Braunschweig, Techn. Universiẗat, Diss., 2000.

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Thesis (MSc)--Stellenbosch University, 2015. / ENGLISH ABSTRACT: South Africa is ranked amongst the top ten for wine production internationally. Viticulture contributes immensely to the economy, which justifies research into the pathogens that may negatively affect wine production. Aster Yellows phytoplasma was reported in South African vineyards in 2010 and has since been an ongoing problem for grape farmers in affected areas. Throughout the world, phytoplasma diseases such as Grapevine Yellows have caused detrimental effects on the vines, often resulting in death. The limited knowledge on prevention and control of the pathogen can be attributed to the lack of full understanding of the epidemiology and accurate diagnosis. The aim of this study was to determine the spatial distribution of Aster Yellows phytoplasma in individual grapevines and to record a possible temporal or seasonal distribution. The recovery phenotype phenomenon was encountered during the study and surveys were conducted in order to determine whether recovery was permanent. In order to perform the studies, a reliable assay to accurately detect the pathogen in grapevines was required. A comparison between three assays was completed in furtherance of deciding which to use for the further experimentation. The three assays included a nested PCR utilizing universal primers, a Real-Time PCR using Syto9 as a double stranded DNA specific dye and a Real- Time PCR with a TaqMan® probe using an identical dilution series. Of the three assays tested, the nested PCR proved to be the most sensitive diagnostic procedure, detecting Aster Yellows phytoplasma in very low titers and was thus used for diagnostics in further experiments. In order to determine the spatial patterns of Aster yellows phytoplasma infection, leaf, petiole, trunk, root and cane samples were taken from three whole grapevine plants. Phloem scrapings obtained from the cane samples yielded more positive results in comparison to the other parts of the plant tested. Not only do phytoplasmas display an erratic spatial distribution, but also have a tendency to change over time. Thirty symptomatic grapevines were sampled over one and a half growing seasons, with results concluding that February yielded the most positive diagnoses. Fifty plants that had been previously pruned back and no longer displayed symptoms were also sampled in 2013 and 2014, and all yielded negative results over both years. This study contributes to comprehension of Aster Yellows phytoplasma epidemiology and ultimately the advancement of accurate diagnosis. / AFRIKAANSE OPSOMMING: Suid-Afrika is internasionaal geposisioneer onder die top tien vir die produksie van wyn. Wingerd dra geweldig by tot die ekonomie, wat navorsing oor die patogene wat wynporduksie negatief beïnvloed, regverdig. Aster Yellows phytoplasmais in 2010 gerapporteer in Suid-Afrikaanse wingerde en is sedertdien 'n deurlopende probleem vir druiweboere in geaffekteerde gebiede. Dwarsdeur die wêreld, het fitoplasma siektes soos Grapevine Yellows ‘n nadelige uitwerking op wingerde, wat dikwels lei tot plantsterftes. Die beperkte kennis oor die voorkoming en beheer van die patogeen kan toegeskryf word aan die gebrek aan begrip van die epidemiologie en akkurate diagnose . Die doel van hierdie studie was om die ruimtelike verspreiding van Aster geel fitoplasma in individuele wingerdstokke te bepaal en 'n moontlike tydelike of seisoenale verspreiding aan te teken. Die herstel-fenotipe verskynsel is tydens die studie teëgekom en opnames is uitgevoer ten einde te bepaal of die herstel permanent was. Ten einde die studie uit te voer , is 'n betroubare toets vereis om die patogeen in wingerde akkuraat te spoor. : Drie toetse is vergelyk (en geëvalueer) vir hulle geskikthed vir gebruik in die studie. Die drie toetse het ingesluit 'n geneste PKR wat gebruik maak van universele primers, 'n in-tydse PKR (real-time PCR) wat Syto9 gebruik as 'n dubbelstring DNS spesifieke kleurstof, en 'n in-tydse PKR met 'n TaqMan® peiler, en is vergelyk met behulp van 'n identiese vedunnings reeks. Van die drie toetse , is die geneste PCR bewys om die mees sensitiewe diagnostiese prosedure te wees , en kon Aster geel fitoplasma in baie lae titers opspoor en is dus gebruik vir die diagnose in verdere eksperimente. Ten einde die ruimtelike patrone van Aster geel fitoplasma infeksie te bepaal, is blaar, blaarsteel, stam, wortel en loot monsters van drie volle wingerdstokke geneem. Floëem skraapsels verkry uit die loot monsters het meer positiewe resultate opgelewer in vergelyking met die ander dele van die plant. Nie net vertoon phytoplasmas 'n wisselvallige ruimtelike verspreiding nie, maar het ook 'n neiging om te verander met verloop van tyd. Dertig simptomaties wingerdstokke is versamel oor een en 'n half groeiseisoene,en die resultate het gewys dat Februarie die meeste positiewe diagnoses het. Monsters is versamel in 2013 en 2014 van vyftig plante wat voorheen teruggesnoei is en nie meer simptome vertoon nie, en alle monsters het negatiewe resultate opgelewer oor beide jare. Hierdie studie dra by tot begrip van Aster geel fitoplasma epidemiologie en uiteindelik die bevordering van akkurate diagnose.

Includes bibliographical references (leaves 158-180) Appendix A. Vineyard disease survey maps -- appendix B. Log linear graphs

Includes bibliographical references (leaves 158-180) / xiii, [220] leaves : ill. (col.), maps ; 30 cm / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Plant Science, 2002

Beet yellows virus (BYV) is a filamentous, positive-strand RNA virus that belongs to the family Closteroviridae. BYV particles encapsidate a 15.5 kb RNA and posses complex polar architecture. A long virion body is formed by the major capsid protein(CP), whereas the minor capsid protein (CPm) assembles a short tail that encapsidates the 5'-terminal region of BYV RNA. In addition to proteins required for viral RNA replication and encapsidation, BYV encodes four proteins whose role in the virus life cycle was unknown. These proteins include a small, 6-kDa, hydrophobic protein (p6), a homolog of the cellular 70-kDa heat shock proteins (Hsp7Oh), a 64-kDa protein (p64), and a 20-kDa protein (p20). It was found recently that Hsp7Oh, p64, and p20 are incorporated into BYV virions, and that Hsp7Oh is required for the virus movement from cell to cell. In this study, we characterized genetic requirements for BYV assembly and cell-to-cell movement, and determined relationships between these two processes. It was demonstrated that in addition to Hsp7Oh, p6, p64, CP, and CPm are each essential, but not sufficient for virus movement. These results indicated that five-component movement machinery of BYV is the most complex among plant viruses. Extensive mutational analysis of CP and CPm revealed strong correlation between abilities of BYV to assemble tailed virions and to move from cell to cell, suggesting that formation of functional virions is a prerequisite for virus translocation. We have found that CPm, Hsp7Oh, and p64 are necessary for the efficient virion tail formation. Assembly of the virion tails and bodies was shown to occur independent of each other and likely to involve two separate packaging signals within the genomic RNA. Our work demonstrated that BYV encodes one conventional movement protein, p6, whose only known function is to mediate virus movement. The other four movement associated proteins of BYV, CP, CPm, Hsp7Oh, and p64 are the virion components, each of which is required for assembly of the tailed, movement-competent virions. Based on these and other data, we propose that BYV and other closteroviruses evolved virion tails as a specialized device for the directional cell-to-cell movement of large RNA genomes. / Graduation date: 2005 / Best scan available.

More than half of the recognized genera of positive strand RNA viruses employ polyprotein processing as one of the strategies for their genome expression. Normally, this processing is mediated by virus-encoded proteinases that belong to the trypsin-like or papain-like family. In particular, papain-like, leader proteinases were found in diverse families of human, animal, plant, and fungal positive strand RNA viruses. In addition to autocatalytic processing, these proteinases play a variety of roles in the virus life cycle. In plant potyviruses, a papain-like helper component-proteinase (HC-Pro) was implicated in genome amplification, cell-to-cell movement, long distance transport, and suppression of host defense. The p29 proteinase encoded by a fungal hypovirus CHV1 was found to be dispensable for virus replication, but it was identified as a major determinant of viral pathogenicity. In an animal equine aterivirus (EAV), a papain-like proteinase nspl was demonstrated to possess a putative zinc finger domain, which functions in subgenomic RNA synthesis, although it is not essential for virus replication. The Lab proteinase of the foot and mouse disease virus (FMDV) is involved in inhibition of cellular mRNA translation and in virus spread in infected animals. In general, it appears that functional plasticity of the papain-like leader proteinases played an important role in the evolution of viral diversity. Here, we examined the functions of a papain-like leader proteinase (L-Pro) in the life cycle of the beet yellows closterovirus (BYV). It was found that L-Pro is required for autoproteolytic processing, genome amplification, virus invasiveness and cell-to-cell movement for BYV. The gene swapping experiments involving several closterviruses, a potyvirus, as well as CHV1, FMDV, and EAV revealed complex functional profiles of the papain-like leader proteinases. The possible mechanisms that underlie L-Pro functions are discussed. / Graduation date: 2002

Thesis (Ph. D.)--University of Wisconsin--Madison, 1958. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 121-126).

Techn. University, Diss., 2000--Braunschweig.

In total, 801 samples of potato leafroll disease were collected and tested for potato leafroll virus (PLRV) and beet western yellows virus (BWYV) in 1986, 1987, and 1988 using triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA) and virus-specific monoclonal antibodies. The samples represented 32 cultivars and originated in eight Canadian provinces and 12 American states. None of the samples tested positive for BWYV, whereas 772 (96.4%) tested positive for PLRV. Neither PLRV nor BWYV could be recovered, with aphid transfers to indicator hosts, from 28 of the 29 samples that tested negative for both viruses. PLRV was recovered from one sample that originally tested negative by TAS-ELISA; the indicator plant tested positive for PLRV by TAS-ELISA. Nucleic acid spot hybridization (NASH) using random primed and cloned cDNA probes was compared with double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) and TAS-ELISA, and aphid transmission tests for detection and identification of PLRV and BWYV in 165 potato leafroll disease samples. All of the samples tested negative for BWYV with each of the assay procedures. PLRV was detected in all of the samples with TAS-ELISA, NASH with a cloned cDNA probe for PLRV, and with aphid transmission to ground cherry (Physalis pubescens). Both DAS-ELISA and NASH using random primed cDNA produced one false-negative result. Shepherd's purse (Capsella bursa-pastoris) was a host for 72% (119/165) of the PLRV isolates. The susceptibility of potato to BWYV was tested by inoculating Russet Burbank with three isolates of BWYV from Canada and four from the United States. Two of the isolates were in a mixed infection with PLRV. None of the isolates were transmitted by Myzus persicae to virus-free potato plants, either by themselves or in association with PLRV. Common weeds were surveyed in the potato-producing areas of British Columbia for PLRV and BWYV. In total, 10,098 weed samples, representing 98 species in 22 plant families, were collected and tested by TAS-ELISA from 1986 to 1989. BWYV was detected in 1% of the samples; the hosts were: chickweed, common groundsel, heart-podded hoary cress, hedge mustard, little western bittercress, prickly lettuce, shepherd's purse, and wild radish. PLRV was detected in three volunteer potato plants, two samples of shepherd's purse, and one black nightshade plant. The low incidence of PLRV in plants other than potato indicates that weeds are of minor importance in the epidemiology of potato leafroll disease in British Columbia. / Land and Food Systems, Faculty of / Graduate