV proved resistant, and FHIA-17 tolerant to Foe 'subtropical' race 4 (VCG 0120) when compared to the highly susceptible Cavendish cultivar 'Williams'. The genetic basis of defence to Foc in FHIA-17, Rose and Calcutta IV was verified using quantitative real-time reverse transcriptase PCR (qRT-RT-PCR). Catalase, POX and PAE were strongly up-regulated in the tolerant and resistant banana varieties. These genes are involved in the oxidative burst and secondary metabolism leading to the phenylpropanoid pathway and cell wall strengthening. Resistance to Foc, 'subtropical' race 4, thus seems to depend on the early recognition of the pathogen and subsequent blocking of its progress into the plant's vascular system. The response of resistant and tolerant banana varieties to Foc was elucidated by analysing the banana transcriptome 6 and 72 hours post inoculation (hpi) using cDNA-AFLP. Seventy-six differentially expressed transcript derived fragments (TDFs) were isolated, sequenced and subjected to BLASTX and BLASTN searches. Many of the sequences were not significantly similar to any other sequences in the databases, but several genes fragments showed homology to defence-related genes. TDFs representing genes such as S-adenosylmethionine synthase (SAMS) and isoflavone reductase, which are potentially involved in the production of cell wall strengthening compounds such as lignin, were identified. Expression patterns of selected TDFs as seen on the cDNA-AFLP gels were confirmed using qRT -PCR. As additional endogenous controls, two TDFs which displayed constitutive expression on the cDNA-AFLP gels, were isolated, sequenced and were optimised for use as endogenous control genes for the normalisation of the qRT-PCR data. In most cases, the expression patterns seen on the cDNA-AFLP gels were replicated by qRT-PCR. This study concludes that the tolerant (FHIA-17) and resistant (Rose and Calcutta IV) banana varieties induced defence-related genes upon attack by Foc and that the cDNA-AFLP technique was further effective in identifying additional defence-related genes. Looking to the future, the greatest understanding of the defence responses induced during the banana/Foe interaction would be obtained by using additional molecular approaches or techniques. It is in some cases sufficient to look exclusively at transcriptomic i.e. cDNA-AFLP and microarray data to study the plant's response, however, a look at differential protein and metabolite expression would complement transcriptomic data and add insight into the fate of certain expressed genes. This would possibly speed up the identification of the defence pathways used by resistant bananas to resist infection by Foc, once identified these pathways can be manipulated in the susceptible plants and thus the generation of a Foc resistant banana could finally become a reality. / Dissertation (MSc)--University of Pretoria, 2008. / Microbiology and Plant Pathology / unrestricted

Presented as Thesis (Ph. D.)--University of Wisconsin--Madison, 1920. / Title from added collective thesis title page. Reprinted from American journal of botany. Part 1: vol. 10, no. 2 (Feb. 1923), Part 2: vol. 10, no. 3 (Mar. 1923). eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

No description available.

Thesis (Ph. D.)(Genetics)--University of Pretoria, 2003. / Title from opening screen (viewed 8th April, 2005). Includes summary. Includes bibliographical references.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1956. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 51-52).

Distribution of Fusarium wilt of muskmelon and watermelon in southwestern Ontario was studied. Particular attention was paid to morphological and physiological variations of the isolates obtained. Morphological variations were based on comparison in culture with a selected standard. Physiological variations were detected by pathogenicity experiments, and a study of assimilation of various carbon and nitrogen compounds. Some further aspects of the biology of the organisms were investigated. An experiment was carried out, employing several muskmelons and watermelon varieties, to compare their resistance under field conditions. / Thesis / Master of Science (MS)

Thesis (M.Sc.)(Microbiology)--University of Pretoria, 2005. / Includes summary. Includes bibliographical references. Available on the Internet via the World Wide Web.

Bananas are susceptible to a diverse range of biotic and abiotic stresses, many of which cause serious production constraints worldwide. One of the most destructive banana diseases is Fusarium wilt caused by the soil-borne fungus, Fusarium oxysporum f. sp. cubense (Foc). No effective control strategy currently exists for this disease which threatens global banana production. Although disease resistance exists in some wild bananas, attempts to introduce resistance into commercially acceptable bananas by conventional breeding have been hampered by low fertility, long generation times and association of poor agronomical traits with resistance genes. With the advent of reliable banana transformation protocols, molecular breeding is now regarded as a viable alternative strategy to generate disease-resistant banana plants. Recently, a novel strategy involving the expression of anti-apoptosis genes in plants was shown to result in resistance against several necrotrophic fungi. Further, the transgenic plants showed increased resistance to a range of abiotic stresses. In this thesis, the use of anti-apoptosis genes to generate transgenic banana plants with resistance to Fusarium wilt was investigated. Since water stress is an important abiotic constraint to banana production, the resistance of the transgenic plants to water stress was also examined. Embryogenic cell suspensions (ECS) of two commercially important banana cultivars, Grand Naine (GN) and Lady Finger (LF), were transformed using Agrobacterium with the anti-apoptosis genes, Bcl-xL, Bcl-xL G138A, Ced-9 and Bcl- 2 3’ UTR. An interesting, and potentially important, outcome was that the use of anti-apoptosis genes resulted in up to a 50-fold increase in Agrobacterium-mediated transformation efficiency of both LF and GN cells over vector controls. Regenerated plants were subjected to a complete molecular characterisation in order to detect the presence of the transgene (PCR), transcript (RT-PCR) and gene product (Western blot) and to determine the gene copy number (Southern blot). A total of 36 independently-transformed GN lines (8 x Bcl-xL, 5 x Bcl-xL G138A, 15 x Ced-9 and 8 x Bcl-2 3’ UTR) and 41 independently-transformed LF lines (8 x Bcl-xL, 7 x BclxL G138A, 13 x Ced-9 and 13 x Bcl-2 3’ UTR) were identified. The 41 transgenic LF lines were multiplied and clones from each line were acclimatised and grown under glasshouse conditions for 8 weeks to allow monitoring for phenotypic abnormalities. Plants derived from 3 x Bcl-xL, 2 x Ced-9 and 5 x Bcl-2 3’ UTR lines displayed a variety of aberrant phenotypes. However, all but one of these abnormalities were off-types commonly observed in tissue-cultured, non-transgenic banana plants and were therefore unlikely to be transgene-related. Prior to determining the resistance of the transgenic plants to Foc race 1, the apoptotic effects of the fungus on both wild-type and Bcl-2 3’ UTR-transgenic LF banana cells were investigated using rapid in vitro root assays. The results from these assays showed that apoptotic-like cell death was elicited in wild-type banana root cells as early as 6 hours post-exposure to fungal spores. In contrast, these effects were attenuated in the root cells of Bcl-2 3’ UTR-transgenic lines that were exposed to fungal spores. Thirty eight of the 41 transgenic LF lines were subsequently assessed for resistance to Foc race 1 in small-plant glasshouse bioassays. To overcome inconsistencies in rating the internal (vascular discolouration) disease symptoms, a MatLab-based computer program was developed to accurately and reliably assess the level of vascular discolouration in banana corms. Of the transgenic LF banana lines challenged with Foc race 1, 2 x Bcl-xL, 3 x Ced-9, 2 x Bcl-2 3’ UTR and 1 x Bcl-xL G138A-transgenic line were found to show significantly less external and internal symptoms than wild-type LF banana plants used as susceptible controls at 12 weeks post-inoculation. Of these lines, Bcl-2 3’ UTR-transgenic line #6 appeared most resistant, displaying very mild symptoms similar to the wild-type Cavendish banana plants that were included as resistant controls. This line remained resistant for up to 23 weeks post-inoculation. Since anti-apoptosis genes have been shown to confer resistance to various abiotic stresses in other crops, the ability of these genes to confer resistance against water stress in banana was also investigated. Clonal plants derived from each of the 38 transgenic LF banana plants were subjected to water stress for a total of 32 days. Several different lines of transgenic plants transformed with either Bcl-xL, Bcl-xL G138A, Ced-9 or Bcl-2 3’ UTR showed a delay in visual water stress symptoms compared with the wild-type control plants. These plants all began producing new growth from the pseudostem following daily rewatering for one month. In an attempt to determine whether the protective effect of anti-apoptosis genes in transgenic banana plants was linked with reactive oxygen species (ROS)-associated programmed cell death (PCD), the effect of the chloroplast-targeting, ROS-inducing herbicide, Paraquat, on wild-type and transgenic LF was investigated. When leaf discs from wild-type LF banana plants were exposed to 10 ìM Paraquat, complete decolourisation occurred after 48 hours which was confirmed to be associated with cell death and ROS production by trypan blue and 3,3-diaminobenzidine (DAB) staining, respectively. When leaf discs from the transgenic lines were exposed to Paraquat, those derived from some lines showed a delay in decolourisation, suggesting only a weak protective effect from the transgenes. Finally, the protective effect of anti-apoptosis genes against juglone, a ROS-inducing phytotoxin produced by the causal agent of black Sigatoka, Mycosphaerella fijiensis, was investigated. When leaf discs from wild-type LF banana plants were exposed to 25 ppm juglone, complete decolourisation occurred after 48 hours which was again confirmed to be associated with cell death and ROS production by trypan blue and DAB staining, respectively. Further, TdT-mediated dUTP nick-end labelling (TUNEL) assays on these discs suggested that the cell death was apoptotic. When leaf discs from the transgenic lines were exposed to juglone, discs from some lines showed a clear delay in decolourisation, suggesting a protective effect. Whether these plants are resistant to black Sigatoka is unknown and will require future glasshouse and field trials. The work presented in this thesis provides the first report of the use of anti-apoptosis genes as a strategy to confer resistance to Fusarium wilt and water stress in a nongraminaceous monocot, banana. Such a strategy may be exploited to generate resistance to necrotrophic pathogens and abiotic stresses in other economically important crop plants.

Thesis (Ph.D.(Plant Pathology)--University of Pretoria, 2006. / Includes bibliographical references. Available on the Internet via the World Wide Web.