Da Graca, John Vincent.
12 September 2014
Avocado sunblotch disease is a graft-transmissible disorder known for over 60 years and has now been recorded in at least eight countries around the world. Affected trees develop yellow, depressed streaks on young stems and fruit, marked rectangular cracking of the mature bark and a decumbent style of growth. Often a tree with symptoms produces completely symptomless shoots, termed 'recovery' growth, which are latently infected. There is a reported 95 to 100% transmission of sunblotch through the seed of such branches, and "the resultant seedlings are themselves symptomless. Indexing for sunblotch to ensure that scion and, in view of seed transmission, especially rootstock material is free of the disease is very important . The standard method used for many years has been to graft tissue onto healthy indicator seedlings and observe for symptom development for 18 months to two years. One aim of the study presented in this thesis was to develop more rapid methods for detecting the sunblotch agent. By conducting the standard indexing method in a glasshouse at controlled high temperatures of 30/28º C (day/ night) and by cutting back the indicator plants every three months, the time was reduced from two years to eight months. While this represents a considerable saving in time, the ideal must be to develop a laboratory diagnostic test that requires no more than a few days, at most, to complete. A comparative study was therefore initiated on the phenol metabolism of healthy and sunblotch-infected avocados to determine whether infection causes any major change that may reliably serve as a marker for diagnostic purposes. Significantly increased peroxidase (PO) and phenylalanine ammonia-lyase (PAL) activities, decreased indoleacetic acid (IAA) oxidase activity and higher sinapic acid levels were detected in bark tissue showing sunblotch symptoms, but not in symptomless 'recovery' growth. In contrast, increased polyphenoloxidase (PPO) activity and isoenzymes, total soluble protein levels, water soluble phenols and reduced ferulic acid levels were found in the bark of all infected trees tested, both with symptoms and symptomless. However, these latter changes have been associated with other plant-virus systems and are therefore not necessarily specific for sunblotch. Neither is any sufficiently large to be definitive as a diagnostic test. Two unidentified phenols were detected in infected, mature bark, but not in infected young bark and leaves. introduced the possibility of rapid disease detection by polyacrylamide gel electrophoresis (PAGE) of extracted RNA's as used for known viroids. In this study the presence of previously reported small molecular weight sunblotchassociated RNA's was confirmed using PAGE methods requiring two to four days to complete. This thesis presents as a further development a more rapid method of PAGE detection of RNA's enabling indexing for sunblotch to be completed in under six hours. Whilst the biochemical studies did not reveal diagnostically meaningful differences between healthy and infected avocados, there were tendencies towards differences between healthy and symptomless carrier tissues, further investigation of which may lead to a future understanding of symptom development and the symptomless condition. These include apparent higher PO and lower PAL activities in symptomless carrier tissue, as well as higher PO isoenzyme a and lower IAA oxidase isoenzyme a activities. General studies on sunblotch-infected avocados showed that fruit from symptomless 'recovery' growth branches are significantly larger and have a higher oil content than those from healthy or diseased branches, the latter finding possibly indicating a more advanced state of maturity of 'recovery' growth fruit due to earlier flowering. The avocado sunblotch agent was shown to have an in vivo thermal inactivation point of 55º C, a temperature higher than the avocado tissue can withstand thereby eliminating the possibility of thermotherapy of infected twigs. In a host range study four lauraceous plant species, Persea Schiedeana, Cinnamomum zeylanicum, C. camphora and Ocotea bullata, were successfully infected with sunblotch by grafting from infected avocado. This is the first demonstration of any host other than avocado. A phanerogametic member of the same family, Cassytha filiformis, was shown to be able to transmit the disease from avocado to avocado. No hosts from other families were found. During an electron microscope study of sunblotch-infected avocado leaf tissue, gross alterations of the chloroplasts in the yellow areas were observed. These changes included organelle swelling, loss of grana and stroma lamellae, rearrangement of remaining membranes and presence of vesicles. Also in the yellow areas paramural bodies were encountered in higher numbers and displaying altered structure than in healthy and symptomless infected leaf tissue. This study on avocado sunblotch disease was successful in both of its aims. Firstly with regard to quicker indexing techniques, the standard method using indicator plants was shortened from two years to eight months, while a rapid, six-hour test based on PAGE analysis, was developed. Secondly, more light has been shed on the biochemical and ultrastructural effects of sunblotch on its host, the avocado, as well as providing information regarding the thermal sensitivity and the host range of the agent. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 1980.
07 October 2005
During the initiation and execution of a rootstock breeding programme to overcome the financially crippling disease, Phytophthora root rot of avocado, various constraints have been identified for both the breeding as well as the screening aspect of the programme. A review of the literature revealed a complex host-pathogen interaction that should be taken into account in the recombination and screening of genetic material. With the detection of beneficial genotypes being the crux of a breeding programme, this dissertation was focused on the screening of rootstock material for tolerance to Phytophthora cinnamomi. Screening should be scientific but at the same time also be time and cost effective. Specific attention was given to (i) the correct medium for screening mass numbers of seedlings, (ii) fast and effective cloning of single selections, and (iii) evaluation of clonal material for tolerance to P. cinnamomi. Soil as a screening medium was compared with three inert hydroponic media as well as one aeroponic system. Only soil was found to be ineffective due to its properties. The other media tested, namely, sand, vermiculite, water and the aeroponic system were equal in performance. The medium to be used will depend on the preference of the breeder as each medium has its own pro's and con's. It was, however, found that the evaluation criterion to be applied depends on the medium that is used. With regard to cloning of single selections, a definite difference with regard to the cloning ability of the different selections was found. An inability to be etiolated was displayed by some of the selections and these could thus not be vegetatively propagated and were not further tested. One of the tolerance mechanisms in the standard cultivar Duke 7, is root regeneration. It was thus expected that this characteristic cloning would give an indication of the rootstock's ability to tolerate P. cinnamomi. This could not be confirmed, but most of the selections did, however, perform better than Duke 7. Comparison of feeder root percentage in non-inoculated and inoculated treatments was not sufficient for facilitating the final selection of candidate rootstocks from a large number of potential clonal selections. Four selections were made, based on the hypothesis that a larger root system will be a better forager and thus enhance the horticultural aspects of the rootstock-scion combination. Valuable information was obtained with regard to various mediums and criteria to be used during mass screening and final screening of clonal selections. This knowledge must be taken into account in the planning of future breeding projects. During this project a total of 38 984 seedlings were screened and four selections were made. For both the nursery and the producer, knowledge of the clonal ability of a potential new rootstock is important from a financial point of view. / Dissertation (MSc (Horticultural Science))--University of Pretoria, 2006. / Plant Production and Soil Science / unrestricted
Schoeman, Margareth Vuyiswa
10 October 2005
A market survey was conducted to determine the incidence of stem-end rot (SE) on avocado fruit obtained from the Pretoria Fresh Produce Market representing the Tzaneen production area. Dothiorella aromatica isolates collected from this survey were compared in terms of physiological characteristics i.e. growth and temperature, carbon and nitrogen utilization and pH response as well as genetic relatedness using random amplified polymorphic DNA (RAPD's). The incidence of SE was found to be as high as 31% and anthracnose 18%. Symptom development was more apparent when fruit was evaluated at the overripe than eating ripe stage. Of the 12 identified fungi isolated from SE lesions, D. aromatica was by far the most frequently isolated fungus. All D. aromatica isolates tested were found to be pathogenic using the fruit inoculation technique. Based on lesion size, isolates were separated into two groups of virulent and less virulent. Most isolates grouped within the one cluster, with only one isolate falling in the second group being less virulent. Although similar groupings were found between physiological tests, a lack of consistency as to which isolate belonged to which group was found. The optimum temperature for growth was 25°C and an initial pH of 6. The mean colony growth rate was 5 mm day-1. Isolates grew at a minimum of eight to a maximum of 27 mm within 24 hours. Isolates grew best on pectin and poorly on sorbitol when used as a carbon source. Urea supported growth best and poor growth was found on casein-amended sources. At a molecular level, the RAPD technique could be used successfully to seperate isolates into three groups based on cluster analyses. OPC02 was the most discrimatory primer and was therefore used in this study. Isolates produced DNA fragments ranging from 1500 bp to 450 bp. The results obtained from RAPDs could not be correlated with the pathogenicity and physiological tests. Future studies should focus on comparing isolates from different avocado production areas and testing different primers for the ability to distinguish between isolates of D. aromatica. / Dissertation (MInst Agrar (Plant Protection))--University of Pretoria, 2005. / Microbiology and Plant Pathology / unrestricted
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