The fungal disease smut (causal agent: Ustilago scitaminea H. & P. Sydow) and stalk borer Eldana saccharina Walker place major constraints on sugarcane agriculture in South Africa. The best approach for management is the introduction of resistant cultivars; however, conventional field-based screening for pest and disease resistance is a lengthy process. This study evaluated in vitro techniques combined with artificial inoculation of 12 week old in vitro plantlets and 8-10 week old embryogenic calli as rapid screening methods. Preliminary investigations were conducted on cultivars with known field ratings to smut and E. saccharina: NCo376, N26 and N39; and 5 „test‟ cultivars, whose identities were undisclosed until completion of experiments, were used to assess the accuracy of protocols. Infective U. scitaminea sporidia generated from teliospores, were used as inocula. Development of a callus protocol was unsuccessful due to sporidial and mycelial overgrowth, despite addition of a contact fungicide, Dithane M-45® (0.025 g/l) and a biocide/fungicide, PPMTM (5 ml/l), to media. Plantlet inoculation by injection, 1 cm above the apical meristem, resulted in 12% and 20% of smut susceptible NCo376 plantlets producing smut whips after 5 weeks when inoculated with 1 x 106 and 1 x 109 sporidia/ml, respectively. Smut whip production in 5 of the 8 (63%) cultivars inoculated with the lower sporidial concentration correlated with their field resistance ratings. In addition, whips harvested from in vitro plantlets were a valuable source of aseptic teliospores for future research. Ongoing work involves inoculation of NCo376 calli with such teliospores and maintenance on medium with PPMTM - emergence of whips from plantlets remains to be assessed. The E. saccharina screening protocol involved surface decontamination of eggs with 1% sodium hypochlorite (NaOCl) for 15 min. Feeding bioassays were conducted by placement of first instar larvae on in vitro plantlets and calli for 3 and 2 weeks, respectively. Larval mass, length and percentage infestation were recorded. Although greater larval size was expected in susceptible compared with resistant cultivars, the results did not support this. Significant differences in plantlet infestation were observed between susceptible (94-98%) and resistant (72-86%) lines. No significant differences were found in the callus feeding bioassay. However, a 24 h callus choice bioassay which investigated larval preference between callus genotypes compared with NCo376, showed significant differences and correctly discerned cultivar susceptibility according to field ratings. / Thesis (M.Sc.)-University of KwaZulu-Natal, Westville, 2010.

Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Grapevine virus E (GVE) is a newly identified virus that has been detected in an established vineyard in South Africa. This virus is a member of the genus Vitivirus, family Flexiviridae. Members of this genus are known to infecte grapevine and are associated with various disease complexes, such as the Rugose wood complex (RWC) and Shiraz disease (SD). However, the role and impact of GVE in South African vineyards are still unknown. It is important to study these viruses to determine how they infect and the possible impact they may have on vine health. The accurate and early detection of grapevine viruses is the first important step in disease management. In this study, reverse transcription-polymerase chain reaction (RT-PCR), double antibody sandwich enzyme linked immunesorbent assay (DAS-ELISA) and quantitative (q)RT-PCR were used for the detection of GVE in the vineyard (Vitis vinifera cv Merlot) where GVE was first identified in South Africa. Reverse transcription-PCR was used for detection and determining the incidence of GVE. The incidence was as low as 3% in the vineyard surveyed. All the GVE positive plants were co-infected with GLRaV-3 and no disease association could therefore be made. Evaluation of the Bioreba Grapevine virus A (GVA) DAS-ELISA kit showed that it did not detect GVE. No cross-reactivity occurred with epitopes of GVE, confirming this kit to be a valid and specific assay for GVA infection. The relative virus titer of GVE was calculated over the growing season of 2010/2011, using qRT-PCR. No fluctuation in virus titer was observed during that growing season. Transmission experiments were performed in an attempt to transfer GVE from grapevine to an alternative host. Three different transmission buffers as well as nine different herbaceous plant species, that have shown to be susceptible to several plant viruses in previous studies, were evaluated. In these experiments, GVE could not be transmitted to any of the herbaceous species. To further characterize GVE, chimeric clones were constructed with GVA. The ORF2 and ORF5 of GVE were cloned into previously constructed GVA ORF2 and ORF5 deletion mutants. Construction of the chimeric clones, 35S-GVA-GR5-ΔORF2-GVE-ORF2 and 35S-GVA-118-ΔORF5-GVE-ORF5 were successful and they were evaluated for their infectivity in N. benthamiana. The 35S-GVA-GR5-ΔORF2-GVE-ORF2 chimera was able to infect and replicate in these plants and disease symptoms such as yellowing of veins and leaf curling were observed. Virus, derived from this vector, was detected by TPIA, RT-PCR and DAS-ELISA. The 35S-GVA-118-ΔORF5-GVE-ORF5 chimeric vector was not able to infect N. benthamiana as no disease symptoms were observed in any of the infiltrated plants and virus was not detected with serological analysis and RT-PCR. This study was aimed at further characterizing the recently identified virus GVE. Here, insight is given into the prevalence of this virus in the vineyard where it was first identified and attempts to biologically characterize GVE were made. / AFRIKAANSE OPSOMMING: Grapevine virus E (GVE) is „n nuut geïndetifiseerde virus wat onlangs in „n gevestigde wingerd in Suid Afrika opgespoor is. Hierdie virus vorm deel van die genus Vitivirus, familie Betaflexiviridae. Spesies in hierdie genus is bekend vir wingerdinfeksies en word met „n verskeidenheid wingerd siektes geassosieer, soos bv. Rugose wood complex (RWC) en Shiraz siekte (SD). Die rol en impak van GVE is nog onbekend. Dit is dus belangrik om die virus te bestudeer om te bepaal hoe dit infekteer en of dit enige impak het op wingerd gesondheid. Akkurate en vroeë opsporing van virusse is die eerste belangrike stap vir virussiekte beheer. In hierdie studie word tru-transkripsie (TT) – polimerase ketting reaksie (PKR), dubbel teenliggaam (DAS) -ensiem gekoppelde immuno-absorberende analise (ELISA) en qTT-PKR gebruik vir die opsporing van GVE in die wingerd (Vitis vinifera cv Merlot) waar dit vroeër in Suid Afrika geïdentifiseer was. Vir opsporing en bepaling van verspreiding is TT-PKR gebruik. Daar is bepaal dat 3% van die wingerd met GVE geïnfekteer is. Al die GVE-positiewe stokke het ook positief getoets vir GLRaV-3 en geen assosiasie met siekte simptome kon gemaak word nie. Evaluering van die Bioreba GVA DAS-ELISA met GVE positiewe stokke het nie GVE opgespoor nie. Geen kruisreaktiwiteit het plaasgevind met epitope van GVE nie en dus is die DAS-ELISA ʼn betroubare toets vir GVA infeksie. Die relatiewe virus titer van GVE was ook bepaal oor die groeiseisoen van 2010/2011 deur qTT-PKR te gebruik. Geen fluktuasie in virus titer gedurende die groeiseisoen is waargeneem nie. Transmissie eksperimente is gedoen om GVE vanaf wingerd na ʼn alternatiewe gasheer oor te dra. Drie verskillende transmissie buffers en tien verskillende sagteplant spesies, wat voorheen vatbaarheid vir plantvirusse getoon het, is gebruik. In die transmissie eksperimente kon GVE nie na enige van die sagteplante oorgedra word nie. Om GVE verder te karakteriseer is hibried-virusse met GVA gemaak. Die leesraam (ORF) 2 en ORF5 van GVE gekloneer in GVA ORF2 en -ORF5 delesie konstrukte, 35S-GVA-GR5-ΔORF2 en 35S-GVA-118-ΔORF5, onderskeidelik (Blignaut, 2009; Du Preez, 2010). Klonering van die hibried konstrukte, 35S-GVA-GR5-ΔORF2-GVE-ORF2 en 35S-GVA-118-ΔORF5-GVE-ORF5, was suksesvol en is in N. benthamiana geëvalueer. Virus afkomstig van die 35S-GVA-GR5-ΔORF2-GVE-ORF2 hibried konstruk, kon plante suksesvol infekteer en kon repliseer binne hierdie plante. Siektesimptome soos vergeling van die are en rolblaar is ook waargeneem in plante geïnfekteer met hierdie hibried konstruk. Plante is getoets met weefsel afdruk immuno analise (TPIA), TT-PKR en DAS-ELISA en is positief gevind vir virus afkomstig van hierdie konstruk. Die 35S-GVA-118-ΔORF5-GVE-ORF5 hibried kon nie N. benthamiana infekteer nie en geen siektesimptome is waargeneem in enige van die plante geïnfiltreer met hierdie konstruk. Serologiese analise en TT-PKR het ook nie virus in die N. benthamiana plante opgespoor nie. Die doel van hierdie studie was om GVE te karakteriseer. In hierdie studie word insig gegee oor die verspreiding van hierdie virus in Suid Afrika en pogings is gemaak om GVE biologies te karakteriseer.

Thesis (MSc)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: South Africa is ranked eighth in the world as far as international wine production is concerned and in terms of area under bearing vines South Africa is ranked 12th. In 2011 the wine industry contributed R4 204.4 million to the South African economy in state revenue from wine products. The importance of viticulture to the economy of South Africa forces the industry to limit the effect of all disease causing pathogens in order to keep their competitive edge. Aster yellows (AY) phytoplasma 16SrI-B subgroup was reported for the first time in grapevine (Vitis vinifera L. (Vitaceae)) in South Africa in 2006. Worldwide phytoplasma diseases of grapevine cause serious damage ranging from lower yields to the death of vines. The lack of knowledge about the epidemiology of AY disease makes it difficult to determine the impact of the disease on the South African wine industry. The aim of this study was to conduct surveys in disease-affected vineyards in the Vredendal region to determine the incidence and spatial distribution of the disease in a variety of cultivars. The field surveys based on visual symptoms of AY disease were confirmed by polymerase chain reaction (PCR). A survey was also conducted in and around AY-infected vineyards in search of possible alternative host plants of the phytoplasma. Spatial distribution of AY-affected vines were analysed using the PATCHY spatial analysis package. A rapid decline of AY-affected Chardonnay eventually leading to the death of vines was observed, confirming the sensitivity of Chardonnay towards grapevine yellows infections. Symptomless AY infections occurred and AY could not be detected in all symptomatic vines, which indicate uneven distribution of AY in individual vines. Molecular analyses using PCR-RFLP showed that all vines sampled in the Vredendal vicinity contained AY phytoplasma only. No phytoplasmas were present in any weeds or other possible host plants tested. Although the mean yearly disease incidences of Chardonnay (29.95%) and Chenin blanc (16.64%) were higher than Pinotage (5.80%) over the four-year survey period, there was no statistically significant difference between the disease incidences of these three cultivars. The mean yearly disease incidence showed a trend over time and the disease incidence of the first year was significantly lower than that of the other years. Chardonnay showed a cumulative disease incidence of 37.77% at the end of the 4-year study which means that Chardonnay vineyards can be 100% AY infected in ten years’ time. Spatial distribution patterns of AY-infected vines were mostly non-random with clustering of disease affected vines along and across vine rows. With the exception of one vineyard, aggregation of AY-affected vines mostly occurred on the edge of vineyards adjacent to infected vineyards. This epidemiological study gives an indication of the sensitivity of the different cultivars towards AY, the tempo of spreading and the future impact of the disease on the South African wine industry. It also contributes valuable information towards the development of a management strategy for grapevine yellows disease in South African vineyards. / AFRIKAANSE OPSOMMING: Suid- Afrika is op agtste op die wêreld ranglys wat internasionale produksie van wyn aan betref, en in terme van oppervlakte onder wingerd, is Suid-Afrika 12de. In 2011 het die wynbedryf R4 204.4 miljoen tot die Suid-Afrikaanse ekonomie bygedra in staats inkomste uit wyn produkte. Die belangrikheid van wingerd tot die ekonomie van Suid-Afrika dwing die bedryf om die effek van alle siekteveroorsakende patogene te beperk, om sodoende hul kompeterende voordeel te behou. Aster vergeling (AY) fitoplasma 16SrI-B subgroep is vir die eerste keer in 2006 in wingerd (Vitis vinifera L. (Vitaceae)) in Suid-Afrika waargeneem. Fitoplasma siektes van wingerd veroorsaak wêreldwyd ernstige skade wat wissel van laer opbrengste tot die afsterf van wingerdstokke. Die gebrek aan kennis oor die epidemiologie van astervergeling siekte maak dit moeilik om die impak van die siekte op die Suid-Afrikaanse wynbedryf te bepaal. Die doel van hierdie studie was om ‘n opname te maak in siekte geaffekteerde wingerde in die Vredendal omgewing om sodoende siekte voorkoms en verspreidingspatrone van die siekte in 'n verskeidenheid van kultivars te bepaal. Die veld opnames, gebaseer op visuele simptome van aster vergeling siekte, was bevestig deur polimerase kettingreaksie (PKR). ‘n Opname is ook in en om aster vergeling geaffekteerde wingerde uitgevoer, op soek na moontlike alternatiewe gasheer plante van die fitoplasma. Verspreidingspatrone van astervergeling geaffekteerde wingerde is ontleed met behulp van die PATCHY ruimtelike analise pakket. 'n Vinnige agteruitgang van AY geaffekteerde Chardonnay, wat uiteindelik gelei het tot die afsterf van wingerde, is waargeneem, wat die sensitiwiteit van Chardonnay teenoor wingerdvergeling infeksie bevestig. Simptoomlose astervergeling fitoplasma infeksies kom voor en astervergeling fitoplasma kon nie opgespoor word in alle simptomatiese wingerdstokke nie, wat op oneweredige verspreiding van AY fitoplasma in individuele wingerdstokke dui. Molekulêre ontledings met behulp van PKR-RFLP het getoon dat alle wingerdstokke, wat in die Vredendal omgewing getoets is, slegs astervergeling fitoplasma bevat. Geen fitoplasmas was teenwoordig in enige onkruide of ander moontlike gasheer plante. Hoewel die gemiddelde jaarlikse siekte voorkoms van Chardonnay (29,95%) en Chenin Blanc (16,64%) oor die vier-jaar opname periode hoër was as dié van Pinotage (5,80%), was daar geen statisties beduidende verskil tussen die siekte voorkoms van hierdie drie kultivars nie. Die gemiddelde jaarlikse siekte voorkoms het 'n tendens oor tyd getoon, en die siekte voorkoms van die eerste jaar was betekenisvol laer as dié van die ander jare. Chardonnay het ‘n kumulatiewe siekte voorkoms van 37.77% aan die einde van die 4-jaar studie getoon, wat beteken dat Chardonnay wingerde binne 10 jaar 100% besmet kan wees met AY. Verspreidingspatrone van AY geaffekteerde wingerdstokke was meestal nie-ewekansig met bondeling van geaffekteerde wingerdstokke in en oor wingerd rye. Bondeling van AY geaffekteerde wingerdstokke het, met die uitsondering van een wingerd, meestal op die kant van wingerde aanliggend aan besmette wingerde, voorgekom. Die epidemiologiese studie gee 'n aanduiding van die sensitiwiteit van die verskillende kultivars ten opsigte van AY, die tempo van die verspreiding en die toekomstige impak van die siekte op die Suid-Afrikaanse wynbedryf. Dit dra ook waardevolle inligting by tot die ontwikkeling van 'n strategie vir die bestuur van wingerdvergeling siekte in Suid-Afrikaanse wingerde.

Cereal crop productivity is hampered when these plants are infested by phloem feeding aphids. A great deal of research has been carried out with the direct aim of a clearer understanding of the mechanism involved in the interaction between aphids and their host plants. Research has directly or indirectly been geared towards enhanced plant productivity and achieving sustainable agriculture. Barley (Hordeum vulgare L.) is an important small grain crop in South Africa, whose crop performance is negatively affected by fluctuations in weather patterns as well as by agricultural pests. One of the insect pests infesting barley is the Russian wheat aphid, Diuraphis noxia Kurdjumov (RWA), of which the two South African biotypes, codenamed RWASA1 and RWASA2, were studied in this thesis. During dry spells, RWA infestation becomes a more serious threat to barley productivity. Resistant plants have been used to combat RWA infestation of small grains. In South Africa, 27 RWA-resistant wheat cultivars are currently used in commercial cultivation, but no resistant barley lines have, unfortunately, been developed, in spite of this grain’s significant economic importance. This informed the study in this thesis, and this interest particularly focussed on three RWA-resistant lines developed by the USDA, testing their performance against South African RWA biotypes, for possible adaptation to South Africa. The aim was thus to examine the plant-aphid interactions, aphid breeding rates, plant damage and sustainability, evidence of resistance or tolerance and finally potential performance under elevated CO2 (a very real climate change threat). Two major avenues of research were undertaken. The first aspect involved examination of structural and functional damage caused by RWASA1 and RWASA2 on the three resistant and a non-resistant line. Aphid population growth and damage symptoms (chlorosis and leaf roll) of infestation by these aphid biotypes were evaluated. This was followed by a structural and functional approach in which the effects of feeding on the transport systems (phloem and xylem) of barley were investigated. Fluorescence microscopy techniques (using aniline blue fluorochrome, a specific stain for callose and 5,6-CFDA, a phloem-mobile probe) were applied to investigate the feeding-related damage caused by the aphids, through an examination of wound callose formation and related to this, the resultant reduction in phloem transport capacity. Transmission electron microscopy (TEM) techniques provided evidence of the extent of the feeding-related cell damage. The second aspect involved a study of the effect of changing CO2 concentrations ([CO2]) on the resistant and susceptible barley cultivars to feeding by the two RWA biotypes. Leaves of plants grown at ambient and two elevated levels of [CO2] were analysed to investigate the effect of changing [CO2] on biomass, leaf nitrogen content and C:N ratio of control (uninfested) and infested plants. The population growth studies showed that the populations of the two RWA biotypes as well as bird cherry-oat aphid (BCA, Rhopalosiphum padi L.) increased substantially on the four barley lines. BCA was included here, as it had been the subject of several previous studies. RWASA2 bred faster than RWASA1 on all lines. The breeding rates of the two RWA biotypes were both suppressed and at near-equivalent levels on the three resistant lines, compared to the non-resistant PUMA. This suggests that the resistant lines possessed an antibiosis resistance mechanism against the feeding aphids. Feeding by the aphids manifested in morphological damage symptoms of chlorosis and leaf roll. The two biotypes inflicted severe chlorosis and leaf roll on the non-resistant PUMA. In the resistant plants, leaf rolling was more severe because of RWASA2 feeding compared to RWASA1 feeding. In contrast, chlorosis symptoms were more severe during RWASA1 feeding than was the case with RWASA2 feeding. Investigation of the effect of aphid feeding on the plants showed that callose was deposited within 24h and that this increased with longer feeding exposure. Wound callose distribution is more extensive in the non-resistant PUMA than in the resistant plants. RWASA2 feeding on the resistant lines caused deposition of more callose than was evident with RWASA1 feeding. During long-term feeding, it was evident that variation in the intensity and amount of wound callose was visible in the longitudinal and transverse veins of the resistant plants. Of the three STARS plants, STARS-9301B had the least callose. Interestingly, wound callose occurred in both resistant and non-resistant plants, in sharp contrast to what has been reported on resistant wheat cultivars that were developed in South Africa. The relative reduction in the wound callose deposited in the resistant line, when compared to the non-resistant lines, suggests the presence of a mechanism in the resistant lines, which may prevent excessive callose formation. Alternatively, the mechanism may stimulate callose breakdown. RWASA2 feeding on the resistant lines deposited more wound callose than RWASA1 feeding. This evidence supports the hypothesis that RWASA2 is a resistance breaking and more aggressive feeder than RWASA1 is; and further underscores the urgent need for development of RWA-resistant barley cultivars. The ultrastructural investigation of the feeding damage showed that the two biotypes caused extensive vascular damage in non-resistant plants. There was extensive and severe cell disruption and often obliteration of cell structure of the vascular parenchyma, xylem and phloem elements. In sharp contrast, among the resistant plants, feeding-related cell damage appeared to be substantially reduced compared to the non-resistant PUMA. Low frequency of damaged cells indicated that majority of the cell components of the vascular tissues were intact and presumed functional. There was evidence of salivary material lining the secondary walls of the vascular tissue, which resulted in severe damage. Within xylem vessels, saliva material impregnated half-bordered pit pairs between the vessels and adjacent xylem parenchyma. This is believed to prevent solute exchange through this interface, thereby inducing leaf stress and vi leaf roll. A notable finding is that RWASA2 effectively induced more cell damage to vascular tissues in the resistant lines than did RWASA1. In general the experimental evidence (see Chapter 5) suggests that the resistant lines are possibly more tolerant (or able to cope with) to RWA feeding. Evidence for this is the reduction of wound callose and at the TEM level, a comparatively less obvious cell damage in the resistant lines, which suggests that they possess antibiosis and tolerance capacity. The apparent reduction of feeding-related cell damage from the TEM study confirmed the disruptive action of the feeding aphids in experiments using the phloem-mobile probe, 5,6-CF. Results showed that feeding by RWASA1 and RWASA2 reducedthe transport functionality of the phloem in all cases, but that RWASA2 feeding caused a more obvious reduction in the rate and distance that 5,6-carboxyfluorescein was transported, than did RWASA1. Investigation of the effect of changing [CO2] on the barley cultivars showed that in the absence of aphids and under elevated CO2 conditions, the plants grew more vigorously. In this series of experiments, the infested plants suffered significant reduction in biomass under ambient (as was expected) and under the two elevated CO2 regimes. Biomass loss was greater at elevated CO2 than under ambient [CO2]. The infested nonresistant PUMA plants showed a more significant biomass loss than did the resistant cultivars. Clearly, the benefits derived from elevated CO2 enrichment was thus redirected to the now-advantaged aphids. Uninfested vii plants showed an increase in leaf nitrogen under the experimental conditions. However, feeding aphids depleted leaf nitrogen content and this was more apparent on plants exposed to RWASA2 than was the case with RWASA1. The end result of this was that C:N ratio of infested plants were higher than uninfested plants. Clearly, the faster breeding rates of the aphids at elevated CO2 caused depletion of N and a resultant deficiency exacerbated chlorosis as well as leaf rolling due to the higher aphid population density under elevated CO2 than at ambient. By 28 days after infestation (DAI), majority of the plants exposed to enriched CO2 treatments had died. A major finding here was thus that although this study demonstrated that elevated CO2 resulted in an increase in biomass, this was detrimentally offset in plants infested by the aphids, with a decline in biomass and loss of functionality leading to plant death at 28DAI. The overriding conclusion from this study is a clear signal that the twin effects of CO2 enrichment (a feature of current climate change) and aphid infestations may precipitate potential grain shortages. A disastrous food security threat looms.

Ph.D. / A number of aspects which might influence the buildup of spider mites on cotton were investigated. In contrast to their numbers on cotton treated with some of the insecticides used to control Heliothis arrniaer (Hübner) in field trials, the numbers of spider mites remained very low in the unsprayed cotton. Predators such as especially the pirate bug, Onus thniroborus (Hesse) appear to play an important role in maintaining mites in untreated plots at low population levels. Predator numbers were significantly reduced by some of the insecticides used. It is, however, not yet clear how the predator complex as a whole affects mite numbers ...

Please read the abstract in the section 00front of this document / Dissertation (Magister Istitutiones Agrariae)--University of Pretoria, 2006. / Plant Production and Soil Science / unrestricted

Albugo tragopogonis is responsible for white rust of sunflower. It was first observed in 1929 in South Africa. Recently however, white rust has resulted in lodging exceeding 80% in some sunflower growing areas. Due to the obligate nature of the pathogen, studies of the biology, epidemiology and control of the disease has until now been limited to field trials and observations. Greenhouse trials are needed to understand the infection process, and to examine any resistance mechanisms used by the plant to defend itself against the pathogen. Presently, there is no practical artificial inoculation technique available and effective storage of the fungus is difficult. The purpose of these studies was to find new storage and inoculation techniques. Once the inoculation technique was optimized, the infection process of A. tragopogonis on susceptible and tolerant sunflower genotypes was examined. Infected leaves were collected from sunflower seedlings at the Grain-Crops Institute in Potchefstroom. Infected leaves were covered with plastic bags and freshly cut stems were placed in a cooler box filled with ice water. Some of the infected leaves were also placed in paper bags and allowed to dry for 24 h. Sporangia were collected using a vacuum device and stored in gelatin capsules at -20°C, -70°C or in liquid nitrogen directly after collection or following desiccation for 24 h. Sunflower seedlings at the four-leaf-stage were inoculated with freshly collected sporangia, or sporangia stored for 3, 5, 9, 12 and 15 mo. A zoospore suspension was prepared by allowing 105 sporangia/ml to germinate in distilled water for 3 h at 10°C. The zoospore suspension was then sprayed onto leaves until they were completely wet with a hand held garden spray bottle. Inoculated seedlings were covered with plastic bags to maintain high humidity and placed at 12°C for 16 h and incubated in a greenhouse until symptom development. Infection levels were assessed 10¬14 d after inoculation, using a scale of 1-5, with 1 indicating resistance and 5 indicating severe infection. Infection with fresh sporangia proved to be very consistent. Sporangia stored in capsules immediately after collection at -70°C after desiccation, produced the highest infection. Low levels of infection resulted from storage in liquid nitrogen or directly at -70°C. It is evident that successful storage may be obtained if the sporangia are dried before storage. These techniques to store and inoculate A. tragopogonis have proven to be reliable. Susceptible and tolerant genotypes were inoculated, using the spray bottle inoculation technique described above, to examine the difference in infection of A. tragopogonis. Leaves used for light microscopy were cut into 20 mm2 and those for scanning electron microscopy were cut into 5x5 mm pieces at 2, 4, 6, 8, 10, 12, 24, 36, 48, 72, 96, 120, 144 and 168 h time intervals after inoculation. The epidermis, palisade parenchyma and spongy parenchyma were chronologically stripped using the double-sided tape method. The material for the light microscope was prepared using the whole-leaf clearing and staining technique, the lactophenol-ethanol-analine blue technique and sectioning with freeze microtome. The material for SEM was prepared according to standard procedures and examined with a JEOL 840 SEM at 5 kV. Both the whole-leaf clearing and staining and the lactophenol-ethanol-aniline blue techniques proved to be unsuitable as most of the tissue was damaged by boiling. Sectioning with the freeze microtome was also unsuccessful. The SEM gave the most transparent results. This method gave us the ability to compare results with previous literature and to compare the infection process between of A. tragopogonis in the susceptible (RHA 358) and the tolerant (HYS 33) genotype. / Dissertation (MSc ( Plant Pathology))--University of Pretoria, 2005. / Microbiology and Plant Pathology / unrestricted

In South Africa the genus, Eucalyptus plays an important role as a major economic component in the forest and mining sector. Unfortunately, this genus has problems due to damage by pests. The Eucalyptus snout beetle, Gonipterus scutellatus, feeds and defoliates the leaves of eucalypts. Plants produce secondary metabolites, which protects them against defoliation by insects and herbivores. This study focuses on the variations of total soluble phenolic content of nine Eucalyptus species between the species and within the species throughout the year. Total soluble phenols were quantified with the Folin-Ciocalteau reagent. There was a general increase in the concentration of total soluble phenols throughout the year. The content of total soluble phenols were generally higher compared to carbohydrates. This experiment needs to be done on a continuous basis so as to formulate a screening method for eucalypt species that are resistant to G. scutellatus. / Dissertation (M Inst Agrar (Sustainable Insect Management))--University of Pretoria, 2005. / Zoology and Entomology / unrestricted

In South Africa, the genus Eucalyptus plays an important role as a plantation tree and hence forms a major economic component in the forest sector. An insect pest of these Eucalyptus species, Gonipterus scutellatus, causes periodic defoliation in the plantations. Plants have extraordinary array of chemicals (secondary metabolites), which defend them from herbivores. This study reports on the seasonal variation of the tannin concentration and carbohydrate content of the leaves of nine Eucalyptus species. Soluble tannins were quantified using Hagerman test and the carbohydrate content with a refractometer. Generally high tannin concentrations were observed during spring and late summer with low concentrations during autumn and winter. During the months of February, April, July tannin concentration was usually found to be higher than the carbohydrate content. The higher concentration of tannin could be linked to the allocation of carbon for the production of tannin instead of carbohydrates for growth. / Dissertation (MInst Agrar)--University of Pretoria, 2006. / Zoology and Entomology / unrestricted

Maize contributes substantially to food security in the Eastern Cape province of South Africa. It is a staple food to many of the province’s rural and urban inhabitants. Insect pests are one of the factors that hamper its productivity and as a result, deprive farmers of good yields. The adverse effects of insecticides and the high cost associated with them and the cost of transgenic seeds are some of the challenges faced by small-scale farmers in rural areas. Alternative control methods which include the use of indigenous techniques to control pests are now sought. A study to identify and document ethnobiological means used by rural farmers to manage insect pests of maize was conducted in the rural areas of the Eastern Cape Province. A total of 217 participants were interviewed on the matter, using semi-structured but detailed questionnaires. Rural farmers due to their linkage to agriculture activities and the fact that they are considered as custodians of agricultural indigenous knowledge were selected as respondents. Only maize producing and IsiXhosa speaking people were chosen to contribute. Main focus was on the demography of respondents, crop production activities and insect pest control. Pretesting of the questionnaire in order to assess the appropriateness of questions and comprehension by both farmers and enumerators was done. Data was analysed using descriptive statistics. Fifty five percent (55 %) of the respondents were females and the highest number of participants was from the Chris Hani District Municipality. Majority of the people were unemployed or pensioners. Most had only attended primary school and the mean age was 59 years. Apart from maize, respondents were cultivating other crops such as cabbage, Swiss chard, potatoes etc. Stalk borers followed by cutworms were the main pests of maize in these areas. Respondents used mainly insecticides, followed by alternative substances, which also included cultural control methods such as planting date manipulation. Few respondents used plants as control agents for insect pests. Some people did not control pests even though they were a problem in their fields. The most used plant was Chenopodium ambrosiodes L, while the most used substance was Madubula (a detergent). The most used insecticide was carbaryl from the carbamite family. Respondents listed different preparation techniques for all the control methods mentioned. These techniques revealed different times of preparation, quantities of ingredients, amounts applied on plants, modes of application and intervals of application. Rural farmers in the study areas used different atypical methods which may play a significant role in pest management today. Some of the products may have a positive influence on agriculture, while some are dangerous to humans and environmental health. Further research which will investigate their potential use in pest control needs to be done.