Doctoral Degrees (Plant Pathology)
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Browsing Doctoral Degrees (Plant Pathology) by browse.metadata.advisor "Lennox, C. L."
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- ItemThe biology of Endophyllum osteospermi, and its use for the biological control of Chrysanthemoides monilifera ssp. monilifera(Stellenbosch : Stellenbosch University, 2004-12) Wood, A. R. (Alan Robert); Crous, P. W.; Lennox, C. L.; Stellenbosch University. Faculty of AgriSciences. Dept. of Plant Pathology.ENGLISH ABSTRACT: Chrysanthemoides monilifera ssp. monilifera is a shrub indigenous to South Africa, which has become a serious weed of native vegetation in Australia. Endophyllum osteospermi is a microcyclic, autoecious, rust fungus that induces witches' brooms on C. monilifera ssp. monilifera. This rust is considered as a candidate biocontrol agent for use against C. monilifera ssp. monilifera in Australia. The vegetative growth and reproductive output of healthy branches on bushes with different levels of E. osteospermi infections were measured at three sites. The growth of healthy branches on infected bushes was 26- 81% less than that of healthy branches on uninfected bushes. The number of buds, flowering capitulae, fruiting capitulae, and cypselas on healthy branches of infected bushes was 35-75%, 45-90%, 15-99%, and 15-90% less, respectively, than those on uninfected bushes. At five sites, the infection levels and number of witches' brooms were determined every two months. The increase in number of witches' brooms per bush ranged between o and 282 within one year, with an average increase per bush of28 (SE ± 4.8) and 39 (SE ± 9.2) during two years. The average simple interest rate (rs) increase of infection levels for all bushes was 0.015 month-I (s.e. ± 0.0041, n = 72) and 0.0098 month" (s.e. ± 0.0073, n = 43) during two years. Aecidioid teliospores germinated between 10 and 20oe, with 15°e as optimum. Light, and particularly near-uv light, stimulated germination. A period of 6 to 8 hours of light was needed to obtain optimum germination levels. The temperature requirements for basidiospore development differed from that of aecidioid teliospore germination. Optimum was at 15°e, but a rapid decrease in basidiospore production occurred at higher temperatures, few developed at 19°e. Two nuclear divisions occurred within 12 hours of germination to produce a metabasidium with three or four nuclei. A third nuclear division occurred in the basidiospores between 24 and 48 hours. Plants inoculated under controlled conditions took 5 to 24 months before witches' brooms began to develop. A Geographic Information System (GIS) approach was used to model the potential distribution of E. osteospermi in South Africa, based on monthly average climate surfaces with parameters derived from the above experiments. The same model was applied to Australia to suggest a potential distribution of the rust if released in Australia. This potential distribution was similar to one generated using the climate matching computer programme CLIMEX©, but gave greater spatial accuracy. Both approaches indicate that E. osteospermi should establish in temperate Australia. Chrysanthemoides species, as well as other South African asteraceaus plants, were monitored for E. osteospermi between 1992 and 2003. Endophyllum osteospermi was recorded on C. monilifera ssp. monilifera, C. monilifera ssp. pisifera, C. monilifera ssp. rotundata, C. monilifera ssp. canescens, C. monilifera ssp. subcanescens, C. incana, an undescribed taxon of Chrysanthemoides, Osteospermum ciliatum, 0. polygaloides and 0. potbergense. Endophyllum dimorphothecae sp. nov. is described on Dimorphotheca cuneata. Aecidium elytropappi, which was recorded on Elytropappus rhinocerostis and Stoebe plumose, is transferred to Endophyllum as E. elytropappi comb. nov. Germination of aecidioid teliospores and penetration by basidiospores were observed on the surface of excised leaves of 32 plant species at 4 days after inoculation. Germinating aecidioid teliospores aborted on 14 plant species, whilst no penetration was attempted on a further 12. Penetration only occurred on 9. Therefore only these 9 plant species need to undergo traditional host specificity testing. Pending these results, E. osteospermi could be safely released in Australia for the biological control of C. monilifera ssp. monilifera.
- ItemOptimisation of imazalil application and green mould control in South African citrus packhouses(Stellenbosch : Stellenbosch University, 2014-04) Erasmus, Arno; Fourie, P. H.; Lennox, C. L.; Stellenbosch University. Faculty of AgriSciences. Dept. of Plant Pathology.ENGLISH ABSTRACT: South Africa is the largest exporter of shipped fresh citrus fruit worldwide. One of the major factors that can lead to substantial losses is postharvest decay. Penicillium digitatum (PD) and P. italicum (PI) are the main wound pathogens, respectively causing green and blue mould decay. PD is more prevalent than PI and therefore also the focus in the majority of research in this field. Imazalil (IMZ) is applied by the majority of citrus packhouses through an aqueous dip treatment, and provides good curative and protective control, as well as sporulation inhibition activity. Two IMZ formulations are in use: the sulphate salt applied in aqueous treatments and the emulsifiable concentrate (EC) applied with wax coatings. The majority of research on IMZ has been done using the EC formulation. The maximum residue limit (MRL) for IMZ on citrus fruit is 5 μg.g-1, whereas 2-3 μg.g-1 is regarded as a biologically effective residue level that should at least inhibit green mould sporulation. A study was conducted to assess the current status of IMZ application in South African packhouses, to determine the adequate residue levels needed to control green mould and inhibit sporulation using IMZ sensitive and resistant isolates, and to study optimisation of modes of IMZ application in citrus packhouses. Factors studied were IMZ concentration, application type (spray vs. dip and drench), exposure time, solution temperature and pH, as well as curative and protective control of PD. The packhouse survey showed that the majority of packhouses applied IMZ in a sulphate salt formulation through a fungicide dip tank, and loaded an IMZ residue of ≈1 μg.g-1. In dip applications, IMZ had excellent curative and protective activity against Penicillium isolates sensitive to IMZ. However, curative control of IMZ resistant isolates was substantially reduced and protective control was lost, even at twice the recommended concentration, nor was sporulation inhibited. The use of sodium bicarbonate (2%) buffered imazalil sulphate solutions at pH ±8, compared with pH ±3 of the unbuffered solutions, markedly increased IMZ residue loading on navel and Valencia oranges and improved curative and protective control of IMZ resistant isolates. Exposure time did not affect IMZ residue loading in IMZ sulphate solutions at pH 3, although the MRL was exceeded after 45 s exposure in pH 8 solutions. Imazalil applied through spray or drench application improved residue loading, but green mould control was less effective than after dip application. IMZ formulation (IMZ sulphate and EC), solution pH (IMZ sulphate at 500 μg.mL-1 buffered with NaHCO3 or NaOH to pH 6 and 8) and exposure time (15 to 540 s) were subsequently investigated in order to improve IMZ residue loading and green mould control on Clementine mandarin, lemon, and navel and Valencia orange fruit. As seen previously, exposure time had no significant effect on residue loading in the unbuffered IMZ sulphate solution (pH 3). No differences were observed between the pH buffers used, but residue loading improved with increase in pH. The MRL was exceeded following dip treatment in IMZ EC (after 75 s exposure time), and IMZ sulphate at pH 8 using NaHCO3 (77 s) or NaOH (89 s) as buffer. The MRL was exceeded after 161 s in IMZ sulphate solutions buffered at pH 6 with either NaHCO3 or NaOH. Green mould control as influenced by residue data was modelled to predict control of IMZ-sensitive and IMZ-resistant PD isolates. From this model the effective residue levels for 95% control of an IMZ-sensitive isolate and of an IMZ-resistant isolate were predicted to be 0.81 and 2.64 ug g-1, respectively. The effects of incubation time (infection age), exposure time, solution pH, wounds size and fruit brushing after dip treatments on residue loading and curative green mould control were also investigated. Exposure time did not have a significant effect on residue loading on fruit dipped in pH 3 solutions of IMZ (< 2.00 μg.g-1). Increasing the pH to 6 resulted in significantly increased residue loading, which increased with longer exposure time, but mostly to levels below the MRL after 180 s. Post-dip treatment brushing reduced residue levels obtained in IMZ pH 3 solutions by up to 90% to levels < 0.5 μg.g-1; however, curative control of the IMZ sensitive isolate was mostly unaffected, but with poor sporulation inhibition. At pH 6, post-dip brushing reduced residues to ≈ 60%; again curative control of the sensitive isolate was unaffected, but with improved sporulation inhibition. Wounded rind sections loaded higher residue levels compared to intact rind sections and large wounds loaded higher levels than small wounds (≈ 10.19, ≈ 9.06 and ≈ 7.91 μg.g-1 for large, small and no wound, respectively). Curative control of infections originating from large wounds was significantly better than those from small wounds. The ability of IMZ to control sensitive green mould infections declined from 6 and 12 h after inoculation on Clementine mandarin fruit of infections induced by small and large wounds, respectively; on navel orange fruit, curative control declined 18 and 36 h after inoculation for the respective wound size treatments. Effective IMZ concentrations that inhibit 50% (EC50) growth of nine PD and five PI isolates were determined in vitro and the IMZ sensitivity of the various isolates categorized according to their EC50 values and resistance (R) factors. Effective residue levels that predicted 50% curative (ER50C) and protective (ER50P) control of these isolates were determined in vivo. All the PI isolates had sensitive EC50 values of 0.005 - 0.050 μg.mL-1. Three PD isolates were sensitive (0.027 – 0.038 μg.mL-1), while one resistant isolate was categorized as low resistant (R-factor of 19), one as moderately resistant (R-factor of 33.2), three as resistant (R-factor of 50 - 57.6) and one as highly resistant (R-factor of 70.7). Sensitive PD isolates had mean ER50C and ER50P values on Valencia orange fruit of 0.29 and 0.20 μg.g-1, and 0.33 and 0.32 μg.g-1 on navel fruit, respectively. ER50 values for resistant isolates did not always correlate with EC50 values and ranged from 1.22 – 4.56 μg.g-1 for ER50C and 1.00 – 6.62 μg.g-1 for ER50P values. ER50P values for resistant isolates could not be obtained on navel orange fruit, but ER50C values (1.42 – 1.65 μg.g-1) were similar to those obtained on Valencia fruit. The PI isolates all behaved similar to the sensitive PD isolates with ER50C and ER50P values on navel and Valencia fruit < 0.38 μg.g-1. Alternative fungicides were assessed for the control of an IMZ sensitive, resistant and highly resistant PD isolates; these included sodium ortho-phenylpenate (SOPP), thiabendazole (TBZ), guazatine (GZT), imazalil (IMZ), pyrimethanil (PYR) and Philabuster® (PLB; a combination of IMZ and PYR), fludioxonil (FLU), azoxystrobin (AZO), Graduate®A+ (a combination of FLU and AZO) and propiconazole (PPZ). Multiple resistance was shown against IMZ, GZT, TBZ and PPZ in both resistant PD isolates. For the sensitive isolates, IMZ, SOPP, TBZ, GZT and PLB provided best curative control, while IMZ, GZT and PLB provided best protective control. For the IMZ-resistant isolates, SOPP, PYR and PLB gave the best curative control, while none of the fungicides provided adequate protective control. Globally, this is the first in-depth study of green and blue mould control with the sulphate formulation of IMZ. Findings from this study are already being implemented by industry. Solution pH is monitored, exposure time is measured and residue loading specific to application method is assessed and interpreted by means of the ER50 values. Aqueous dip applications performed best in terms of curative control, and IMZ residue loading in wound sites was most important for curative control. Other studies confirmed this and showed that IMZ is better protectively applied with wax coatings. The practical impact of IMZ resistance has been highlighted as resistant isolates infections could never be adequately controlled. IMZ alternative fungicides were assessed and SOPP, TBZ, GZT, PYR and/or PLB could be used to reduce the development and impact of IMZ resistance.
- ItemVirulence spectrum, molecular characterisation and fungicide sensitivity of the South African Rhynchosporium secalis population(Stellenbosch : Stellenbosch University, 2000-12) Robbertse, Barbara; Crous, P. W.; Lennox, C. L.; Stellenbosch University. Faculty of AgriSciences. Dept. of Plant Pathology.ENGLISH ABSTRACT: Barley leaf scald, caused by Rhynchosporium secalis, is the most important disease of barley (Hordeum vulgare) in the Western Cape province of South Africa. The disease was first reported from South Africa in 1937. The present study is the first attempt to characterise the South African R. secalis population. Topics such as pathogenesisrelated proteins, virulence spectra, variability of pathotypes, sources of variation, host resistance, breeding strategies, molecular characterisation and fungicide sensitivity are summarised in Part 1 of this dissertation. In succeeding Parts the focus is on the characteristics of the local R. secalis population regarding virulence spectrum, DNA polymorphisms, in vitro as well as in vivo fungicide sensitivity. These aspects are treated as separate entities, leading to some duplication which is unavoidable. In Part 2 the virulence spectra of 50 R. secalis isolates from a population in the. Western Cape province were determined. Twenty-one races were detected using 17 differential barley cultivars. The two most prevalent races, namely races 4 and 7 had three and four virulence genes respectively. Both race 4 and 7 were virulent on the most susceptible cultivars, namely West China, Steudelli, C.I.8618 and C.I.2226. Considering the resistance genes reported for cultivars Atlas 46, Turk, and C.I.3515 which showed no susceptible cultivar-pathogen interaction, it would appear that the Rh- Rh3-Rh4 complex is primarily involved in conferring resistance to the local R. secalis isolates. A total of 20 races (47 isolates) characterised in Part 2 were selected for further characterisation by means of DNA fingerprinting. In Part 3 an anonymous multilocus DNA probe was used to characterise the genotypic structure of these isolates by means of RFLP analysis. No correlation between any particular fingerprint pattern, race, district, field or lesion was found. The two most prevalent races, 4 and 7, did not share the same genotypes, even when isolated from the same field or lesion. The genotypic diversity of the isolates studied was 46.5% of the theoretical maximum diversity. The high level of genotypic variation observed in the South African R. secalis population resembled the genotypic diversity observed in other cereal pathogens with known sexual structures. Although no teleomorph has yet been observed, these data suggest that sexual recombination may operate within the local population of R. secalis. In South Africa barley scald is primarily controlled by means of fungicides. The continued use of fungicides on cereal crops results in the build-up of fungicide resistance in the population, which could lower the efficacy of these compounds. These aspects were investigated in Part 4, where isolates (collected during 1993 to 1995) were evaluated in vitro for sensitivity to triadimenol, tebuconazole, flusilazole and propiconazole. The sensitivity fluctuated but in 1995 isolates were significantly less sensitive towards triadimenol than in the previous two years. In a second experiment, isolates collected from two fields with a 5-6 year-history of triadimenol seed treatments and tebuconazole applications, were evaluated for their fungicide sensitivity. A significant positive correlation was observed between tebuconazole and triadimenol sensitivity among,R. secalis populations from these fields. However, such a correlation was not found within the R. secalis population collected during 1993-1995 where shorter crop rotation patterns and a range of fungicides was applied. In a third experiment, the fungicide sensitivity of local R. secalis isolates was evaluated towards two new triazole fungicides, namely bromuconazole and triticonazole. Correlation coefficients observed between these new triazoles and those previously applied in South Africa were not significantly positive. The lack of significant cross-resistance has important practical implications regarding the management of fungicide resistance. In Part 5, isolates with different minimum inhibitory concentration (MIC) towards tebuconazole in vitro (1, 3 and 10 ug/ml) were compared in vivo. The aim of this study was to determine how MIC values would influence virulence (leaf area affected) and sporulation. Results indicated that all isolates were equally fit to induce lesions and sporulate in the absence of tebuconazole. Thus no fitness cost was associated with the degree of tebuconazole sensitivity in the present study. All R. secalis isolates were able to induce lesions on tebuconazole treated leaves, but differed significantly with respect to the percentage leaf area affected. Isolates, least sensitive (MIC = 10 ug/rnl) towards tebuconazole were more adapted on tebuconazole treated leaves, being able to repeatedly cause larger lesions than sensitive R. secalis isolates (MIC = 1 ug/rnl), Sporulation was not significantly different between isolates on lesions of untreated or tebuconazole treated leaves. Larger leaf areas affected and adequate sporulation suggest that a less sensitive population would result in more disease in tebuconazole treated fields. In conclusion, this study revealed the variability associated with the South African R. secalis population regarding virulence spectrum and genotypic structure. The data in this study suggest that it is likely that the local population will easily adapt to newly introduced, single gene resistance. For more durable resistance, higher levels of quantitative resistance should be introduced. This type of resistance is, however, more difficult to identify and incorporate than single gene resistance. Consequently, barley scald control will remain dependent on the efficacy of fungicide applications. Furthermore, the lack of cross-resistance and low frequency of resistant isolates indicates a low risk for the development of fungicide resistance in the local R. secalis population. Other factors such as current crop rotation practices and the range of fungicides being ~pplied also contribute to this low risk level. However, the status of these factors can change over time. The in vivo tebuconazole sensitivity study has indicated that a resistant field population of R. secalis may be able to build-up. It is, therefore, necessary to monitor the fungicide sensitivity of R. secalis isolates at timely intervals with view to successful barley cultivation in the future.