No description available.

No description available.

No description available.

Thesis (MScAgric (Plant Pathology))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT: A survey was undertaken on apple and pear trees in the Western Cape Province to determine the aetiology of trunk diseases with reference to trunk diseases occurring on grapevine. Grapevine trunk diseases cause the gradual decline and dieback of vines resulting in a decrease in the vine’s capability to carry and ripen fruit. In recent years, viticulture has been expanding into several of the well established pome fruit growing areas. The presence of trunk pathogens in pome fruit orchards may affect the health of the pome fruit trees as well as cause a threat to young vineyards planted in close proximity to these potential sources of viable inoculum. Several genera containing species known to be involved in trunk disease on pome fruit and grapevine were found, including Diplodia, Neofusicoccum, Eutypa, Phaeoacremonium and Phomopsis. Diplodia seriata and D. pyricolum, were isolated along with N. australe and N. vitifusiforme. Four Phaeoacremonium species, P. aleophilum, P. iranianum, P. mortoniae and P. viticola, two Phomopsis species linked to clades identified in former studies as Phomopsis sp. 1 and Phomopsis sp. 7, and Eutypa lata were found. In addition, Paraconiothyrium brasiliense and Pa. variabile, and an unidentified Pyrenochaetalike species were found. Of these the Phaeoacremonium species have not been found on pear wood and it is a first report of P. aleophilum occurring on apple. This is also a first report of the Phomopsis species and Eutypa lata found occurring on pome trees in South Africa Two new coelomycetous fungi were also found including a Diplodia species, Diplodia pyricolum sp. nov., and a new genus, Pyrenochaetoides gen. nov. with the type species, Pyrenochaetoides mali sp. nov., were described from necrotic pear and apple wood. The combined ITS and EF1-α phylogeny supported the new Diplodia species, which is closely related to D. mutila and D. africana. The new species is characterised by conidia that become pigmented and 1-septate within the pycnidium, and that are intermediate in size between the latter two Diplodia species. Phylogenetic inference of the SSU of the unknown coelomycete provided bootstrap support (100%) for a monophyletic clade unrelated to known genera, and basal to Phoma and its relatives. Morphologically the new genus is characterised by pycnidial with elongated necks that lack setae, cylindrical conidiophores that are seldomly branched at the base, and Phoma-like conidia. The phylogenetic results combined with its dissimilarity from genera allied to Phoma, lead to the conclusion that this species represents a new genus. A pathogenicity trial was undertaken to examine the role of these species on apple, pear and grapevine shoots. N. australe caused the longest lesions on grapevine shoots, while Pyrenochaetoides mali, Pa. variabile, D. seriata and P. mortoniae caused lesions that were significantly longer than the control inoculations. On pears, D. pyricolum and N. australe caused the longest lesions, followed by D. seriata and E. lata. On apples, the longest lesions were caused by N. australe and P. iranianum. D. seriata, D. pyricolum, E. lata, N. vitifusiforme, Pa. brasiliense, P. aleophilum and P. mortoniae also caused lesions on apple that were significantly longer than the control. The study demonstrated that close cultivation of grapevine to apple and pear orchards may have inherent risks in terms of the free availability of viable inoculum of trunk disease pathogens. / No Afrikaans abstract available.

Thesis (MSc)--University of Stellenbosch, 2005. / ENGLISH ABSTRACT: In the field of agriculture, plant pathogens are a major concern because of the severe damage these organisms cause to crops yearly. Fundamental studies regarding plant pathogens and their modes of action made it possible for researchers in the field of molecular biology to investigate pathogens further on a molecular level. Botrytis cinerea, has been used to great effect as a model system to investigate various aspects regarding pathogenesis, also on a molecular level. Molecular research done on B. cinerea over the last few years has shown that the endopolygalacturonases (EPGs) of this fungus are key role players in pathogenesis. This hydrolytic enzyme family of six members, encoded by the Bcpg1-6 genes, are important in breaking down the complex cell wall polymers of host plants, enabling the fungus to penetrate its host sufficiently. It has been shown that both BcPG1 and 2 are crucial for virulence of B. cinerea. A leucine-rich repeat inhibitor protein situated in the cell wall of various plant species, the polygalacturonase-inhibiting protein (PGIP), has been proven to interact with and inhibit EPGs, and thus the necrotic actions of B. cinerea. From literature it was clear that specific data regarding individual interactions of fungal EPGs with PGIPs are lacking currently. Furthermore, most experiments regarding the effects of EPG as well as interaction and inhibition studies of EPGs and PGIPs, rely on in vitro methods, without the possibility to contextualize the results on an in vivo or in planta level. The scope of this study was to specifically address the issues of individual EPG:PGIP interactions and the use of possible in vivo methodology by using EPGs from a highly virulent South African strain of B. cinerea and the grapevine VvPGIP1 that has been previously isolated in our laboratory. This PGIP, originally isolated from Vitis vinifera cv Pinotage, has been shown to inhibit a crude EPG extract from this strain with great efficiency. The approach taken relied on heterologous over-expression of the individual Bcpg genes and the isolation of pure and active enzymes to evaluate the inhibition of the EPGs with VvPGIP1. The genes were all successfully over-expressed in Saccharomyces cerevisiae with a strong and inducible promoter, but active enzyme preparations have been obtained only for the encoding Bcpg2 gene, as measured with an agarose diffusion assay. The in vitro PGIP inhibition assay is also based on the agarose diffusion assay and relies on activity of the EPGs to visualize the inhibiting effect of the PGIP being tested. The active EPG2, however, was not inhibited by VvPGIP1 when tested with this assay. The EPG encoding genes from B. cinerea were transiently over-expressed also in Nicotiana benthamiana by using the Agrobacterium-infiltration technique. Transgene expression was confirmed by Northern blot analysis and EPG-related symptoms were observed five to eight days post-infiltration. Differential symptoms appeared with the various EPGs, providing some evidence that the symptoms were not random events due to the infiltration or a hypersensitive response. Moreover, the symptoms observed for EPG2 was similar to those that were reported recently by another group on the same host. In spite of the expression data and the clear symptoms that developed, active preparations, as measured with the agarose diffusion plate asay, could only be obtained for EPG2 again. In our search for a possible in vivo method to detect and quantify EPG activity and inhibition by PGIPs, we tested and evaluated a technique based on chlorophyll fluorescence to detect the effect of EPGs on the rate of photosynthesis. Our results showed that the over-expression of these genes reduced the rate of electrons flowing through photosystem II, indicating metabolic stress occurring in the plant. We used the same technique to evaluate possible interaction between VvPGIP1 respectively with BcPG1 and 2 and found that the co-expressing of the Vvpgip1 gene caused protection of the infiltrated tissue, indicating inhibition of EPG1 and 2 by VvPGIP1. For EPG2, the observed interaction and possible inhibition by VvPGIP1 is the first report to our knowledge of an interaction between this specific EPG2 and a PGIP. Moreover, to further elucidate the in planta interaction between VvPGIP1 and the EPGs from the South African B. cinerea strain, we tested for possible interactions by making use of a plant two-hybrid fusion assay, but the results are inconclusive at this stage. Previous studies in our laboratory have shown that several natural mutations exist between PGIP encoding genes from different V. vinifera cultivars. Based on this finding and the fact that these natural mutations could result in changes with regard to EPG inhibition and ultimately disease susceptibility, we isolated an additional 37 PGIP encoding genes from various grapevine genotypes, some of which are known for their resistance to pathogens. Combined, these results make a valuable contribution to understand plant pathogen interactions, specifically in this case by modeling the interactions of pathogen and plant derived proteins. The possibility to use in vivo methods such as chlorophyll fluorescence to follow these interactions on an in planta level, provides exciting possibilities to strenghten and contextualize in vitro results. / AFRIKAANSE OPSOMMING: Plantpatogene organismes veroorsaak jaarliks erge skade aan landbougewasse en word dus as ’n ernstige probleem in die landbousektor beskou. Diepgaande studies wat handel oor plantpatogene en hul metodes van infeksie het dit vir molekulêre bioloë moontlik gemaak om patogene nou ook op molekulêre vlak verder te bestudeer. Botrytis cinerea is baie effektief as modelsisteem gebruik om verskeie aspekte van patogenese verder te bestudeer, ook op ‘n molekulêre vlak. Molekulêre navorsing op B. cinerea, het getoon dat die endopoligalakturonases (EPGs) van dié swam kernrolbelangrik in patogenese is. Hierdie sesledige hidrolitiese ensiemfamilie word gekodeer deur die Bcpg1-6 gene en is belangrik vir die afbraak van die komplekse selwandpolimere van plantgashere, om suksesvolle gasheerpenetrasie te veroorsaak. Daar is aangetoon dat beide BcPG1 en 2 essensieël vir virulensie van die patogeen is. ’n Leusienryke-herhalings inhibitorproteïen wat in die selwand van verskeie plantspesies voorkom, die poligalakturonase-inhiberende proteïen (PGIP), het interaksie met en inhibeer EPGs en gevolglik ook die nekrotiserende aksies van B. cinerea. Uit die literatuur is dit duidelik dat spesifieke inligting aangaande individuele interaksies van fungiese EPGs met PGIPs tans nog ontbreek. Verder word daar op in vitro metodologie staatgemaak wannneer die effekte van EPGs asook die interaksie en inhibisie met PGIPs bestudeer word, sonder om die konteks van die in vivo- of in planta-omgewing in ag te neem. Die fokus van hierdie studie was om aspekte van individuele EPG:PGIP interaksies, asook die moontlike gebruik van in vivo metodologie te bestudeer deur EPGs, afkomstig van ’n hoogs virulente Suid-Afrikaanse ras van B. cinerea en die wingerd VvPGIP1, wat vroeër in ons laboratorium geïsoleer is, te gebrruik. Hierdie PGIP wat uit Vitis vinifera cv Pinotage geïsoleer is, inhibeer ’n kru EPG-ekstrak van bogenoemde ras baie effektief. Die benadering wat gevolg is het op die ooruitdrukking van die individuele Bcpg-gene in heteroloë sisteme staatgemaak en die gevolglike isolering van suiwer en aktiewe ensieme om EPG-inhibisie deur VvPGIP1 te beoordeel. Al die gene is suksesvol in Saccharomyces cerevisiae ooruitgedruk onder ’n sterk induseerbare promotor, maar volgens ’n agarose-diffundeerbare toets kon aktiewe ensiempreparate slegs vir die enkoderende Bcpg2 verkry word. Die in vitro PGIP-inhibisie toets is ook op die gemelde toets gebasseer en vereis EPG-aktiwiteit om die inhiberende effek van die PGIP, te visualiseer. Die aktiewe EPG2 is egter nie deur VvPGIP1 geïnhibeer met die aanleg van hierdie toets nie. Die EPG-enkoderende gene van B. cinerea is ook tydelik in Nicotiana benthamiana ooruitgedruk deur gebruik te maak van ’n Agrobacterium-infiltrasietegniek. Transgeenuitdrukking kon met die Noordelike kladtegniek bevestig word en EPG-verwante simptome is vyf tot agt dae na infiltrasie waargeneem. Verskillende simptome vir die verskillende EPGs is waargeneem, wat aanduidend is dat die simptome nie lukrake gevolge van die infiltrasies, of ’n hipersensitiewe respons is nie. Verder kon die simptome wat EPG2 vertoon het, gekorreleer word met dié wat onlangs deur ’n ander groep op dieselfde gasheer waargeneem is. Ten spyte van die ekspressiedata en die waargenome simptome, kon aktiewe ensiempreparate op die agarose-diffundeerbare toets, weereens slegs vir EPG2 waargeneem word. ’n Metode wat gebasseer is op chlorofilfluoressensie is getoets en geëvalueer as ’n moontlike in vivo metode om EPG aktiwiteit en inhibisie deur PGIPs waar te neem en te kwantifiseer. Die resultate het bevestig dat die ooruitdrukking van hierdie gene die elektronvloeitempo deur fotosisteem II verminder het wat ’n aanduiding is dat metaboliese stres in die plant heers. Dieselfde tegniek is gebruik om die moontlike interaksies tussen BcPG1 en 2 en VvPGIP1 te bestudeer en het aangetoon dat die mede-uitdrukking van die Vvpgip1-geen aanleiding gee tot ’n beskermende effek van die geinfiltreerde weefsel, wat aanduidend is van inhibisie van EPG1 en 2 deur VvPGIP1. In die geval van EPG2 is hierdie interaksie en moontlike inhibisie met ’n PGIP die eerste waarneming in die verband. In ’n verdere poging om die in planta-interaksie tussen VvPGIP1 en die EPGs van die Suid-Afrikaanse B. cinerea ras uit te klaar, is ’n plantgebasseerde twee-hibriede toets aangelê, maar geen klinkklare resultate kon verkry word nie. Vorige werk het bevestig dat verskeie natuurlike mutasies in PGIP-enkoderende gene, afkomstig van verskillende V. vinifera kultivars, voorkom. Hierdie resultaat en die feit dat hierdie mutasies verskille in EPG inhibisie en uiteindelik vatbaarheid vir siektes kan beïnvloed, het aanleiding gegee tot die isolering van ’n verdere 37 PGIP-enkoderende gene uit ‘n verskeidenheid druifplantgenotipes, sommige waarvan juis bekend vir hul weerstand teen patogene is. Die gekombineerde resultate wat in dié studie verkry is, maak ’n waardevolle bydrae tot die verstaan van plant-patogeeninteraksies, spesifiek met die modelering van interaksies van patogeen- en plantgebasseerde proteïene. Die moontlikheid om in vivo-metodes soos chlorofilfluoressensie te gebruik in in planta-analises, is besonder bemoedigend om in vitro-resultate te versterk en ook in konteks te plaas.

Thesis (M.Sc. (Biochemistry, Microbiology & Biotechnology)) - Rhodes University, 2008.

Thesis (MSc (Genetics))--University of Stellenbosch, 2009. / Worldwide, the rust diseases cause significant annual wheat yield losses (Wallwork 1992; Chrispeels & Sadava 1994). The utilization of host plant resistance to reduce such losses is of great importance particularly because biological control avoids the negative environmental impact of agricultural chemicals (Dedryver et al. 1996). The wild relatives of wheat are a ready source of genes for resistance to disease and insect pests. A large degree of gene synteny still exists among wheat and its wild relatives (Newbury & Paterson 2003). It is therefore possible to transfer a chromosome segment containing useful genes to a homologous region in the recipient genome without serious disruption of genetic information. Special cytogenetic techniques are employed to transfer genes from the wild relatives to the wheat genomes (Knott 1989). Unfortunately the transfer of useful genes may be accompanied by the simultaneous transfer of undesirable genes or redundant species chromatin which has to be mapped and removed (Feuillet et al. 2007). DNA markers are extremely useful for the characterisation and shortening of introgressed regions containing genes of interest (Ranade et al. 2001), and may also be used for marker aided selection of the resistance when the genes are employed commercially. Eight wheat lines containing translocations/introgressions of wild species-derived resistance genes were developed by the Department of Genetics (SU). These lines are presently being characterized and mapped and attempts are also being made to shorten the respective translocations. This study aimed to find DNA markers for the various translocations and to convert these into more reliable SCAR markers that can be used in continued attempts to characterize and improve the respective resistance sources. A total of 260 RAPD and 21 RGAP primers were used to screen the eight translocations and, with the exception of Lr19, it was possible to identify polymorpic bands associated with each translocation. However, it was not possible to convert all of these into more reliable SCAR markers. The primary reason for this was the low repeatability of most of the bands. Certain marker fragments turned out to be repeatable but could not be converted successfully. Some of the latter can, however, be used directly (in RAPD or RGAP reactions) as markers. The Lr19 translocation used in the study (Lr19-149-299) is a significantly reduced version of the original translocation and failure to identify polymorphisms associated with it can probably be ascribed to its small size. The following numbers of markers (direct and converted into SCARs) were Worldwide, the rust diseases cause significant annual wheat yield losses (Wallwork 1992; Chrispeels & Sadava 1994). The utilization of host plant resistance to reduce such losses is of great importance particularly because biological control avoids the negative environmental impact of agricultural chemicals (Dedryver et al. 1996). The wild relatives of wheat are a ready source of genes for resistance to disease and insect pests. A large degree of gene synteny still exists among wheat and its wild relatives (Newbury & Paterson 2003). It is therefore possible to transfer a chromosome segment containing useful genes to a homologous region in the recipient genome without serious disruption of genetic information. Special cytogenetic techniques are employed to transfer genes from the wild relatives to the wheat genomes (Knott 1989). Unfortunately the transfer of useful genes may be accompanied by the simultaneous transfer of undesirable genes or redundant species chromatin which has to be mapped and removed (Feuillet et al. 2007). DNA markers are extremely useful for the characterisation and shortening of introgressed regions containing genes of interest (Ranade et al. 2001), and may also be used for marker aided selection of the resistance when the genes are employed commercially. Eight wheat lines containing translocations/introgressions of wild species-derived resistance genes were developed by the Department of Genetics (SU). These lines are presently being characterized and mapped and attempts are also being made to shorten the respective translocations. This study aimed to find DNA markers for the various translocations and to convert these into more reliable SCAR markers that can be used in continued attempts to characterize and improve the respective resistance sources. A total of 260 RAPD and 21 RGAP primers were used to screen the eight translocations and, with the exception of Lr19, it was possible to identify polymorpic bands associated with each translocation. However, it was not possible to convert all of these into more reliable SCAR markers. The primary reason for this was the low repeatability of most of the bands. Certain marker fragments turned out to be repeatable but could not be converted successfully. Some of the latter can, however, be used directly (in RAPD or RGAP reactions) as markers. The Lr19 translocation used in the study (Lr19-149-299) is a significantly reduced version of the original translocation and failure to identify polymorphisms associated with it can probably be ascribed to its small size. The following numbers of markers (direct and converted into SCARs) were v identified: S8-introgression (Triticum dicoccoides) = one RAPD and two SCARs; S13-translocation (Aegilops speltoides) = four RAPDs, three RGAPs and five SCARs; S15-translocation (Ae. peregrina) = one RAPD and two SCARs; S20-translocation (Ae. neglecta) = two RAPDs, two RGAPs and one SCAR. The markers are already being employed in current projects aiming to map and shorten these translocations. Some of the markers can be combined in multiplex reactions for more effective mass screening. No repeatable markers could be identified for the four remaining translocations (S12 from Ae. sharonensis; S14 from Ae. kotschyi; Smac from Ae. biuncialis and Lr19-149-299 from Thinopyrum ponticum).

Thesis (MSc (Genetics))--University of Stellenbosch, 2011. / Includes bibliography / ENGLISH ABSTRACT: This study investigated the use of antimicrobial peptides (AMPs) as possible source of resistance against a range of pathogens in grapevine. Whilst the ultimate aim would be to express AMPs in grapevine, the development of transgenic grapevine is time consuming and therefore pre-screening of potential AMPs is necessary. These small molecules, of less than 50 amino acids in length, are expressed by almost all organisms as part of their non-specific defence system. In vitro pre-screening of AMP activity is valuable but is limited since the activity on artificial media may differ from the AMP activity in planta. These tests are also restricted to pathogens which can be cultured in vitro. These limitations can be overcome by using transient expression systems to determine the in planta activity of AMPs against pathogens of interest. In this study transient systems were used to express AMPs in developed plant tissue to test their efficacy against grapevine pathogens such as Agrobacterium vitis, Xylophilus ampelinus and aster yellows phytoplasma. Aster yellows phytoplasma, which was recently discovered in local vineyards, is known to cause extensive damage and therefore pose a great threat to the South African grapevine industry. To study the in planta effect of AMPs against the abovementioned pathogens, transient expression vectors were constructed expressing either of the AMPs D4E1 or Vv-AMP1. D4E1 is a synthetically designed AMP known to be active against bacteria and fungi, while Vv-AMP1, isolated from grapevine berries, has already shown activity against fungi. In a transient approach in grapevine, the expression of foreign genes from viral and non-viral vectors was confirmed by expression of the marker genes β-glucuronidase and Green Fluorescent Protein, while tissue-printing immunoassays confirmed viral replication and systemic spread in Nicotiana benthamiana. The viral vectors were based on the phloem-limited virus grapevine virus A. Only Agrobacterium-mediated 35S transient expression vectors were used for AMP in planta activity screening since the viral-mediated expression in grapevine was insufficient for screening against A. vitis and X. ampelinus as it was restricted to phloem tissues after whole-leaf infiltration. No phytoplasma-infected material could be established and as a result AMP activity screening was only performed against the A. vitis and X. ampelinus. Quantification of the bacteria was performed by qPCR. Vv-AMP1 did not show activity against either of the two bacteria in planta while D4E1 was found to be active against both. The observed in planta activity of D4E1 correlated with the in vitro activity as measured in an AMP plate bioassay. In contrast to in vitro screenings, the in planta AMP activity screening might give a more accurate representation of the potential antimicrobial activity of the peptide in a transgenic plant environment. This study proved that transient expression systems can be used as a pre-screening method of AMP activity in planta against grapevine pathogens, allowing the screening of various AMPs in a relatively short period of time before committing to transgenic grapevine development. / AFRIKAANSE OPSOMMING: Hierdie studie het die gebruik van antimikrobiese peptiede (AMPe) as 'n moontlik bron van weerstand teen 'n reeks van patogene in wingerd ondersoek. Alhoewel die uiteindelike doel sal wees om AMPe uit te druk in wingerd, is transgeniese wingerd ontwikkeling tydrowend en daarom is vooraf evaluering van potensiële AMPe nodig. Hierdie klein molekules, van minder as 50 aminosure in lengte, word uitgedruk deur amper alle organismes as deel van hul nie-spesifieke verdedigingsisteem. In vitro vooraf evaluering van AMP aktiwiteit is van waarde, maar is beperk aangesien die aktiwiteit op kunsmatige media mag verskil van die AMP-aktiwiteit in planta. Hierdie toetse is ook beperk tot patogene wat in vitro gekweek kan word. Hierdie beperkinge kan oorkom word deur gebruik te maak van tydelike uitdrukkingsisteme om die in planta aktiwiteit van AMPe te bepaal teen patogene van belang. In hierdie studie is tydelike uitdrukkingsisteme gebruik om AMPe uit te druk in ontwikkelde plantweefsel om hul effektiwiteite te toets teen wingerdpatogene soos Agrobacterium vitis, Xylophilus ampelinus en aster yellows fitoplasma. Aster yellows fitoplasmas, wat onlangs in plaaslike wingerde ontdek is, is bekend vir die uitgebreide skade wat hul aanrig en hou daarom 'n groot bedreiging in vir die Suid-Afrikaanse wingerd industrie. Om die in planta effek van AMPe teen die bogenoemde patogene te bestudeer is tydelike uitdrukkingsvektore ontwikkel wat die AMPe D4E1 of Vv-AMP1 uitdruk. D4E1 is 'n sinteties-ontwerpte AMP wat aktief is teen bakterieë en fungi, terwyl Vv-AMP1, wat uit druiwekorrels geïsoleer is, alreeds aktiwiteit teen fungi getoon het. In 'n tydelike uitdrukkingsbenadering in wingerd is die uitdrukking van transgene, vanaf virus of nie-virus gebaseerde vektore, bevestig deur die uitdrukking van die merker gene β-glukuronidase en die Groen Fluoresserende Proteïen, terwyl weefsel afdrukkings-immunotoetse virus replisering en sistemiese beweging in Nicotiana benthamiana bevestig het. Die virusvektore was gebaseer op die floëem-beperkte virus, wingerdvirus A. Slegs Agrobacterium-bemiddelde 35S tydelike uitdrukkingsvektore is gebruik om die AMP in planta aktiwiteit te bepaal aangesien die virus-bemiddelde uitdrukking in wingerd onvoldoende was vir evaluering teen A. vitis en X. ampelinus weens die beperking tot die floëem weefsel na infiltrering van die totale blaar. Geen fitoplasma geïnfekteerde materiaal kon gevestig word nie, en daarom is AMP aktiwiteitsevaluering slegs teen A. vitis en X. ampelinus uitgevoer. Kwantifisering van die bakterieë is deur middel van qPCR uitgevoer. Vv-AMP1 het geen aktiwiteit getoon teen enige van die bakterieë in planta nie, terwyl D4E1 aktief was teen beide. Die waargenome in planta aktiwiteit van D4E1 het ooreengestem met die in vitro aktiwiteit soos bepaal deur 'n AMP plaat bio-toets. In kontras tot in vitro evaluering kan die in planta AMP-aktiwiteit evaluering 'n meer akkurate voorspelling bied van die potensiële antimikrobiese aktiwiteite van die peptied in 'n transgeniese plant omgewing. Hierdie studie het bewys dat tydelike uitdrukkingsisteme gebruik kan word as 'n voorafgaande evalueringsmetode vir AMP in planta aktiwiteit teen wingerdpatogene, wat die evaluering van 'n verskeidenheid AMPe in 'n relatiewe kort tydperk toelaat voor verbintenis tot die ontwikkeling van transgeniese wingerd.

The challenges faced by the agricultural sector especially around improving production yields using environmentally friendly solutions have received market attention. Biological intervention can range from application of biological products to enhance the nutritional value of crops or to control plant pathogens. Biostart, a biological product that demonstrated growth enhancement when applied in lettuce crops is currently in the market. The product is comprised of a consortium of bacterial isolates (Bacillus licheniformis, Brevibacillus laterosporus and Bacillus laterosporus) but the contribution of the individual isolates to growth enhancement had not been elucidated. Green house experiments on lettuce seedlings with individual and mixed treatments were commissioned to determine such contribution. There was either no or marginal growth enhancement observed in the experiments. The results showed that the product was effective as a consortium and not as individual isolates. Further isolation and screening for potential Bacilli with antifungal properties was undertaken. An isolate identified as Bacillus subtilis that demonstrated inhibition against a wide spectrum of fungi, and especially the phytopathogenic Verticillium dahliae and Fusarium oxysporum, was successfully identified. The isolate was cryo-preserved and cultivated to significant levels at bench scale. A characterized comparison of different putative products with known systematic fungicide showed potential application even of heat treated products. The product showed control V. dahliae when tested in green houses with potatoes and tomatoes as test crops. This isolate has been targeted for further development as a biological control product.

Thesis (M. Tech) --Central University of Technology, Free State, 2008 / Aphids (Hemiptera: Aphididae) are amongst the most destructive insects in agricultural crop production systems. This reputation stems from their complex life cycles which are mostly linked to a parthenogenetic mode of reproduction, allowing them to reach immense population sizes within a short period of time. They are also notorious as important and efficient vectors of several plant viral diseases. Their short fecund life cycles allow them to be pests on crops with a short growth period, e.g. lettuce (Lactuca sativa L.). It is common practice to provide this crop with some degree of protection from environmental extremes on the South African Highveld. Shadehouses are popular in this regard, but aphids are small enough to find their way into these structures, and their presence on lettuce is discouraged due to phytosanitary issues. In addition, the excessive use of insecticides is criticized due to the negative influence on human health, and because aphids can rapidly develop resistance. This necessitates the use of alternative control options in order to suppress aphid numbers. Biological control is popular in this regard and the use of predatory ladybirds (Coleoptera: Coccinellidae) is a popular choice. This study investigated the aphid and coccinellid species complex encountered under varying shadehouse conditions on cultivated head lettuce in the central Free State Province (South Africa). Their seasonality was also examined, along with variations in their population size throughout a one-year period. Finally, the impact of varying aphid populations on some physical characteristics of head lettuce was examined, and recommendations for aphid control (using naturally occurring coccinellid predators) were made. Two shadehouse structures were evaluated during this study. One was fully covered with shade netting and designed to exclude the pugnacious ant, Anoplolepis custodiens (Hymenoptera: Formicidae), while the other was partially covered with shade netting (on the roof area) allowing access to the ants. Six cycles of head lettuce were planted and sampled four times during each cycle. These were scheduled to monitor the seedling, vegetative and heading stage of lettuce. Four important aphid species were recorded on the lettuce, namely Acyrthosiphon lactucae, Nasonovia ribisnigri, Myzus persicae and Macrosiphum euphorbiae. Both structures harboured similar aphid and coccinellid species, but their population dynamics differed. A. lactucae dominated in the absence of A. custodiens in the fully covered structure (whole study), while N. ribisnigri dominated in the partially covered structure in the presence of these ants during the warmer months (December – January). M. euphorbiae replaced this species as the dominant species in the absence of A. custodiens (April – September). M. persicae occured during the winter (May – August) in the fully covered structure. Promising coccinellid predators were Hippodamia variegata and Scymnus sp. 1, and to a lesser extent, Exochomus flavipes and Cheilomenes lunata. However, the fully covered structure hampered the entrance of the larger adult coccinellid species, resulting in their lower occurrence. Aphid and coccinellid activity peaked during the summer months (October – January), and the fully covered structure attained the highest aphid infestation levels and coccinellid larval numbers during this time. On the other hand, aphid numbers were higher in the partially covered structure during the cooler months of the year (April – July) and this structure also harboured more adult coccinellids. In most cases, aphid infestation levels did not affect the amount of leaves formed. However, symptomatic damage in terms of head weight reduction did occur under severe infestation levels. Specific environmental conditions within a shadehouse structure concurrently contributed to this reduction, with less favourable conditions accelerating this condition. Results from this study have shown that even though the type of shadehouse structure does not influence the insect species complex found on lettuce, it does have an influence on detrimental and beneficial insect population dynamics. Aphid species infesting lettuce have been identified, along with coccinellid predators that could potentially be used in their control. Both types of structures had advantages and disadvantages, and therefore, decisions concerning shadehouses should not be focused on which type of structure to use, but rather which type of structure to use during different seasons of the year.