Thesis (MSc)--University of Stellenbosch, 2007. / ENGLISH ABSTRACT: Presently, the observed variation in symptoms of Potato leafroll virus (PLRV) infection in potato cultivars in South Africa cannot be reconciled with PLRV symptoms obtained 10-15 years ago, even if the different interactions between the pathogen and the cultivar are taken into account. In an effort to analyze this variation, mutations in the coat protein (CP) gene of South African isolates of PLRV were assessed. The CP gene of PLRV isolates from different areas within South Africa was amplified by reverse transcription-polymerase chain reaction (RT-PCR), cloned and sequenced. Significant sequence variation in the CP gene was found within the analyzed South African isolates of PLRV. Phylogenetic analysis revealed two major clades with most South African isolates and an Australian and North American isolate grouped together and the remainder grouped with isolates from diverse countries worldwide. The deduced amino acid sequences from representatives of these two clades indicated differences in CP threedimensional structure. In an effort to produce recombinant PLRV CP for the production of antibodies specific for South African isolates of PLRV for use in enzyme-linked immunosorbent assay (ELISA), the CP gene of a South African isolate of PLRV was subcloned into a bacterial expression vector (pET14-b). Expression of full length recombinant PLRV CP was attempted in Escherichia coli strains BL21(DE3)pLysS, Rosetta-gami B(DE3)pLysS and Rosetta-2(DE3)pLysS. As this was not successful, the PLRV CP gene was subcloned in another expression vector (pGEX) for expression as an N-terminal fusion protein with glutathione-S-transferase (GST) in E. coli strains BL21(DE3)pLysS and Rosetta-2(DE3)pLysS. The recombinant GST-PLRV CP fusion protein was purified and used for antibody production in rabbits. Using western blots, the effectiveness of antibodies produced to recombinant GST-PLRV CP fusion protein was assessed for PLRV recognition. It was found that antibodies to the recombinant GST-PLRV CP fusion protein were more effective for the detection of GST than PLRV CP and that production of antibodies to the cleaved PLRV CP product would be necessary if antibodies are required for ELISA applications. / AFRIKAANSE OPSOMMING: Huidiglik kan die waargeneemde simptome van infeksie met aartappelrolbladvirus (Potato leafroll virus, PLRV) in aartappelkultivars in Suid-Afrika nie vereenselwig word met PLRV simptome wat 10-15 jaar gelede verkry was nie, selfs al word die verskillende interaksies tussen die patogeen en kultivar in ag geneem. In ‘n poging om hierdie variasie te analiseer, was mutasies in die mantelproteïen (CP) geen van Suid-Afrikaanse isolate van PLRV bepaal. Die CP geen van PLRV isolate van verskillende areas in Suid-Afrika was ge-amplifiseer met behulp van die tru transkripsie-polimerase ketting reaksie (RT-PCR), gekloneer en die nukleotiedvolgorde bepaal. Noemenswaardige nukleotied variasie is in die CP gene van die ge-analiseerde Suid-Afrikaanse isolate van PLRV gevind. Filogenetiese analises het gedui op twee hoof klades met die meeste van die Suid-Afrikaanse isolate wat saam met ‘n Australiese en Noord-Amerikaanse isolaat gegroepeer en die res wat met isolate van verskillende lande wêreldwyd gegroepeer. Die afgeleide aminosuurvolgordes van verteenwoordigers van bogenoemde twee klades het gedui op verskille in die CP driedimensionele struktuur. In ‘n poging om rekombinante PLRV CP te produseer vir die produksie van antiliggame spesifiek teen Suid-Afrikaanse isolate van PLRV om in “enzyme-linked immunosorbent assay” (ELISA) te gebruik, was die CP geen van ‘n Suid-Afrikaanse isolaat van PLRV gesubkloneer in ‘n bakteriële ekspressie vektor (pET14-b). Daar was gepoog om vollengte rekombinante PLRV CP in die Escherichia coli rasse BL21(DE3)pLysS, Rosetta-gami B(DE3)pLysS en Rosetta- 2(DE3)pLysS te produseer. Aangesien dit nie suksesvol was nie, was die PLRV CP gesubkloneer in ‘n ander ekspressie vektor (pGEX) sodat die proteïen as ‘n N-terminale fusie proteïen met “glutathione-S-transferase” (GST) in E. coli rasse BL21(DE3)pLysS en Rosetta- 2(DE3)pLysS geproduseer kon word. Die rekombinante GST-PLRV CP fusie proteïen was gesuiwer en gebruik vir antiliggaam produksie in konyne. Die effektiwiteit van die antiliggame wat teen rekombinante GST-PLRV CP fusie proteïen geproduseer was vir PLRV herkenning is deur middel van “western blots” geanaliseer. Dit was gevind dat antiliggame teen die rekombinante GST-PLRV CP fusie proteïen meer effektief was vir die herkenning van GST as PLRV CP. Gevolglik sal dit nodig wees om antiliggame teen die gesnyde PLRV CP produk te maak vir gebruik in ELISA.

Graduation date: 1998

In the past 10 years we have witnessed the beginnings of the study of soil ecology as a unified science, and the general realization by soil scientists, farmers, and land managers that many of the most important economic aspects of soil health are controlled by biological factors. This research focuses on alterations in a tropical soil microarthropod community under differing intensive agricultural protocols: native vegetation, pastures and cropland, during June, July and August 1998. The effect of pesticides in potato cultivation was also studied. In the Ecuadorean montane forest, 361 morphospecies of soil arthropods, were classified during the three sampling months. August was the month with highest abundance and diversity. Acari, Coleoptera, Collembola, Diptera and Homoptera were the most abundant orders present in all the three types of land management. The native vegetation had the most abundant and diverse representation of all soil arthropod taxa compared to the pastures and croplands. Coleoptera, Diptera, Diplopoda, Diplura and Hemiptera were significantly more diverse in native vegetation than in pastures and croplands. The most abundant functional groups were the fungivores, herbivores and predators. The abundance of functional groups was significantly higher in the native vegetation for predators, herbivores and detritivores. 115 morphospecies of soil arthropods were identified in the study of arthropod response to pesticides in a complete randomized potato plot. Seasonal effects were documented for Acari, Collembola, Diptera, and Homoptera. Predators were most abundant in July and fungivores decreased in September. Neither arthropod orders nor functional groups showed a significant change in abundance between different treatments. Only Homoptera showed an increase in its abundance in the third sampling date and only in the Antracol plots. The potato plants in the whole block showed poor productivity, suggesting that the whole system was stressed by the fungal pest. / Graduation date: 2000

No description available.

Potato ranks fourth after wheat, rice and corn as a major food crop. It is an excellent source of nutrients and at the same time relatively inexpensive, therefore it is the mainstay in the diets of people in both developed and developing countries. Potato losses and quality degradation due to the effect of storage variables or processing conditions must be reduced to increase the world food supply. To accomplish these objectives, studies have been conducted to evaluate: (a) quality changes in potatoes during cooking and frying, (b) quality changes in raw potatoes as influenced by storage conditions, (c) changes in cooking quality of potatoes as influence by storage conditions, (d) changes in frying quality of potatoes as influenced by storage conditions, and finally (e) quality changes in potatoes as influenced by Erwinia carotovora ssp. carotovora infection (Ecc, casual agent of soft rot disease in potato). / Kinetics of quality changes during cooking and frying of potatoes were evaluated. Potatoes were cooked at 80--100°C or fried at 160--190°C for selected times and their texture and color were evaluated. Results showed that texture values of cooked potatoes decreased with the progress of cooking, and the rate of texture changes at each temperature was found to be consistent with two pseudo first-order kinetic mechanisms. Textural values of fried potatoes increased with frying time and followed a first order kinetic model. Cooked potatoes were less bright, more red and less yellow in color as compared to raw samples. A modified first order model was used to characterize the color changes kinetics of both cooked and fried potatoes based on the changes occurring between the initial and a maximum or minimum value. 10 min cooking at 100°C and 10 min frying at 180°C were considered to give the designed cooked and fried products, respectively. / The changes in quality characteristics of potatoes as a function of storage variables (temperature and time) were evaluated. Potatoes were stored at five temperatures (4, 8, 12, 16 and 20°C) for selected duration (at least 5 time intervals) and different physico-chemical quality parameters were evaluated. Potatoes remained healthier when they stored at lower temperatures due to absence of sprouts and visible spoilage. They became softer and darker by passage of time.

Many parasitic diseases display symptoms beyond the areas invaded by the parasite itself. This suggests that toxic metabolic products of the pathogen diffusing in front of its advancing growth brings about derangements in unoccupied tissues at times far remote from the focus of infection. [...]

No description available.

Potato leafroll virus (PLRV) is one of the most destructive potato viruses in South Africa. In order to establish resistance against PLRV in commercial potato cultivars, the coat protein (CP) gene of the virus was previously isolated, cloned and subcloned into the plant expression vector pBI121 in both the sense and antisense orientations (BURGER, unpublished results). The pBI121 constructs containing the PLRV-CP gene were subsequently transferred to Agrobacterium tumefaciens LBA 4404 in a triparental mating process with the helper plasmid pRK2013. Two A. tumefaciens- mediated transformation methods for potatoes were investigated, viz. vacuum infiltration and leaf disk transformation. In addition, optimal transformation and regeneration conditions were identified for potato cultivars Late Harvest and BP[1] In total, 27 transgenic potato lines containing the PLRV-CP, β-glucoronidase (GUS) and nptII (neomycin phosphotransferase II) trans genes were generated under kanamycin selection. Transgenic plants grown in the glasshouse appeared to be phenotypically normal, and no differences in ploidy level in comparison to non-transformed plants could be established. Stable transgene insertion into the genome of the transgenic plants was verified using PCR and Southern blot analysis. Expression of the GUS transgene was investigated using a fluorometric assay (JEFFERSON et al. 1987), and it was found that orientation of the inserted PLRV-CP gene upstream from the GUS gene had a direct influence on the levels of GUS expression. The expression of the PLRV-CP gene was analysed using DAS-ELISA and immunoblot detection. Coat protein could not be detected in either assay. RNA dot blots were used successfully to show PLRV-CP expression in transgenic potato plants at the mRNA level. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1995.

No description available.

No description available.