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1
Brown, J. K., D. Goldstein, and M. R. Nelson. "Plant Virus Diseases that Threaten Cucurbit Production in Arizona." College of Agriculture, University of Arizona (Tucson, AZ), 1986. http://hdl.handle.net/10150/214137.
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A number of plant viruses were isolated from diseased cucurbits in Arizona during 1982 -85. Watermelon mosiac virus 2, cucumber mosaic virus, and squash mosaic virus are previously recognized viral pathogens in Arizona and in most years are not considered economically threatening to cucurbit production. Three newly described plant viruses (lettuce infectious yellows, watermelon curly mottle and zucchini yellow mosaic) however, have the potential to, or already have, incited serious epidemics in Arizona. Losses are heaviest with these diseases when insect vector levels build up early in the growing season and plants become infected during critical developmental stages.
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Jeffries, Alex Craig. "The study at the molecular level of the New Zealand isolate of Lucerne transient streak sobemovirus and its satellite RNA." Title page, contents and summary only, 1993. http://web4.library.adelaide.edu.au/theses/09PH/09phj47.pdf.
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Király, Lóránt. "Interactions between cauliflower mosaic virus isolates and nicotiana species that determine systemic necrosis /." free to MU campus, to others for purchase, 1997. http://wwwlib.umi.com/cr/mo/fullcit?p9841160.
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Cole, Anthony Blaine Thomas. "Investigations into the hypersensitive response of Nicotiana species to virus infections /." free to MU campus, to others for purchase, 2001. http://wwwlib.umi.com/cr/mo/fullcit?p3012960.
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Balcı, Evrim Doğanlar Sami. "Genetic characterization of cucumber mosaic virus(CMV)resistance in tomato and pepper." [s.l.]: [s.n.], 2005. http://library.iyte.edu.tr/tezler/master/biyoloji/T000388.pdf.
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Yu, Weichang. "CAMV gene VI protein : a virulence factor and the host responses in Arabidopsis /." free to MU campus, to others for purchase, 2002. http://wwwlib.umi.com/cr/mo/fullcit?p3075411.
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Torok, Valeria Anna. "Biological and molecular variation among isolates of pea seed borne mosaic virus." Title page, contents and abstract only, 2001. http://web4.library.adelaide.edu.au/theses/09PH/09pht686.pdf.
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Corrigendum inserted at the back. Includes bibliographical references (leaves 133-158). Ch. 1. General introduction -- ch. 2. General materials and methods -- ch. 3. Biological characterisation of Australian PSbMV isolates -- ch. 4. Developing nucleic acid based diagnostics for PSbMV -- ch. 5. Detection of PSbMV isolates by RT-PCR and RFLP analysis -- ch. 6. Developing an internal control for PSbMV RT-PCR -- ch. 7. Molecular analysis of the PSbMV VPG -- ch. 8. PSbMV sequence and phylogenetic analysis -- ch. 9. General discussion Sixteen pea seed borne mosaic virus (PSbMV) isolates were collected between 1995 and 1998. These isolates were biologically distinct yet serologically indistinguishable. The conclusion is that PSbMV is widespread and occurs at a low incidence in Australia. Reports sequence information on new isolates of PSbMV which has allowed genomic regions to be identified which distinguish PSbMV pathotypes and isolates; and, to the development of PSbMV nucleic acid hybridisation and RT-PCR assays.
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PANNO, Stefano. "IDENTIFICATION OF THE MAIN DESTRUCTIVE PLANT-VIRUS-DISEASES OF HORTICULTURAL CROPS IN SICILY AND DEVELOPMENT OF NEW DIAGNOSTIC TECHNIQUES." Doctoral thesis, Università degli Studi di Palermo, 2014. http://hdl.handle.net/10447/91005.
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Chauhan, Ramola. "A study of filamentous viruses in maize and smallgrains." Master's thesis, University of Cape Town, 1985. http://hdl.handle.net/11427/22013.
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Bibliography: pages 175-184.The occurrence of maize dwarf mosaic virus (MDMV) in field grown maize was investigated. For this purpose, maize showing mosiac symptoms was collected from different maize growing areas in South Africa by Prof. M.B. von Wechmar. These samples from Transvaal, Orange Free State and Natal were then investigated for the presence of MDMV and possible strains of this virus. Three virus isolates were purified and partially characterised. These isolates were serologically compared together with a fourth isolate SCMV 4975, obtained from the U.S., to establish strain relationships.
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Rechcigl, Nancy A. "Ultrastructural cytology of peanut infected with peanut stripe virus." Thesis, Virginia Polytechnic Institute and State University, 1986. http://hdl.handle.net/10919/91063.
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Two isolates of peanut stripe virus (PStV), stripe and blotch, were compared ultrastructurally in peanut (Arachis hypogaea L. 'Florigiant') at several stages of leaf expansion. Ultrathin sections of young leaves infected with either isolate of PStV revealed pinwheel inclusions attached to the cell wall near plasmodesmata. The cytoplasm of infected cells were highly vesiculated. Virus particles amassed in crystalline arrays were observed in blotch infected cells. Virus particles were observed along the arms of pinwheel inclusions. Scroll inclusions appeared in PStV infected cells at a later stage of leaf expansion. In more mature leaves, pinwheel and scroll inclusions occurred in the cytoplasm in association with mitochondria. Virus particles were observed free in the cytoplasm as well as concentrated in linear arrays along the inner surface of the tonoplast. Membrane and organelle degradation was evident in cells infected with either isolate of the virus. Numerous cytoplasmic inclusions and virus particles were observed in cells from light green areas of the leaf. Cells from dark green areas did not contain cytoplasmic inclusions and contained few if any virus particles. Particle measurements show stripe and blotch isolates to have a mean length of 753 nm and 747 nm for leaf dip preparations and 746 nm and 745 nm for partially purified preparations, respectively. Both isolates had a modal length of 750 nm, regardless of the extraction procedure.
The relative virus titer of each isolate was determined in peanut leaves at five stages of leaf expansion and in dark green and light green areas of infected leaves. Virus titer increased significantly from the closed to the fully expanded stage, at which time the virus titer peaked and then decreased slightly. Virus titer was consistently higher in leaves infected with the blotch isolate at all expansion stages. Virus titer was also higher in cells from light green areas of the leaf than from dark green areas of the leaf, regardless of isolate.M.S.
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Maino, Macquin Kilagi. "The development of a serological-based diagnostic test for Dasheen mosaic potyvirus (DsMV)." Thesis, Queensland University of Technology, 2003. https://eprints.qut.edu.au/37139/7/37139_Digitised%20Thesis.pdf.
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Dasheen mosaic potyvirus (DsMV) is an important virus affecting taro. The virus has been found wherever taro is grown and infects both the edible and ornamental aroids,
causing yield losses of up to 60%. The presence of DsMV, and other viruses,prevents the international movement of taro germplasm between countries. This has
a significant negative impact on taro production in many countries due to the inability to access improved taro lines produced in breeding programs. To overcome
this problem, sensitive and reliable virus diagnostic tests need to be developed to enable the indexing of taro germplasm. The aim of this study was to generate an
antiserum against a recombinant DsMV coat protein (CP) and to develop a serological-based diagnostic test that would detect Pacific Island isolates of the virus.
The CP-coding region of 16 DsMV isolates from Papua New Guinea, Samoa, Solomon Islands, French Polynesia, New Caledonia and Vietnam were amplified,cloned and sequenced. The size of the CP-coding region ranged from 939 to 1038
nucleotides and encoded putative proteins ranged from 313 to 346 amino acids, with the molecular mass ranging from 34 to 38 kDa. Analysis ofthe amino acid sequences
revealed the presence of several amino acid motifs typically found in potyviruses,including DAG, WCIE/DN, RQ and AFDF. When the amino acid sequences were
compared with each other and the DsMV sequences on the database, the maximum variability was21.9%. When the core region ofthe CP was analysed, the maximum variability dropped to 6% indicating most variability was present in the N terminus. Within seven PNG isolates ofDsMV, the maximum variability was 16.9% and 3.9% over the entire CP-coding region and core region, respectively. The sequence ofPNG isolate P1 was most similar to all other sequences. Phylogenetic analysis indicated that almost all isolates grouped according to their provenance. Further, the seven
PNG isolates were grouped according to the region within PNG from which they were obtained.
Due to the extensive variability over the entire CP-coding region, the core region ofthe CP ofPNG isolate Pl was cloned into a protein expression vector and expressed as a recombinant protein. The protein was purified by chromatography and SDS-PAGE and used as an antigen to generate antiserum in a rabbit. In western blots, the antiserum reacted with bands of approximately 45-47 kDa in extracts from purified DsMV and from known DsMV -infected plants from PNG; no bands were observed using healthy plant extracts. The antiserum was subsequently incorporated into an indirect ELISA. This procedure was found to be very sensitive and detected DsMV in sap diluted at least 1:1,000. Using both western blot and ELISA formats,the antiserum was able to detect a wide range ofDsMV isolates including those from Australia, New Zealand, Fiji, French Polynesia, New Caledonia, Papua New Guinea, Samoa, Solomon Islands and Vanuatu. These plants were verified to be infected with DsMV by RT-PCR. In specificity tests, the antiserum was also found to react with sap from plants infected with SCMV, PRSV-P, PRSV-W, but not with PVY or CMV -infected plants.
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Maree, H. J. (Hans Jacob). "The expression of Dianthin 30, a ribosome inactivating protein." Thesis, Stellenbosch : Stellenbosch University, 2003. http://hdl.handle.net/10019.1/53633.
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Thesis (MSc)--Stellenbosch University, 2003.ENGLISH ABSTRACT: Ribosome inactivating proteins (RIPs) are currently classified as rRNA N-glycosidases, but also have polynucleotide: adenosine glycosidase activity. RIPs are believed to have anti-viral and anti-fungal properties, but the exact mechanism of these proteins still need to be elucidated.The mechanism of resistance however, appears to be independent of the pathogen. For resistance the RIP terminates virus infected plant cells and stops the reproduction and spread of the virus. Transgenic plants containing RIPs should thus be resistant to a wide range of viruses. The ultimate goal of the larger project of which this forms part is the development of virus resistant plants. To monitor the expression of a RIP in a transgenic plant a detection method had to be developed. Antibody detection of the RIP was decided upon as the most cost effective method. The RIP, Dianthin 30 from Dianthus caryophyllus (carnation), was used and expressed in bacterial and insect expression systems. The bacterial expression experiments were done using the pET expression system in BL21(DE3)pLysS cells. The expression in this system yielded recombinant protein at a very low concentration. Expression experiments were also performed in insect tissue culture with the baculovirus vector BAC-TO-BAC™.With this system the expression was also too low to be used for the production of antibodies. A Dianthin 30 specific peptide was then designed and then produced by Bio-Synthesis. This peptide was then used to raise antibodies to detect Dianthin 30. These antibodies were tested on Dianthus caryophyllus proteins. To establish if this detection method was effective to monitor the expression in plants, tobacco plants were transformed with Agrobacterium tumefaciens containing Dianthin 30 in the pART27 plant expression vector. The putative transformed plants were analysed with peR and Southern blots.
AFRIKAANSE OPSOMMING: Tans word Ribosomale-inaktiverende proteïene (RIPs) geklassifiseer as rRNA N-glikosidase wat ook polinukleotied: adenosien glikosidase aktiwiteit bevat. Daar word geglo dat RIPs anti-virale en anti-fungus eienskappe bevat, maar die meganisme van beskerming word nog nie ten volle verstaan nie. Dit is wel bewys dat die meganisme van weerstand onafhanklik is van die patogeen. Virus geinfekteerde plantselle word deur die RIP gedood om die voortplanting en verspreiding te bekamp en sodoende word weerstand bewerkstellig. Transgeniese plante wat dan 'n RIP bevat sal dus weerstandbiedend wees teen 'n wye spektrum virusse. Die hoofdoel van die breër projek, waarvan die projek deel uitmaak: is die ontwikkeling van virusbestande plante. Om die uitdrukking van die RIP in die transgeniese plante te kontroleer, moes 'n deteksie metode ontwikkel word. Die mees koste effektiewe deteksie metode is met teenliggame. Die RIP, Dianthin 30 from Dianthus caryophyllus (angelier) was gebruik vir uitdrukking in bakteriele- en insekweefselkultuur. Die bakteriele uitdrukkingseksperimente was gedoen met die pET uitdrukkings sisteem III BL21(DE3)pLysS selle. Die uitdrukking in die sisteem het slegs rekombinante proteïene gelewer in uiters lae konsentrasies. Uitdrukkingseksperimente was ook gedoen in insekweefselkultuur met die baculovirus vektor BAC-To- BACTM. Met die sisteem was die uitdrukking ook veels te laag om bruikbaar te wees vir die produksie van teenliggame. Daar is toe 'n peptied ontwerp wat Dianthin 30 kan verteenwoordig vir die produksie van teenliggame. Die teenliggame is getoets teen Dianthus caryophyllus proteïene. Om vas te stel of die deteksiemetode wel die uitdrukking van Dianthin 30 sal kan monitor, is tabak ook getransformeer met Dianthin 30. Die transformasies is gedoen met die hulp van Agrobacterium tumefaciens en die pART27 plant uitdrukkings vektor. Die plante is getoets met die polimerase ketting reaksie en Southern klad tegnieke.
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Malan, Stefanie. "Real time PCR as a versatile tool for virus detection and transgenic plant analysis." Thesis, Stellenbosch : University of Stellenbosch, 2009. http://hdl.handle.net/10019.1/1921.
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Thesis (MSc (Genetics))--University of Stellenbosch, 2009.ENGLISH ABSTRACT: South Africa is regarded as one of the top wine producing countries in the world. One of the threats to the sustainability of the wine industry is viral diseases of which Grapevine leafroll-associated virus 3 (GLRaV-3) and Grapevine virus A (GVA) are considered to be the most important and wide spread. Scion material is regularly tested for viruses; however scion material is often grafted onto rootstocks that have questionable phytosanitary status. Virus detection in rootstocks is challenging due to low and varying titres, but is imperative as a viral control mechanism. An additional viral control mechanism is the use of transgenic grapevine material which offers resistance to grapevine infection. The objective of this project was to establish a detection system using real time PCR (qPCR) techniques, to accurately and routinely detect GLRaV-3 and GVA in rootstock propagation material. qPCR would furthermore be used to perform molecular characterisation of transgenic plants containing a GLRaV-3 antiviral ΔHSP-Mut construct. A severely infected vineyard (Nietvoorbij farm) in the Stellenbosch area was screened throughout the grapevine growing season to investigate virus prevalence throughout the season and to determine the optimal time for sensitive virus detection. A large scale screening of nursery propagation material for GLRaV-3 infection was also conducted. The qRT-PCR results were compared to DAS-ELISA results to compare the efficacy and sensitivity of the two techniques. For the severely infected vineyard, the ability to detect GLRaV-3 increased as the season progressed towards winter. qRT-PCR was more sensitive and accurate in detecting GLRaV-3 than DASELISA, as the latter technique delivered numerous false positive results later in the season. The best time to screen for GLRaV-3 in the Western Cape region was from the end of July to September. For the nursery screenings, our qRT-PCR results were compared to the results of the DAS-ELISA performed by the specific nurseries. No GLRaV-3 infection was detected in the specific samples received from the two different nurseries. The results for all the samples correlated between the two techniques. This confirms that the propagation material of these nurseries has a healthy phytosanitary status with regards to GLRaV-3. However, the detection of GVA in the severely infected vineyard yielded inconsistent results. Detection ability fluctuated throughout the season and no specific trend in seasonal variation and virus titre fluctuation could be established. The highest percentage of GVA infected samples were detected during September, April and the end of July. Previously published universal primers were used for the detection of GVA, but further investigation indicated that they might not be suitable for sensitive detection of specific GVA variants present in South Africa. Vitis vinifera was transformed with a GLRaV-3 antiviral construct, ΔHSP-Mut. SYBR Green Real time PCR (qPCR) and qRT-PCR were utilised as alternative methods for molecular characterisation of transgenic plants. The qPCR and Southern blot results correlated for 76.5% of the samples. This illustrated the ability of qPCR to accurately estimate transgene copy numbers. Various samples were identified during qRT-PCR amplification that exhibited high mRNA expression levels of the transgene. These samples are ideal for further viral resistance studies. This study illustrated that the versatility of real time PCR renders it a valuable tool for accurate virus detection as well as copy number determination.
AFRIKAANSE OPSOMMING: Suid Afrika word geag as een van die top wyn produserende lande ter wereld. Die volhoubaarheid van die wynbedryf word onder andere bedreig deur virus-infeksies. Grapevine leafroll associated virus 3 (GLRaV-3) en Grapevine virus A (GVA) is van die mees belangrike virusse wat siektes veroorsaak in Suid-Afrikaanse wingerde. Wingerd bo-stok materiaal word gereeld getoets vir hierdie virusse, maar hierdie materiaal word meestal geënt op onderstokmateriaal waarvan die virus status onbekend is. Virus opsporing in onderstokke word egter gekompliseer deur baie lae en variërende virus konsentrasies, maar opsporing in voortplantingsmateriaal is ‘n noodsaaklike beheermeganisme vir virus-infeksie. Die doel van die projek was om ‘n opsporingsisteem te ontwikkel via kwantitatiewe PCR (qPCR) tegnieke vir akkurate en gereelde toetsing van GLRaV-3 en GVA in onderstokmateriaal. qPCR sal ook verder gebruik word vir molekulêre karakterisering van transgeniese plante wat ‘n GLRaV-3 antivirale ΔHSP-Mut konstruk bevat. ‘n Hoogs geïnfekteerde wingerd was regdeur die seisoen getoets om seisoenale fluktuasies in viruskonsentrasie te ondersoek en om die optimale tydstip vir sensitiewe virus opsporing te bepaal. ‘n Grootskaalse toetsing van kwekery voortplantingsmateriaal vir GLRaV-3 infeksie was ook uitgevoer. Die qRT-PCR resultate is met die DAS-ELISA resultate vergelyk om die effektiwiteit en sensitiwiteit van die twee tegnieke te vergelyk. Vir die hoogs geïnfekteerde wingerd het die GLRaV-3 opsporing toegeneem met die verloop van die seisoen tot en met winter. qRT-PCR was meer sensitief en akkuraat as DAS-ELISA in die opsporing van GLRaV-3, weens verskeie vals positiewe resultate wat later in die seisoen deur die laasgenoemde tegniek verkry is. Die beste tyd om vir GLRaV-3 te toets is vanaf einde Julie tot September. Tydens die kwekery toetsings was qRT-PCR resultate met die DAS-ELISA resultate van die spesifieke kwekerye vergelyk. Geen GLRaV-3 infeksie was waargeneem in die spesifieke monsters wat vanaf die kwekerye ontvang is nie. Die resultate van die twee tegnieke het ooreengestem vir al die monsters wat v getoets is. Dit het bevestig dat die voortplantingsmateriaal van hierdie kwekerye gesonde fitosanitêre status met betrekking tot GLRaV-3 gehad het. Die opsporing van GVA in die geïnfekteerde wingerd het egter wisselvallige resultate gelewer. Opsporing van die virus het ook regdeur die seisoen gefluktueer en geen spesifieke neiging in seisoenale opsporingsvermoë kon gemaak word nie. Die hoogste persentasie GVA geïnfekteerde monsters was waargeneem tydens September, April en die einde van Julie. Voorheen gepubliseerde universele inleiers was gebruik vir die opsporing van GVA, maar verdere ondersoeke het getoon dat hierdie inleiers nie noodwendig geskik is vir sensitiewe opsporing van GVA variante wat teenwoordig is in Suid-Afrika nie. Vitis vinifera was getransformeer met ‘n GLRaV-3 antivirale konstruct, ΔHSP-Mut. SYBR Green Real time PCR (qPCR) en qRT-PCR was ingespan as alternatiewe metodes vir molekulêre karaterisering van transgeniese plante. Die qPCR en Southern-klad resultate het ooreengestem vir 76.5% van die monsters. Dit illustreer die vermoë van qPCR om akkurate kopie-getalle van transgene te bepaal. Verskeie plante is geïdentifiseer tydens qRT-PCR amplifisering wat hoë vlakke van transgeen mRNA uitdrukking getoon het. Hierdie monsters is ideaal vir verdere virus weerstandbiedendheids studies. Hierdie studie het die veelsydigheid van real time PCR bewys en getoon dat dit ‘n kosbare tegniek is vir akkurate virus opsporing sowel as kopie-getal bepaling.
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Mkhize, Thokozani M. "The detection of cherry leaf-roll nepovirus and the use of molecular markers for germplasm identification in walnuts (Juglans regia L.)." Thesis, Stellenbosch : Stellenbosch University, 2003. http://hdl.handle.net/10019.1/53624.
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Thesis (MSc)--Stellenbosch University, 2003.ENGLISH ABSTRACT: The aim of this study was to combine two common diagnostic tools: serological kits and genetic fingerprinting to identify cherry leaf-roll nepovirus (CLRV), and to establish a marker system to characterize walnut germplasm. The detection of plant viruses is difficult. Restrictions are imposed for quarantine purposes on the importation of plant material from foreign countries. Modern techniques such as a PCR based screening method for CLRV are required to ensure material do not harbour viruses. A primer pair was designed to amplify a 430 bp non-coding homologous region. For the choice of primers, consensus sequences were considered and areas where the sequence data shared 98.5% homology, were chosen. The sensitivity of this detection method was 100-fold higher when compared to the ELISA. The PCR fragment was verified by nucleotide sequencing. AFLP technology was used to identify polymorphic fragments for 6 walnut cultivars and a rootstock, and SCARs were developed from AFLP specific bands. The AFLP technique distinguished all the walnut cultivars and the rootstock. However, conversion of AFLP fragments to SCAR markers for the development of a simple robust technique for cultivar discrimination, was not successful. Using 27 AFLP primer combinations, polymorphic fragments as high as 47.8% were scored. The reason for the lack of efficient conversion was as the result of the AFLP technique. The SCAR primers were generated from sequences internal to the AFLP primers but the specificity of the markers was in the AFLP primers not the internal sequence. In this study using AFLP, walnut cultivars were found to be closely related. The AFLP primer pairs used, provided polymorphic fragments. From these fragments, 7 SCAR markers were developed. It was expected that these SCARs derived from the AFLP markers would detect slight differences between cultivars. The Paradox SCAR marker was the only one that could divide the cultivars into two groups. When Chandler SCAR products were digested with the restriction enzyme Rsal, the same banding pattern as that of Paradox SCAR products was observed.
AFRIKAANSE OPSOMMING: Die doel van hierdie studie was om twee algemene opsporingstegnieke te kombineer: serologiese toetsstelle en genetiese vingerafdrukke om cherry leaf-roll nepovirus (CLRV) te eien en om In merkersisteem te ontwikkel wat okkerneut kiemplasma kan karakteriseer. Die opsporing van plant virusse is baie moeilik. As gevolg van kwarantyn vereistes, word daar beperkinge geplaas word op die invoer van plant materiaal vanuit die buiteland. Moderne tegnieke soos hierdie een wat op PKR berus, word benodig om te verseker dat CLRV nie in plantmateriaal teenwoordig is nie. In Stel inleiers is ontwerp wat In 430 bp nie-koderende homoloë area amplifiseer. Hiervoor is konsensus volgordes bestudeer en slegs die volgordes wat 98,5% homologie getoon het, is gekies. In vergelyking met ELISA was die sensitiwiteit van hierdie deteksie metode 100 maal beter. DNA volgordebepaling is op die resulterende fragment gedoen om die PKR produk te verifieer. AFLP tegnologie is gebruik om polimorfiese fraqmente vir 6 okkerneut kultivars en 'n onderstok te identifiseer en SCARs is uit hierdie fragmente ontwikkel. Die AFLP tegniek kon tussen al die okkerneut kultivars en die onderstok onderskei. Die omskakeling van die AFLP fragmente in SCAR merkers om sodoende In eenvoudige kragtige tegniek vir kultivar onderskeiding te ontwikkel, was egter nie suksesvol nie. Met die gebruik van 27 AFLP inleier kombinasies, kon polimorfiese fragmente van so hoog as 47.8% verkry word. Die rede hoekom omskakeling onsuksesvol was lê by die aard van die AFLP tegniek. Die SCAR inleiers is ontwikkel uit volyordes intern tot die AFLP inleiers, maar die spesifisiteit van die merkers het juis in die AFLP inleiers gelê en nie in die interne volgordes nie. In hierdie studie, met die gebruik van AFLP, is gevind dat okkerneut kultivars baie naby verwant is. Die AFLP inleierstelle wat gebruik is, het polimorfiese fragmente gelewer. Uit hierdie fragmente is 7 SCAR merkers ontwikkel. Daar is verwag dat die SCARs wat uit die AFLP merkers ontwikkel is, klein verskille tussen kultivars sou opspoor. Dit was egter net die Paradox SCAR merker wat die kultivars in twee groepe kon verdeel. Restriksie ensiem vertering met Rsalop die Chandler SCAR produkte het dieselfde bandpatrone as die van die Paradox SCAR produkte gelewer.
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Sassi, Giovanna. "Relative quantification of host gene expression and protein accumulation upon turnip mosaic potyvirus infection in tobacco." Thesis, McGill University, 2004. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=81433.
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Turnip mosaic virus (TuMV) infects a variety of crops, worldwide, including the economically relevant Brassicacea family. It was previously demonstrated that TuMV infection in tobacco protoplasts leads to an overall decrease of host protein. However, it remains unclear whether this phenomenon is due to the repression of plant gene transcription during the infection period or due to viral inhibition of host translation. In this study, quantification of various transcripts and protein products from infected tobacco was performed via real-time RT-PCR and ELISA. In comparison to the gamma-tubulin endogenous control, gene expression for the tobacco H3, HSP70 and granule-bound starch synthase was affected by TuMV infection with time.Tobacco protein accumulation in whole leaf tissues was also significantly affected by increase of virus particles.
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Srinivasan, Indira. "Isolation and detection of bean yellow mosaic, clover yellow vein and peanut stunt viruses from Trifolium L. species." Thesis, This resource online, 1992. http://scholar.lib.vt.edu/theses/available/etd-09122009-040402/.
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Shih, Sharon Min-Hsuan Biotechnology & Biomolecular Sciences Faculty of Science UNSW. "Transient viral infection of plant tissue culture and plants for production of virus and foreign protein." Awarded by:University of New South Wales, 2007. http://handle.unsw.edu.au/1959.4/34967.
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This work was aimed to investigate the basic viral infection protocols mainly focusing on Nicotiana benthamiana hairy root cultures and wild-type tobacco mosaic virus (TMV). The application of transgenic virus containing the gene for green fluorescent protein (GFP) for foreign protein production in plant tissue cultures and whole plants was also studied. The effect on viral accumulation of the form of plant tissue culture used, such as hairy roots, shooty teratomas and suspended cells, was investigated. Viral infection was shown to have no effect on culture growth and morphology. Hairy root cultures are a superior host for viral propagation and production in vitro. The maximum specific rate of viral accumulation occurred mainly during the root growth phase. The average maximum virus concentration in the hairy roots was 0.82 ?? 0.14 mg g-1 dry weight and virus protein represented a maximum of approximately 6% of total soluble protein in the root biomass. Proportional scale-up of TMVinfected hairy roots in shake flasks and bioreactors can be achieved without changing the average virus concentration accumulated in the hairy roots. The level of viral accumulation was much lower in N. benthamiana hairy roots infected with transgenic virus containing GFP (TMVGFPC3) compared with TMV and low levels or no GFP was detected. Viral accumulation and GFP production in whole plants was studied using different generations of transgenic TMV-GFPC3 virus. Hybrid viruses with the foreign gene GFPC3 deleted may have been formed in successive TMV-GFPC3 generations, resulting in the loss of GFP production and enhanced viral infectivity. In vitro generated RNA transcript and first generation TMV-GFPC3 were found to be more suitable for infection than the second generation TMV-GFPC3. However, the accumulation of GFP and virus concentration did not occur at the same ratio. Provided a more genetically stable transgenic viral vector is used for infection, transient viral infection of hairy roots can be a potential alternative system for foreign protein production than plants grown in the field as the containment or safety issues can be addressed.
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Lennefors, Britt-Louise. "Molecular breeding for resistance to rhizomania in sugar beets /." Uppsala : Department of Plant Biology and Forest Genetics, Swedish University of Agricultural Sciences, 2006. http://epsilon.slu.se/2006106.pdf.
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Narita, Nobuyoshi 1961. "Epidemiologia do "Cowpea aphid borne mosaic virus" (CABMV) em maracujazeiros na região produtora da Alta Paulista, SP /." Botucatu : [s.n.], 2007. http://hdl.handle.net/11449/103210.
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Resumo: Dos vírus que infectam o maracujazeiro no Brasil, atualmente o Cowpea aphid borne mosaic virus (CABMV), é considerado fator limitante à cultura. Dependendo da velocidade de disseminação e idade com que as plantas são infectadas no campo, a cultura torna-se comercialmente improdutiva. O presente estudo teve como objetivo, avaliar a diversidade e a dinâmica populacional dos afídeos na região da Alta Paulista, SP e a possibilidade de transmissão do vírus pela semente. Assim, quatro locais (Leste e Oeste da cidade de Marília e Municípios de Ocauçú e Guaimbê) foram monitorados durante 24 meses com armadilhas amarelas de água do tipo Moericke. Constatou-se nas quatro regiões a predominância do gênero Aphis. Outras espécies coletadas foram Myzus persicae, Geopenphigus flocculosus, Brevicoryne brassicae, Rhopalosiphum spp, Dysaphis spp e Lipaphis erysimi. A flutuação populacional de formas aladas do gênero Aphis, caracterizou-se por apresentar maiores revoadas em maio, junho, agosto e setembro. As espécies de Aphis (A. fabae, A. gossypii, A. spiraecola) devem ser os principais vetores do CABMV na região. Plantios novos, ao lado de plantações infectadas, tornam-se infectadas em três meses. Nos testes de transmissão através de sementes, do total de 13056 semeadas oriundas de plantas doentes, germinaram 10592, e em avaliações visuais dois meses após a germinação, não foram observadas plantas sintomáticas, indicando a não transmissão pela semente.Abstract: From the viruses were described infecting passionfruit plants in Brasil, and the Cowpea aphid borne mosaic virus (CABMV), is considered the most hazardous. Depending on the spread velocity of aphids and the age that the plants are infected, the crops doesnt produce commercial fruits. The present study was designed to evaluate the diversity and dynamic population of aphids in the Alta Paulista, SP region and aspects of seed transmission. For this, four regions (East and West of Marília city, Guaimbê and Ocauçú) were monitored for 24 months using yellow water Moerick trap. The predominance of the genus Aphis was observed in the four evaluated areas. Other species founded in the area were: Myzus persicae, Geopenphigus flocculosus, Brevicoryne brassicae, Rhopalosiphum spp, Dysaphis spp and Lipaphis erysimi. The population curve of alate Aphis spp showed the highest frequency of flights during May, June, August and September. The Aphis spp (A. fabae, A. gossypii, A. spiraecola) probably is the most important vector of the CABMV in the region. New crops near old infected plants, were infected in three months. To evaluate properties of seed transmission, from 13056 collected from infected plants, 10592 were germinated and evaluated during two months for the presence of visual symptoms. No plants with simptoms were observed indicating no seed transmission.
Orientador: Marcelo Agenor Pavan
Coorientador: Valdir Atsushi Yuki
Banca: Renate Krause Sakate
Banca: Aloisio Costa Sampaio
Banca: Alexandre Levi R. Chaves
Banca: Hugo Kuniyuki
Doutor
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Beltrame, André Boldrin. "Efeito de cianobactérias e algas eucarióticas na resistência de plantas de fumo contra o Tobacco mosaic virus (TMV)." Universidade de São Paulo, 2006. http://www.teses.usp.br/teses/disponiveis/11/11135/tde-02032006-155032/.
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As algas produzem uma grande diversidade de compostos com atividade biológica, inclusive que agem diretamente sobre vírus ou como indutores de fitoalexinas. Em vista disso, foi investigada a redução de sintomas causados por Tobacco mosaic vírus (TMV) em plantas de fumo tratadas com cianobactérias ou algas eucarióticas, além de se tentar elucidar o modo de ação das algas no patossistema estudado. Quando as plantas de fumo foram tratadas dois dias antes da inoculação, foi verificado que suspensões dos isolados 004/02, 008/02, 061/02, Anabaena sp. e Nostoc sp. 61, bem como as preparações do conteúdo intracelular do isolado 004/02 (4 C) e do filtrado do meio de cultivo do isolado 061/02 (61 M) apresentaram efeito na redução dos sintomas de TMV em plantas de fumo, cultivar TNN. Além disso, foi estudado o efeito direto das algas sobre as partículas de vírus. Os resultados mostraram que os isolados Anabaena sp., Nostoc sp. 21, Nostoc sp. 61 e 090/02 apresentam compostos que agem diretamente sobre o TMV. Para tentar elucidar o mecanismo de ação das algas no patossistema estudado, diversos parâmetros bioquímicos foram investigados. Foi detectado que a preparação 4 C aumentou a atividade de peroxidases e que todos os tratamentos analizados reduziram a atividade de β-1,3-glucanase em folhas de fumo a partir do quarto dia após o tratamento das plantas. Por sua vez, as suspensões dos isolados 008/02 e 061/02 e a preparação 61 M proporcionaram maior acúmulo de superóxido, enquanto que a preparação 4 C reduziu o acúmulo de peróxido de hidrogênio, em relação aos controles água destilada e meio de cultura BG 11, 37 horas após a inoculação do vírus. Em vista disso, as algas podem ser utilizadas como agentes de controle biológico, por apresentar ação direta sobre fitopatógenos ou alterarem o metabolismo de plantas, o que pode estar associado com a sintese de compostos de defesa.Algae produce several different compounds that show biological activity, including ones with antiviral activity or that act as phytoalexin inducers. Thus, it was investigated the reduction of Tobacco mosaic virus (TMV) symptoms on tobacco plants treated with cyanobacteria or eukaryotic algae, and it was studied the way of action of algae on the studied pathosystem. When the tobacco plants were treated two days before the inoculation, it was verified that the suspension of 004/02, 008/02, 061/02 Anabaena sp., and Nostoc sp. 61 strains as well as the intracellular preparation of 004/02 strain (4 C) and the medium filtrated from 061/02 strain (61 M) reduced TMV symptoms on tobacco plants, cultivar TNN. Furthermore, it was studied the direct effect of the algae suspensions on virus particles. The results showed that Anabaena sp., Nostoc sp. 21, Nostoc sp. 61 and 090/02 strains have compounds with direct activity on TMV. To try to elucidate the way of the action of algae, on the studied pathosystem, several biochemical parameters were investigated. It was seen that the preparation 4 C increase peroxidase activity and all treatments decrease β-1,3-glucanase activity on tobacco leaves from the forth day on after the treatment. Moreover, 008/02 and 061/02 strains and the 61 M preparation caused higher superoxide accumulation, and the preparation 4 C decreased hydrogen peroxide accumulation when compared to the controls distilled water and BG 11 medium 37 hour after virus inoculation. In this way, the algae could be a biocontrol agents, because it shows direct action on phytopathogens and/or change the metabolism of the plants, that could be associated with the synthesis of deffence compounds.
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Narita, Nobuyoshi [UNESP]. "Epidemiologia do Cowpea aphid borne mosaic virus (CABMV) em maracujazeiros na região produtora da Alta Paulista, SP." Universidade Estadual Paulista (UNESP), 2007. http://hdl.handle.net/11449/103210.
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Dos vírus que infectam o maracujazeiro no Brasil, atualmente o Cowpea aphid borne mosaic virus (CABMV), é considerado fator limitante à cultura. Dependendo da velocidade de disseminação e idade com que as plantas são infectadas no campo, a cultura torna-se comercialmente improdutiva. O presente estudo teve como objetivo, avaliar a diversidade e a dinâmica populacional dos afídeos na região da Alta Paulista, SP e a possibilidade de transmissão do vírus pela semente. Assim, quatro locais (Leste e Oeste da cidade de Marília e Municípios de Ocauçú e Guaimbê) foram monitorados durante 24 meses com armadilhas amarelas de água do tipo Moericke. Constatou-se nas quatro regiões a predominância do gênero Aphis. Outras espécies coletadas foram Myzus persicae, Geopenphigus flocculosus, Brevicoryne brassicae, Rhopalosiphum spp, Dysaphis spp e Lipaphis erysimi. A flutuação populacional de formas aladas do gênero Aphis, caracterizou-se por apresentar maiores revoadas em maio, junho, agosto e setembro. As espécies de Aphis (A. fabae, A. gossypii, A. spiraecola) devem ser os principais vetores do CABMV na região. Plantios novos, ao lado de plantações infectadas, tornam-se infectadas em três meses. Nos testes de transmissão através de sementes, do total de 13056 semeadas oriundas de plantas doentes, germinaram 10592, e em avaliações visuais dois meses após a germinação, não foram observadas plantas sintomáticas, indicando a não transmissão pela semente.
From the viruses were described infecting passionfruit plants in Brasil, and the Cowpea aphid borne mosaic virus (CABMV), is considered the most hazardous. Depending on the spread velocity of aphids and the age that the plants are infected, the crops doesn t produce commercial fruits. The present study was designed to evaluate the diversity and dynamic population of aphids in the Alta Paulista, SP region and aspects of seed transmission. For this, four regions (East and West of Marília city, Guaimbê and Ocauçú) were monitored for 24 months using yellow water Moerick trap. The predominance of the genus Aphis was observed in the four evaluated areas. Other species founded in the area were: Myzus persicae, Geopenphigus flocculosus, Brevicoryne brassicae, Rhopalosiphum spp, Dysaphis spp and Lipaphis erysimi. The population curve of alate Aphis spp showed the highest frequency of flights during May, June, August and September. The Aphis spp (A. fabae, A. gossypii, A. spiraecola) probably is the most important vector of the CABMV in the region. New crops near old infected plants, were infected in three months. To evaluate properties of seed transmission, from 13056 collected from infected plants, 10592 were germinated and evaluated during two months for the presence of visual symptoms. No plants with simptoms were observed indicating no seed transmission.
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Nogueira, Diêgo Rodrigues Soares. "Produção e avaliação de anti-soro policlonal visando a detecção do Pepper yellow mosaic virus." Universidade Federal de Viçosa, 2014. http://locus.ufv.br/handle/123456789/4436.
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Pepper yellow mosaic virus (PepYMV) naturally infects sweet pepper and tomato plants, leading to severe losses since its first report in Brazil (Brasília, DF, 2002). Molecular and serological methods can be used to detect the pathogen. Serological methods for viral detection require the use of a high quality antiserum, offering good sensitivity and specificity. Traditionally, purified viral particles are used as immunogens. However, the purification process is very laborious and the final preparation may have unsatisfactory purity and/or concentration. Thus, the aim of this work was to produce a polyclonal antiserum against the recombinant capsid protein (CP) of PepYMV to allow its use both for diagnosis and for studies of the interaction of the PepYMV CP with other viral proteins and host factors. The coding sequence of the capsid protein gene of PepYMV was cloned into an expression vector (pRSET-A) and transformed into Escherichia coli strain BL21::DE3 for in vitro expression. The recombinant protein, fused to a histidine tag, was purified under denaturing conditions by affinity chromatography using a Ni-NTA column. The purified recombinant protein was dialyzed under renaturing conditions. Its integrity and identity were confirmed by polyacrylamide gel electrophoresis and mass spectrometer analyses. New Zealand rabbits were immunized with increasing amounts of the recombinant protein. The sensitivity and specificity of the antisera were analyzed by Western blot and indirect ELISA assays. The antisera raised against the recombinant CP showed good specificity and sensibility, proving to be a reliable tool for the detection of PepYMV.
O Pepper yellow mosaic virus (PepYMV), agente causal do mosaico amarelo do pimentão e do tomateiro, desde seu primeiro relato no Brasil no ano de 2002 em Brasília, DF, vem se disseminando em regiões produtoras de pimentão e tomate causando perdas substanciais ao produtores dessas culturas. Para identificação dessa enfermidade algumas ferramentas são utilizadas, dentre elas, podemos destacar os métodos moleculares e sorológicos, sendo estes últimos mais utilizados por apresentarem alta especificidade e sensibilidade, além de possuir um custo relativamente baixo. Para a produção de anti-soro, tradicionalmente, utilizam-se partículas virais concentradas como imunógenos. No entanto, o processo de purificação é muito trabalhoso e pode apresentar pureza e concentrações insatisfatórias. Desta forma, o objetivo deste trabalho foi produzir um anti-soro policlonal, a partir da proteína capsidial recombinante do PepYMV, que permita sua utilização tanto para diagnose quanto para estudos de interação da proteína capsidial do PepYMV com outras proteínas virais e fatores do hospedeiro. A sequência do gene da proteína capsidial do PepYMV foi clonada em vetor de expressão (pRSET-A) e transformada em Escherichia coli, linhagem BL21::DE3, para expressão in vitro. A proteína expressa fusionada a uma cauda de histidina foi purificada sob condições desnaturantes por cromatografia de afinidade em coluna de resina Ni-NTA. Em seguida a proteína purificada foi dialisada sob condições renaturantes e sua integridade e identidade foram confirmadas por gel de poliacrilamida a 12% e análise de espectrometria de massa. Dois coelhos da raça Nova Zelândia foram imunizados com quantidades crescentes da proteína recombinante dialisada adicionados do adjuvante de Freud incompleto na proporção 1:1. A sensibilidade e a especificidade do anti-soro foram testados por Western blot e ELISA indireto. O anti-soro produzido apresentou boa especificidade e sensibilidade, provando ser uma ferramenta confiável para a diagnose do PepYMV.
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Northfield, John. "Aspects to T-cell phenotype during infection with HIV, CMV and Hepatitis C virus." Thesis, University of Oxford, 2008. http://ora.ox.ac.uk/objects/uuid:283098ce-e24d-4099-8826-07dcc75381f2.
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This work concerns itself with understanding the organisation of cellular immune responses to three major human pathogens - HIV, CMV and Hepatitis C (HCV). Each was studied to form three projects, each undertaken with a different approach - arrived at independently - and largely owing their origins to opportunity and circumstance as much as design. Each project led to exploration of a particular aspect of T-cell phenotype (that is the expression of particular molecular markers on T-cells) and its’ broader biological significance. I found that T-cell phenotype was strongly linked to the magnitude of T-cell responses (CMV) and the ability of T-cells to control infection (HIV). Finally I explored the significance of expression of a molecule known as CD161 on the surface of HCV specific CD8+ T-cells, indicating a phenotype of T-cell that may not follow the ‘normal rules’ applicable to T-cells in general.
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Santos, Mateus de Almeida. "Tentativas de purificação e produção de antissoro contra o vírus da morte súbita dos citros e isolamento do CSDaV em plantas herbáceas." Universidade de São Paulo, 2011. http://www.teses.usp.br/teses/disponiveis/11/11135/tde-12092011-152557/.
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A morte súbita dos citros (MSC) foi identificada em 2001, no município de Comendador Gomes, Minas Gerais, e desde então foi responsável pela perda de 4 milhões de plantas na região sul do Triângulo Mineiro, e no norte e noroeste do estado de São Paulo. É uma doença de combinação copa/porta-enxerto, afetando principalmente laranjeira doce enxertada em limoeiro Cravo, e que culmina na morte das plantas. Passados dez anos do seu relato, até hoje não se tem conhecimento exato do agente causal e dos possíveis vetores. Sabe-se, todavia, que em todas as plantas com morte súbita encontram-se o vírus da tristeza dos citros (Citrus tristeza virus - CTV) e um vírus do Gênero Marafivirus, Família Tymoviridae, denominado Citrus sudden death associated virus (CSDaV). Devido esse fato, há a necessidade de separá-los para testar os postulados de Koch para o CSDaV. O principal objetivo deste trabalho foi tentar isolar o CSDaV para verificar o seu real envolvimento com a MSC. Também se procurou purificar esse vírus para a produção de antissoro policlonal para trabalhos de diagnose da doença. Para a purificação do CSDaV foi utilizado o protocolo de purificação do Potato leaf roll virus, porém os resultados não foram satisfatórios devido a constante presença do CTV. Tentativas de remoção do CTV por meio de imunoprecipitação com antissoro homólogo não foram satisfatórias. O antissoro produzido reagiu indistintamente com extratos de plantas infectadas com o CSDaV e o CTV. O CSDaV foi transmitido mecanicamente para plantas de Nicotiana sp., N. clevelandii, Chenopodium amaranticolor e C. quinoa, causando principalmente infecção localizada. A presença do vírus foi confirmada por RT-PCR e a sua identidade por meio do sequenciamento de nucleotídeos do produto da amplificação. Tentativas de transmissão do CSDaV usando inóculo de extrato de folhas de plantas do campo , por meio da Cuscuta sp., através de cortes no tronco das plantas com lâmina embebida no inóculo, com pulgões Toxoptera citricida aparentemente virulíferos somente para o tymovirus e por meio da inoculação de sementes de citros deram resultados negativos.Citrus sudden death (CSD) disease was identified in 2001, at Comendador Comes County, State of Minas Gerais, Brazil. Since then the disease has caused the death of 4 million trees in Southwestern Minas Gerais State and Northern São Paulo State. This new and destructive disease affects sweet orange as well as other species, varieties, and hybrids when grafted on Rangpur (Citrus limonia). Then years after the first report on CSD, the causal agent and possible vector(s) have not been precisely identified. It is known, however, that all disease trees are infected with Citrus tristeza virus (CTV) and Citrus sudden death associate virus (CSDaV), which is a member of the Genus Marafivirus, Famíly Tymoviridae. Due to this, it is necessary to separate these pathogens, in order to complete Kochs postulated for the CSDaV. The main purpose of the present work was to try to isolate the CSDaV to verify its role on CSD disease. In addition, attempts were done to purify the virus and produce polyclonal antiserum for disease diagnosis. Purification was carried out as described for Potato leaf roll virus, but results were not suitable due to the constant presence of CTV. Efforts to remove CTV by immunoprecipitation with homologous antiserum did not succeed. The produced antiserum reacted indistinctly with extracts of plants infected with both viruses. SCDaV was mechanically transmitted to Nicotiana sp., N. clevelandii, Chenopodium amaranticolor, and C. quinoa, causing mainly local infection. Infection was confirmed by RT-PCR and virus identity was determined by nucleotide sequence of the amplified fragment. Efforts to transmit CSDaV, using as inoculum extract from field infected plants, by means of Cucucuta sp., by incisions on the trunk of the test-plants, with Toxoptera citricida apparently viruliferous only for the tymovirus, and by means of citrus seed inoculation gave negative results.
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Amaral, Ingrid. "Biologia e tabela de vida de Brevipalpus yothersi (Acari: Tenuipalpidae) oriundos de diferentes regiões citrícolas do Estado de São Paulo /." Jaboticabal, 2016. http://hdl.handle.net/11449/137926.
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Orientador: Daniel Junior de AndradeBanca: Marineide Rosa Vieira
Banca: Renato Beozzo Bassanezi
Resumo: O ácaro Brevipalpus yothersi Baker é vetor da leprose dos citros, principal doença viral da citricultura mundial. Informações sobre a biologia de B. yothersi são essenciais para compreender a dinâmica populacional do ácaro no campo e inferir se mudanças no manejo do pomar em função da região pode alterar a biologia do ácaro. O objetivo do trabalho foi determinar a biologia e elaborar a tabela de vida de fertilidade de B. yothersi coletados em diferentes regiões citrícolas do estado de São Paulo. Os experimentos foram realizados no Laboratório de Acarologia, pertencente à Faculdade de Ciências Agrárias e Veterinárias - FCAV/UNESP, Jaboticabal - SP. Os ácaros foram coletados em pomares cítricos das regiões de Barretos, Jales e Santa Cruz do Rio Pardo, posteriormente, em laboratório, foram multiplicados em frutos de laranja. Os parâmetros biológicos avaliados foram duração das fases de desenvolvimento, oviposição, período de incubação, viabilidade dos ovos, longevidade, taxa líquida de reprodução (Ro), tempo médio de geração (T), taxa intrínseca de crescimento populacional (rm) e taxa finita de crescimento populacional (λ). Estes parâmetros foram avaliados em dois experimentos, o primeiro consistiu na biologia de B. yothersi em frutos isentos de resíduos de produtos fitossanitários à 23±1ºC e o segundo sob frutos com resíduo de espirodiclofeno à 25±1ºC. As observações foram realizadas diariamente, pela manhã e ao fim da tarde. A duração do desenvolvimento, longevidade, período d... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The mite Brevipalpus yothersi Baker is the vector of the citrus leprosis, major viral disease of citrus worldwide. Information about B. yothersi's biology are essential to understanding the population dynamics of the mite in the field and infer whether changes in orchard management by region can change the mite biology. The objective was to determine the biology and prepare the fertility life table of B. yothersi collected in different citrus regions of São Paulo state. The experiments were performed in Acarology Laboratory, belonging to the Faculty of Agricultural and Veterinary Sciences - FCAV/UNESP, Jaboticabal - SP. The mites were collected in citrus orchards in the regions of Barretos, Jales and Santa Cruz do Rio Pardo, later in the laboratory were multiplied in orange fruits. The biological parameters assessed were duration of the stages of development, oviposition, incubation period, egg viability, longevity, net reproductive rate (Ro), mean generation time (T), intrinsic rate of increase (rm) and finite rate increase (λ). These parameters were evaluated in two experiments, the first consisted the biology of B. yothersi in fruits free of residues of pesticides at 23 ± 1°C and the second consisting of the biology of B. yothersi under fruit with spirodiclofen residue at 25 ± 1°C . The observations were performed daily, in the morning and in the afternoon. The duration of the development, longevity, pre-oviposition period, oviposition rate and number of B. yothersi eggs s... (Complete abstract click electronic access below)
Mestre
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Al-Kaff, Nadia Saleh Ahmed. "Biological and molecular diversity of cauliflower mosaic virus." Thesis, University of East Anglia, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.240834.
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Pinto, Yvonne Maria. "Characterization of a Nigerian isolate of rice yellow mottle virus." Thesis, Imperial College London, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.243556.
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Padmini, de Silva D. P. "Studies of black pepper (Piper nigrum L.) virus disease in Sri Lanka." Thesis, University of Reading, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308101.
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Ali, Akhtar. "Pathology and molecular comparison of a range of pea seed-borne mosaic virus isolates." Title page, contents and summary only, 1999. http://web4.library.adelaide.edu.au/theses/09ACP/09acpa398.pdf.
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Copies of author's previously published articles inserted. Bibliography: leaves 128-143. This thesis describes the development of serological and nucleic acid based diagnostic methods for pea-seed borne mosaic virus (PSbMV), the isolation of specific effects on infected pea plants, the collection and biological comparison of new PSbMV isolates from Pakistan, the cloning and sequencing of specific parts of the genome of selected isolates, nucleotide and amino acid sequence comparisons between selected isolates, and the development of a ribonuclease protection assay (RPA) for identifying genomic differences among the PSbMV isolates. It is the first comparison of a range of geographically different isolates of PSbMV on the basis of both biological and molecular properties.
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Saqib, Muhammad. "Studies on new plant phytoplasma and viruses infections and molecular dissection of virus resistance using Medicago truncatula." Thesis, Saqib, Muhammad (2008) Studies on new plant phytoplasma and viruses infections and molecular dissection of virus resistance using Medicago truncatula. PhD thesis, Murdoch University, 2008. https://researchrepository.murdoch.edu.au/id/eprint/288/.
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The work presented in this thesis is in two areas - study of novel pathogens resulting from new encounters between crop and native species and 'mining' for plant virus resistance genes in the model legume Medicago truncatula.
The history of agriculture in Western Australia (WA) is less than 150 years old. All major broadacre and horticultural crops grown in WA have been introduced from overseas. These introduced horticultural and field crops potentially carry pathogens which may be transferred to infect native vegetation. Conversely, cultivated plants are vulnerable to infection by pathogens present in indigenous plants. This potential for new disease encounters is compounded by expansion of agriculture to crop new land and by predicted climate changes. These changes may provide selective advantage to a particular pest or disease, enabling infection to increase and so increase crop losses or damage native species. Global trade in agricultural produce also increases the potential for introduction of exotic pathogens. The focus of the first part of the research was to look for new pathogens of crops and native plants in WA.
A series of field trips to study diseases in horticultural crops and native vegetation were made in the agricultural regions of Carnarvon, Broome, Kununurra, Perth and the surrounding metropolitan area. Although the initial focus was on virus diseases, the work expanded to study phytoplasma-associated diseases, because of their widespread occurrence and clear symptoms.
In the agricultural region around Kununurra the potyvirus Bean common mosaic virus (BCMV) was found infecting Phaseolus vulgaris crops. Sequencing of isolates collected provided the first reliable molecular confirmation of the presence of BCMV in Australia.
In joint work with K. Bayliss three commercial Paulownia tree plantations near Perth were found exhibiting symptoms of Witches'-Broom disease. The Paulownia trees were found to be associated with 'Candidatus Phytoplasma australiense' 16SrXII group. Chickpeas in the Kununurra region were found with symptoms of stunting, little leaf and proliferating branches and tested positive for phytoplasma. Sequencing confirmed the presence of a phytoplasma with high similarity to the 16SrII group 'Ca Phytoplasma aurantifolia' (peanut witches broom group). This is the first molecular evidence for a phytoplasma-associated disease in chickpea. Red clover (Trifolium pratense), several other pasture legumes and paddy melon (Cucumis myriocarpus) with symptoms of diminished leaf size, pallor, rugosity, leaf deformation, shoot proliferation and stunting were observed amongst pasture plots in south-western Australia. All species with these symptoms were positive for a phytoplasma resembling 'Ca Phytoplasma australiense, 16SrXII group. This association was confirmed for red clover and paddy melon by subsequent nested PCR and sequence analysis. This is the first time that 'Ca. Phytoplasma australiense, 16SrXII group, has been reported infecting these hosts in southern WA. Snakebean (Vigna unguiculata var. sesquipedalis) and tomato (Lycopersicon esculentum) plants with phytoplasma-like symptoms were found in the horticultural region at Broome. The symptoms on snakebean were typical of phytoplasma disease. Sequence analysis identified that the agent associated with the symptoms as a strain of sweet potato little leaf strain V4 (SPLL-V4) phytoplasma (16SrXII group, strain of 'Ca Phytoplasma australiense'). SPLL phytoplasma has not been reported in snakebean or tomato in this isolated agricultural region. In a survey in the Gascoyne region phytoplasma-like symptoms were found in tomato, eggplant and papaya. Previously in this region plants had been found to be associated with peanut witches broom phytoplasma 16SrII group 'Ca Phytoplasma aurantifolia'. Phytoplasma-like symptoms which included bunchy growth, witches' broom and 'little leaf' were observed in Allocasuarina fraseriana (Western Sheoak, Casuarina) and Acacia saligna (Acacia, Orange Wattle) trees in Kings Park and Botanic Garden Perth WA. Phytoplasma-associated disease was confirmed for the first time in native Australian casuarina and acacia trees in WA. Based on the identification of these phytoplasma associated diseases in WA, phytoplasma-associated diseases can be divided into two zones, because phytoplasma 16SrII group was found mostly in the north west of WA and the 16SrXII group in the south west of WA. This work has added to knowledge of the extent and distribution of phytoplasma disease in WA: it is concluded that crop-associated phytoplasma disease originated from native vegetation.
The aim of the second part of the research was to screen and map a virus resistance gene in the model legume M. truncatula to better understand host/pathogen interactions of legume-infecting viruses. Natural resistance genes found in M. truncatula could then be used to locate similar genes in grain legumes (e.g. chickpea and lupins) for practical applications. M. truncatula is a model legume which has a relatively small genome. International consortia have been established to develop genomic resources for M. truncatula. The M. truncatula core collection (from SARDI, South Australia) totalling 230 accessions was screened for resistance/susceptibility to four legume-infecting viruses: Alfalfa mosaic virus (AMV), Cucumber mosaic virus (CMV), Bean yellow mosaic virus (BYMV) and Subterranean clover mottle virus (SCMoV). Five plants from each of the 230 phenotypically distinct members of the M. truncatula core collection were challenged with one isolate of each virus using infectious sap together with five uninoculated control plants for each accession. The symptoms that developed were recorded and virus presence was confirmed by ELISA for inoculated and systemic leaves. Accessions that were potentially resistant were retested to check for escapes. The result from this screen was that 5 accessions were potentially resistant to AMV, 56 to BYMV, 21 to CMV and 42 to SCMoV. The remaining accessions were susceptible to all four viruses with symptoms which ranged from no apparent symptoms (symptomless systemic infection) to highly susceptible and plant death. In continuing work with DAFWA (Dr R. Jones) accessions potentially resistant to AMV, BYMV and CMV are being challenged with additional isolates to check for the presence of genes providing broader resistance.
The Sobemovirus SCMoV was chosen for further study because it is the most widespread viral pathogen of subterranean clover pastures in Australia. It is also a high titre, mechanically transmitted virus which gave the least escapes on infection. SCMoV has a linear, single-stranded positive-sense RNA genome of 4.25 Kb. Making use of natural resistance is an effective means to reduce pasture losses caused by SCMoV. From the screen of the core collection of M. truncatula, amongst the lines resistant to SCMoV a single dominant hypersensitive resistance was detected in line DZA-315. To accelerate mapping of the SCMoV resistance gene, an F8 RIL population of a cross between the resistant line (DZA-315) and a susceptible line (Jemalong-J6, A-17) was sourced and obtained from INRA Toulouse. A total of 166 RILs were phenotyped for resistance and susceptibility to SCMoV. Resistant and susceptible lines showed parental phenotypic symptoms with 84 being susceptible and 82 being resistant. This indicated the presence of a single resistance (R) gene. This phenotypic data was combined with genotypic data (76 polymorphic molecular markers) already available for this RIL population to provide a framework map. Mapmaker and Mapmanager mapping programs were used to locate the position of the resistance gene. This framework map indicated a position for the resistance gene on the long arm of chromosome 6.
Additional polymorphic SSR markers flanking the R gene locus on chromosome 6 were used to map the position of the R gene more closely. These SSR markers were developed from a parental cross of M. truncatula line A17 and A20 at UC Davis and from a parental cross between line A17 and DZA 315 developed at INRA Toulouse. Ten new polymorphic SSR markers were identified and located on the long arm of chromosome 6 after analysis of the F8 RIL population. When combined with the other phenotypic and genotypic data a more accurate map position for the SCMoV R gene was obtained. The results indicate that the R gene to SCMoV is located on the long arm of M. truncatula chromosome 6 between position 35 to 38 centimorgans (cM). The closest marker to the SCMoV R gene is marker mtic153 which is about 2.3 cM away. From existing maps of M. truncatula most of the R genes located in this region are of the TIR-NBS-LRR type and occur in R gene clusters. A series of BACs that span the region of interest have been identified in which SCMoV R gene should be present.
M. truncatula has been used as a model legume to study a number of symbiotic (e.g. rhizobium) and pathogenic interactions (e.g. fungal and nematode), but this is the only example of its use to study legume-virus interactions. The results obtained indicate the potential of using M. truncatula as a model to study resistance response to other legume viruses and provide a firm basis for identifying the hypersensitive R gene that confers resistance to SCMoV.
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Saqib, Muhammad. "Studies on new plant phytoplasma and viruses infections and molecular dissection of virus resistance using Medicago truncatula." Saqib, Muhammad (2008) Studies on new plant phytoplasma and viruses infections and molecular dissection of virus resistance using Medicago truncatula. PhD thesis, Murdoch University, 2008. http://researchrepository.murdoch.edu.au/288/.
Full textAbstract:
The work presented in this thesis is in two areas - study of novel pathogens resulting from new encounters between crop and native species and 'mining' for plant virus resistance genes in the model legume Medicago truncatula.
The history of agriculture in Western Australia (WA) is less than 150 years old. All major broadacre and horticultural crops grown in WA have been introduced from overseas. These introduced horticultural and field crops potentially carry pathogens which may be transferred to infect native vegetation. Conversely, cultivated plants are vulnerable to infection by pathogens present in indigenous plants. This potential for new disease encounters is compounded by expansion of agriculture to crop new land and by predicted climate changes. These changes may provide selective advantage to a particular pest or disease, enabling infection to increase and so increase crop losses or damage native species. Global trade in agricultural produce also increases the potential for introduction of exotic pathogens. The focus of the first part of the research was to look for new pathogens of crops and native plants in WA.
A series of field trips to study diseases in horticultural crops and native vegetation were made in the agricultural regions of Carnarvon, Broome, Kununurra, Perth and the surrounding metropolitan area. Although the initial focus was on virus diseases, the work expanded to study phytoplasma-associated diseases, because of their widespread occurrence and clear symptoms.
In the agricultural region around Kununurra the potyvirus Bean common mosaic virus (BCMV) was found infecting Phaseolus vulgaris crops. Sequencing of isolates collected provided the first reliable molecular confirmation of the presence of BCMV in Australia.
In joint work with K. Bayliss three commercial Paulownia tree plantations near Perth were found exhibiting symptoms of Witches'-Broom disease. The Paulownia trees were found to be associated with 'Candidatus Phytoplasma australiense' 16SrXII group. Chickpeas in the Kununurra region were found with symptoms of stunting, little leaf and proliferating branches and tested positive for phytoplasma. Sequencing confirmed the presence of a phytoplasma with high similarity to the 16SrII group 'Ca Phytoplasma aurantifolia' (peanut witches broom group). This is the first molecular evidence for a phytoplasma-associated disease in chickpea. Red clover (Trifolium pratense), several other pasture legumes and paddy melon (Cucumis myriocarpus) with symptoms of diminished leaf size, pallor, rugosity, leaf deformation, shoot proliferation and stunting were observed amongst pasture plots in south-western Australia. All species with these symptoms were positive for a phytoplasma resembling 'Ca Phytoplasma australiense, 16SrXII group. This association was confirmed for red clover and paddy melon by subsequent nested PCR and sequence analysis. This is the first time that 'Ca. Phytoplasma australiense, 16SrXII group, has been reported infecting these hosts in southern WA. Snakebean (Vigna unguiculata var. sesquipedalis) and tomato (Lycopersicon esculentum) plants with phytoplasma-like symptoms were found in the horticultural region at Broome. The symptoms on snakebean were typical of phytoplasma disease. Sequence analysis identified that the agent associated with the symptoms as a strain of sweet potato little leaf strain V4 (SPLL-V4) phytoplasma (16SrXII group, strain of 'Ca Phytoplasma australiense'). SPLL phytoplasma has not been reported in snakebean or tomato in this isolated agricultural region. In a survey in the Gascoyne region phytoplasma-like symptoms were found in tomato, eggplant and papaya. Previously in this region plants had been found to be associated with peanut witches broom phytoplasma 16SrII group 'Ca Phytoplasma aurantifolia'. Phytoplasma-like symptoms which included bunchy growth, witches' broom and 'little leaf' were observed in Allocasuarina fraseriana (Western Sheoak, Casuarina) and Acacia saligna (Acacia, Orange Wattle) trees in Kings Park and Botanic Garden Perth WA. Phytoplasma-associated disease was confirmed for the first time in native Australian casuarina and acacia trees in WA. Based on the identification of these phytoplasma associated diseases in WA, phytoplasma-associated diseases can be divided into two zones, because phytoplasma 16SrII group was found mostly in the north west of WA and the 16SrXII group in the south west of WA. This work has added to knowledge of the extent and distribution of phytoplasma disease in WA: it is concluded that crop-associated phytoplasma disease originated from native vegetation.
The aim of the second part of the research was to screen and map a virus resistance gene in the model legume M. truncatula to better understand host/pathogen interactions of legume-infecting viruses. Natural resistance genes found in M. truncatula could then be used to locate similar genes in grain legumes (e.g. chickpea and lupins) for practical applications. M. truncatula is a model legume which has a relatively small genome. International consortia have been established to develop genomic resources for M. truncatula. The M. truncatula core collection (from SARDI, South Australia) totalling 230 accessions was screened for resistance/susceptibility to four legume-infecting viruses: Alfalfa mosaic virus (AMV), Cucumber mosaic virus (CMV), Bean yellow mosaic virus (BYMV) and Subterranean clover mottle virus (SCMoV). Five plants from each of the 230 phenotypically distinct members of the M. truncatula core collection were challenged with one isolate of each virus using infectious sap together with five uninoculated control plants for each accession. The symptoms that developed were recorded and virus presence was confirmed by ELISA for inoculated and systemic leaves. Accessions that were potentially resistant were retested to check for escapes. The result from this screen was that 5 accessions were potentially resistant to AMV, 56 to BYMV, 21 to CMV and 42 to SCMoV. The remaining accessions were susceptible to all four viruses with symptoms which ranged from no apparent symptoms (symptomless systemic infection) to highly susceptible and plant death. In continuing work with DAFWA (Dr R. Jones) accessions potentially resistant to AMV, BYMV and CMV are being challenged with additional isolates to check for the presence of genes providing broader resistance.
The Sobemovirus SCMoV was chosen for further study because it is the most widespread viral pathogen of subterranean clover pastures in Australia. It is also a high titre, mechanically transmitted virus which gave the least escapes on infection. SCMoV has a linear, single-stranded positive-sense RNA genome of 4.25 Kb. Making use of natural resistance is an effective means to reduce pasture losses caused by SCMoV. From the screen of the core collection of M. truncatula, amongst the lines resistant to SCMoV a single dominant hypersensitive resistance was detected in line DZA-315. To accelerate mapping of the SCMoV resistance gene, an F8 RIL population of a cross between the resistant line (DZA-315) and a susceptible line (Jemalong-J6, A-17) was sourced and obtained from INRA Toulouse. A total of 166 RILs were phenotyped for resistance and susceptibility to SCMoV. Resistant and susceptible lines showed parental phenotypic symptoms with 84 being susceptible and 82 being resistant. This indicated the presence of a single resistance (R) gene. This phenotypic data was combined with genotypic data (76 polymorphic molecular markers) already available for this RIL population to provide a framework map. Mapmaker and Mapmanager mapping programs were used to locate the position of the resistance gene. This framework map indicated a position for the resistance gene on the long arm of chromosome 6.
Additional polymorphic SSR markers flanking the R gene locus on chromosome 6 were used to map the position of the R gene more closely. These SSR markers were developed from a parental cross of M. truncatula line A17 and A20 at UC Davis and from a parental cross between line A17 and DZA 315 developed at INRA Toulouse. Ten new polymorphic SSR markers were identified and located on the long arm of chromosome 6 after analysis of the F8 RIL population. When combined with the other phenotypic and genotypic data a more accurate map position for the SCMoV R gene was obtained. The results indicate that the R gene to SCMoV is located on the long arm of M. truncatula chromosome 6 between position 35 to 38 centimorgans (cM). The closest marker to the SCMoV R gene is marker mtic153 which is about 2.3 cM away. From existing maps of M. truncatula most of the R genes located in this region are of the TIR-NBS-LRR type and occur in R gene clusters. A series of BACs that span the region of interest have been identified in which SCMoV R gene should be present.
M. truncatula has been used as a model legume to study a number of symbiotic (e.g. rhizobium) and pathogenic interactions (e.g. fungal and nematode), but this is the only example of its use to study legume-virus interactions. The results obtained indicate the potential of using M. truncatula as a model to study resistance response to other legume viruses and provide a firm basis for identifying the hypersensitive R gene that confers resistance to SCMoV.
32
Reyes, Castro Guillermo. "Studies on cocoyam (Xanthosoma spp.) in Nicaragua, with emphasis on Dasheen mosaic virus /." Uppsala : Dept. of Plant Biology and Forest Genetics, Swedish University of Agricultural Sciences, 2006. http://epsilon.slu.se/200607.pdf.
Full text
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Fourie, Michelle Louise. "The potential of wheat, maize, lucerne, and soybean as plant borders to reduce aphid-transmitted virus incidence in seed potatoes." Pretoria : [s. n.], 2008. http://upetd.up.ac.za/thesis/available/etd-09042009-172734/.
Full text
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Webb, Kathleen A. "The effect of a homoeopathic preparation in the control of tobacco mosaic virus." Thesis, 1997. http://hdl.handle.net/10321/2144.
Full textAbstract:
A dissertation submitted in partial compliance with the requirements for the Masters Degree in Technology: Homoeopathy, Technikon Natal, 1997.Most economically important crop plants may become infected with viruses. Several of these virus diseases are limiting factors in agricultural production and have contributed to serious economic and social hardship in many countries, especially in tropical and subtropical regions. Homoeopathic microdoses have been investigated for their role in the control of virus diseases, with good results. However, few of the studies contain statistical analyses. The object of this study was to assess the effect of a homoeopathic preparation of a leaf infected with tobacco mosaic virus (TM Viricum) in the contol of tobacco mosaic virus (TMV). The potencies used were 6CH, 12CH, 30CH and 200CH. iv Trays of 24 tomato seedlings per tray were the subjects of this study. Tomato plants were systemically infected with TMV. Four trays were used per treatment. There was an uninoculated and an inoculated control group. The rest of the test population was divided into two groups. The
M
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Na, Hong. "3'-terminal RNA structures regulate tomato bushy stunt virus replication /." 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:NR29514.
Full textAbstract:
Thesis (Ph.D.)--York University, 2007. Graduate Programme in Biology.Typescript. Includes bibliographical references. Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:NR29514
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Jeffries, Alex Craig. "The study at the molecular level of the New Zealand isolate of Lucerne transient streak sobemovirus and its satellite RNA / Alex Craig Jeffries." Thesis, 1993. http://hdl.handle.net/2440/21450.
Full textAbstract:
Also available as microfiche.Header title of microfiche :"The study of the molecular level of the NZ isolate of LTSV and its satellite RNA"
Bibliography: leaves 102-125.
iv, 126 leaves, [30] leaves of plates : ill. ; 30 cm.
Thesis (Ph.D.) -- University of Adelaide, Dept. of Biochemistry and Dept. of Plant Science, 1994.
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Ibaba, Jacques Davy. "Characterization of potato virus Y (PVY) isolates infecting solanaceous vegetables in KwaZulu-Natal (KZN), Republic of South Africa (RSA)." Thesis, 2009. http://hdl.handle.net/10413/613.
Full textAbstract:
Potato virus Y (PVY) is an economically important virus worldwide. In South Africa, PVY has been shown to be a major limiting factor in the production of important solanaceous crops, including potato (Solanum tuberosum L.), pepper (Capsicum annuum L.), tomato (Lycopersicon esculentum Mill.) and tobacco (Nicotiana spp). The variability that PVY displays, wherever the virus occurs, merits the study of the isolates occurring in KwaZulu-Natal (KZN) in the Republic of South Africa (RSA). This characterization will provide a clear understanding of strains/isolates from local vegetables and how they relate to the other PVY strains already identified, as well as information that can be used to manage the diseases they cause. Hence, the aim of this project was to study the biological and genetic properties of PVY isolates infecting potato, tomato and pepper in KZN. Enzyme-linked immunosorbent assay (ELISA) using monoclonal antibodies and reverse transcription polymerase chain reaction (RT-PCR) using primers specific to all PVY strains were used to detect the virus in plant material showing PVY-like symptoms collected from various locations in KZN. A total of 39 isolates (18 isolates infecting tomato, 12 infecting potato and 9 infecting pepper) were further differentiated into strains by means of ELISA using strain specific antibodies and RT-PCR using primers specific to the different strains of PVY identified around the world. All PVY isolates infecting tomato and pepper tested positive for the ordinary PVYO strain with both ELISA and RT-PCR. PVY isolates infecting potato were more diverse and comprised the PVYN, PVYNTN and PVYNWilga strains, with mixed infections noted in some cases. The biological properties were studied by mechanically inoculating Chenopodium quinoa, Nicotiana tabacum cv Xanthi, N. tabacum cv Samsun, N. glutinosa, and N. rustica with leaf extracts from plants infected with the different PVY strains detected in this study. All inoculated C. quinoa plants did not show symptoms. All tobacco plants showing symptoms were tested for the presence of PVY by means of ELISA using monoclonal antibodies targeting all strains and electron microscopy using the leaf dip technique. Not all the inoculated tobacco tested positive with ELISA. The symptoms observed were therefore divided into PVY-related and PVY non- related. PVY-related symptoms included vein clearing, mosaic chlorosis, stunting, and vein necrosis. PVY non-related symptoms included wrinkles and leaf distortions. Potyvirus-like particles of about 700 nm were observed under the transmission electron microscope (TEM) from plants showing PVY-related symptoms while rod shaped viral particles of sizes varying between 70 and 400 nm were observed from plants showing non-PVY related symptoms. A portion of the virus genome (1067 bp) covering part of the coat protein gene and the 3’ non-translated region (NTR) of three PVYO isolates infecting tomato, one PVYO isolate infecting pepper and one PVYNWilga isolate infecting potato were amplified, cloned and sequenced. The 5’ NTR, P1, HC-Pro and part of P3 regions (2559 bp) of a PVYN isolate infecting potato were also amplified, cloned and sequenced. Sequence data was compared with selected PVY sequences from different geographical locations around the world. These were available on the NCBI website and subsequently used for phylogenic analyses. The sequenced genomic regions of the PVYN isolate were found to be 99% similar to the New Zealand PVYN isolate (GenBank accession number: AM268435), the Swiss PVYN isolate CH605 (X97895) and the American PVYN isolate Mont (AY884983). Moreover, the deduced amino acid sequence comparison of the genomic regions of the PVYN isolate revealed the presence of five distinct amino acids residues. The three amino acid residues (D205, K400, and E419), which determine the vein necrosis phenotype in tobacco, were also identified. The coat protein and 3’ NTR sequences of all KZN PVYO isolates infecting pepper and tomato were closely similar to each other than to KZN PVYNWilga isolate infecting potato. The phylogenic analysis clustered the KZN PVYN isolate with the European sublineage N, PVYNWilga isolate infecting potato with the American PVYO isolate Oz (EF026074) in the O lineage and all PVYO isolates infecting tomato and pepper in a new sublineage within the O lineage. Taken together, these results point to the presence of PVY in solanaceous vegetables cultivated in KZN and they lay the foundation for the formulation of effective control measure against PVY diseases in KZN.Thesis (M.Sc.) - University of KwaZulu-Natal, Pietermaritzburg, 2009.
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Saeed, Muhammad. "The role of a geminiviral DNA β satellite in viral pathogenicity and movement." 2006. http://hdl.handle.net/2440/37886.
Full textAbstract:
Geminiviruses ( family Geminiviridae ) have circular single - stranded genomes encapsidated in twinned quasi - isometric particles and are responsible for major crop losses worldwide. The largest genus, Begomovirus, comprises viruses transmitted by the whitefly Bemisia tabaci. Most begomoviruses have bipartite genomes, termed DNA A and DNA B. The DNA A component encodes proteins required for viral DNA replication and encapsidation whereas the DNA B encodes two proteins that are essential for systemic movement. A small number of begomoviruses have a monopartite DNA genome that resembles the DNA A of bipartite begomoviruses. This DNA carries all gene functions for replication and pathogenesis. Specific small circular single - stranded DNA satellites containing a single open reading frame ( ORF ), termed DNA β, have recently been found in association with certain monopartite begomovirus infections. They comprise about 1350 nucleotides and require a helper begomovirus for replication and encapsidation. DNA β contributes to the production of symptoms and enhanced helper virus accumulation in certain hosts. This study examines the role of DNA β satellite in viral pathogenicity and movement in the host plant. Infectivity analysis of Tomato leaf curl virus and DNA β having mutation in the C1 and V1 ORF indicated that the complementary - sense ORF, βC1, is responsible for inducing disease symptoms in Nicotiana tabacum. An ORF present on the plus strand, βV1, appeared to have no role in pathogenesis. Tobacco plants transformed with the βC1 ORF under the control of the Cauliflower mosaic virus 35S promoter, or with a dimeric DNA β exhibited severe disease - like phenotypes, while plants transformed with a mutated version of βC1 appeared normal. Northern blot analysis of RNA from the transgenic plants using strand - specific probes identified a single complementary - sense transcript. The transcript carried the full βC1 ORF encoding a 118 amino acids product. It mapped to the DNA β nucleotide ( nt ) position 186 - 563 and contained a polyadenylation signal 18 nt upstream of the stop codon. A TATA box was located 43 nt upstream of the start codon. These results indicate that βC1 protein is responsible for DNA β induced disease symptoms. Tomato leaf curl New Delhi virus ( ToLCNDV ) is a bipartite begomovirus in which both DNA A and DNA B are required for systemic infection. Inoculation of tomato plants with ToLCNDV DNA A alone induced local but not systemic infection whereas co - inoculation with DNA A and the DNA β resulted in systemic infection. The presence of both DNA A and the DNA β in systemically infected tissues and the absence of DNA B was confirmed by probe hybridization. DNA β containing a disrupted βC1 ORF did not mobilize the DNA A for systemic infection. Co - inoculation of plants with DNA A and a construct of βC1 ORF, under the control of the Cauliflower mosaic virus 35S promoter, resulted in the systemic movement of the DNA A. βC1 fused to GFP accumulated around and inside the nucleus, at the periphery of tobacco and onion epidermis cells and co - localized with the endoplasmic reticulum. This distribution would be consistent with βC1 mediating intra cellular transport from the nucleus to the plasma membrane. These results showed that the βC1 protein can replace the functions of DNA B in allowing the systemic movement of a bipartite geminivirus DNA A.Thesis (Ph.D.)--School of Agriculture, Food and Wine, 2006.
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Saeed, Muhammad. "The role of a geminiviral DNA β satellite in viral pathogenicity and movement." Thesis, 2006. http://hdl.handle.net/2440/37886.
Full textAbstract:
Geminiviruses ( family Geminiviridae ) have circular single - stranded genomes encapsidated in twinned quasi - isometric particles and are responsible for major crop losses worldwide. The largest genus, Begomovirus, comprises viruses transmitted by the whitefly Bemisia tabaci. Most begomoviruses have bipartite genomes, termed DNA A and DNA B. The DNA A component encodes proteins required for viral DNA replication and encapsidation whereas the DNA B encodes two proteins that are essential for systemic movement. A small number of begomoviruses have a monopartite DNA genome that resembles the DNA A of bipartite begomoviruses. This DNA carries all gene functions for replication and pathogenesis. Specific small circular single - stranded DNA satellites containing a single open reading frame ( ORF ), termed DNA β, have recently been found in association with certain monopartite begomovirus infections. They comprise about 1350 nucleotides and require a helper begomovirus for replication and encapsidation. DNA β contributes to the production of symptoms and enhanced helper virus accumulation in certain hosts. This study examines the role of DNA β satellite in viral pathogenicity and movement in the host plant. Infectivity analysis of Tomato leaf curl virus and DNA β having mutation in the C1 and V1 ORF indicated that the complementary - sense ORF, βC1, is responsible for inducing disease symptoms in Nicotiana tabacum. An ORF present on the plus strand, βV1, appeared to have no role in pathogenesis. Tobacco plants transformed with the βC1 ORF under the control of the Cauliflower mosaic virus 35S promoter, or with a dimeric DNA β exhibited severe disease - like phenotypes, while plants transformed with a mutated version of βC1 appeared normal. Northern blot analysis of RNA from the transgenic plants using strand - specific probes identified a single complementary - sense transcript. The transcript carried the full βC1 ORF encoding a 118 amino acids product. It mapped to the DNA β nucleotide ( nt ) position 186 - 563 and contained a polyadenylation signal 18 nt upstream of the stop codon. A TATA box was located 43 nt upstream of the start codon. These results indicate that βC1 protein is responsible for DNA β induced disease symptoms. Tomato leaf curl New Delhi virus ( ToLCNDV ) is a bipartite begomovirus in which both DNA A and DNA B are required for systemic infection. Inoculation of tomato plants with ToLCNDV DNA A alone induced local but not systemic infection whereas co - inoculation with DNA A and the DNA β resulted in systemic infection. The presence of both DNA A and the DNA β in systemically infected tissues and the absence of DNA B was confirmed by probe hybridization. DNA β containing a disrupted βC1 ORF did not mobilize the DNA A for systemic infection. Co - inoculation of plants with DNA A and a construct of βC1 ORF, under the control of the Cauliflower mosaic virus 35S promoter, resulted in the systemic movement of the DNA A. βC1 fused to GFP accumulated around and inside the nucleus, at the periphery of tobacco and onion epidermis cells and co - localized with the endoplasmic reticulum. This distribution would be consistent with βC1 mediating intra cellular transport from the nucleus to the plasma membrane. These results showed that the βC1 protein can replace the functions of DNA B in allowing the systemic movement of a bipartite geminivirus DNA A.Thesis (Ph.D.)--School of Agriculture, Food and Wine, 2006.
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Fajolu, Oluseyi Lydia. "Genetic variability of Hosta virus X in hosta." 2009. http://etd.utk.edu/2009/May2009Theses/FajoluOluseyiLydia.pdf.
Full text
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Torok, Valeria Anna. "Biological and molecular variation among isolates of pea seed borne mosaic virus / Valeria Anna Torok." Thesis, 2001. http://hdl.handle.net/2440/21692.
Full textAbstract:
Corrigendum inserted at the back.Includes bibliographical references (leaves 133-158).
xvi, 158 leaves : ill., col. map ; 30 cm.
Sixteen pea seed borne mosaic virus (PSbMV) isolates were collected between 1995 and 1998. These isolates were biologically distinct yet serologically indistinguishable. The conclusion is that PSbMV is widespread and occurs at a low incidence in Australia. Reports sequence information on new isolates of PSbMV which has allowed genomic regions to be identified which distinguish PSbMV pathotypes and isolates; and, to the development of PSbMV nucleic acid hybridisation and RT-PCR assays.
Thesis (Ph.D.)--University of Adelaide, Dept. of Applied and Molecular Ecology, 2001
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Halgren, Anne B. "Characterization, epidemiology, and ecology of a virus associated with black raspberry decline." Thesis, 2006. http://hdl.handle.net/1957/29931.
Full textAbstract:
The objective of this study was to characterize an unknown agent associated with
decline in black raspberry (Rubus occidentalis) in Oregon. A virus was found
consistently associated with decline symptoms of black raspberries and was named Black
raspberry decline associated virus (BRDaV). Double stranded RNA extraction from
BRDaV-infected black raspberry revealed the presence of two bands of approximately
8.5 and 7 kilobase pairs, which were cloned and sequenced. The complete nucleotide
sequences of RNA 1 and RNA 2 are 7581 nt and 6364 nt, respectively, excluding the 3'
poly(A) tails. The genome structure was identical to that of Strawberry mottle virus
(SMoV), with the putative polyproteins being less than 50% identical to that of SMoV
and other related sequenced viruses. The final 189 amino acids of the RNA-dependent-
RNA-polymerase (RdRp) reveal an unusual indel with homology to AlkB-like protein
domains, suggesting a role in repair of alkylation damage. This is the first report of a
virus outside the Flexiviridae and ampeloviruses of the Closteroviridae to contain these
domains. An RT-PCR test was designed for the detection of BRDaV from Rubus tissue.
BRDaV is vectored non-persistently by the large raspberry aphid Amphorophora
agathonica, the green peach aphid Myzus persicae, and likely nonspecifically by other
aphid species. Phylogenetic analysis of conserved motifs of the RdRp, helicase, and
protease regions indicate that BRDaV belongs to the Sadwavirus genus.
To assess the rate of spread BRDaV, four newly planted fields of black
raspberries (Rubus occidentalis) in Oregon were studied for three years. In an effort to
characterize the suspected complexity of synergistic interactions between BRDaV and
other Rubus-infecting viruses, the prevalence of ten additional Rubus viruses was also
monitored in the study fields. The timing of BRDaV infection as it relates to aphid
populations and flights was also determined. Testing of nearby vegetation identified
several symptomless Rubus hosts of BRDaV, as well as detection in multiple cultivars of
black raspberry and several non-Rubus weeds. It was determined that BRDaV spreads
rapidly with a low aphid threshold and consistently is associated with decline of black
raspberries in Oregon.Graduation date: 2006
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Cook, Meridith Ayn. "Development of new tools for the application of biotechnology to agricultural improvement and assessing risks of biotechnology and its products." Diss., 2008.
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Mariote, David. "Response to selection for downy mildew (Peronosclerospora sorghi) and maize streak virus resistance in three quality protein maize populations in Mozambique." Thesis, 2007. http://hdl.handle.net/10413/748.
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Gichuru, Lilian Njeri. "Breeding investigations on utility of maize streak virus resistant germplasm for hybrid development in the tropics." Thesis, 2014. http://hdl.handle.net/10413/10694.
Full textAbstract:
Maize (Zea mays L.) supports millions of livelihoods in sub-Saharan Africa (SSA) in terms of
food and feed. Production of the crop is however limited by several factors, among these, maize
streak virus (MSV) disease. Although extensively studied, MSV remains a serious problem in
SSA due to several challenges in breeding MSV resistant maize varieties. These include
integration of MSV resistant germplasm from different backgrounds, reliance on a few resistant
sources, and genotype x environment interactions. This study was designed to assess the
breeding potential of several MSV resistant lines in hybrid combinations. Understanding
architecture of genetic divergence and background of these genotypes would greatly aid in
breeding high yielding and stable MSV resistant hybrids. Experiments were conducted during
2010 to 2012 seasons in Kenya. Diallel crosses and SSR markers were used to characterize
MSV resistant maize inbred lines from three programs of CIMMYT, KARI and IITA.
In general, this study revealed that MSV is still an important problem in Kenya with high
incidence and severity levels in the farmers’ fields. The levels of MSV resistance in locally
grown hybrids needs to be improved. Farmers challenged breeders to develop new hybrids that
combine early maturing, high yield potential and MSV resistance.
The study was successful in identifying the best eight inbred lines for use in breeding new maize
hybrids with MSV resistance. The nature of gene effects was established for the first time, in
particular the role of epistasis and G x E in conditioning MSV resistance in hybrids. Results
indicate serious implications for previous models that ignored epistasis in studying MSV
resistance in maize. The inbreds Z419, S558, CML509 and Osu23i, displayed high levels of
epistasis for MSV resistance. Unless strong sources of MSV resistance, such as MUL114 and
CML509, are used, breeding resistant hybrids will require parents that carry dominant
resistance genes. The additive-dominance model was adequate to explain northern leaf blight
(NLB) resistance in hybrids, indicating fewer complications in breeding NLB resistant hybrids.
The study also reveals that SSR genetic distance data can be used to predict hybrid
performance, especially when the correct set of markers is used. Many previous studies have
not found any significant relationship between genetic distance and heterosis, due to large
G x E and use of a wrong set of markers. The diallel analysis and SSR data established the
important heterotic groups, which will be exploited for efficient development of MSV resistant
maize hybrids. These strategies will be recommended to programs that emphasize MSV
resistance in maize hybrids.Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.
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Domola, Mapula Julia. "Survey and characterisation of sweet potato viruses in South Africa." Diss., 2004. http://hdl.handle.net/2263/24205.
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Sow, Mounirou El-Hassimi. "Genetic diversity of Oryza species in Niger ; screening and breeding for resistance to rice yellow mottle virus (RYMV)." Thesis, 2012. http://hdl.handle.net/10413/8520.
Full textAbstract:
Rice is a staple food in many West African countries, including Niger. However, both regional and national rice production have failed to meet demand due to several constraints, among which is the Rice yellow mottle virus (RYMV). Moreover, attempted intensification of rice cultivation and the introduction of modern cultivars are encouraging farmers towards abandoning local landraces for high yielding, but often susceptible varieties. The study was primarily oriented towards rice pre-breeding, and identifying priorities for rice breeding in Niger in relation to farmers' preferences and their environment. A secondary aim was the development and evaluation (for release at the regional level) of new breeding lines with resistance to RYMV. This study aimed to: 1) Establish farmers' perception of rice varieties as well as the main constraints on rice production in Niger and particularly those posed by RYMV; 2) Create a collection of rice species from Niger for ex- situ conservation, and to determine the phenotypic variability within this collection; 3) Determine the genetic diversity and population structure of the collection; 4) Screen the collection for resistance to RYMV, so that new sources of resistance could be detected; 5) Improve five elite varieties from West Africa for resistance to RYMV using marker-assisted selection (MAS). The germplasm collection and PRA of this study were conducted in 2008 and 2009 in Niger, while the field and the laboratory researches were conducted in 2008 and 2009 at the Africa Rice Center (AfricaRice) in Benin. For the PRA, data was obtained from a semi-structured group discussion carried out in 14 villages, individual questioning of 153 farmers and visits to farmers' field and storage facilities. The local farmers' union was the only formal seed dissemination system. Seed exchanges between farmers and the use of seeds from previous harvests were important. The RYMV and the bacterial leaf blight (BLB) were cited as the prevalent biotic stresses in the irrigated agrosystem, where the varieties IR1529-680-3 and Waihidjo were found to be the most popular. Flood, birds and hippopotamus were the most damaging agents in the lowland cropping system, and the landrace Degaulle/ D5237 was the preferred variety. Apart from the yield, farmers preferred varieties with good grain quality (milling quality and good taste), high market value, stress tolerance (drought, flood, disease, birds, rodents), and those recommended by the local farmers' association. These findings should be included in breeding goals, seed production and dissemination systems. During collection, a total of 270 rice accessions were assembled, comprising the two cultivated rice species Oryza sativa L. and O. glaberrima Steud. and its two wild relatives Oryza barthii A. Chev. and O. longistaminata Chev. et Roehr. The region of the Niger River and its tributary (the Dallol Maouri) provided the majority (80.7%) of the accessions. Apart from a few wild O. barthii accessions, the accessions found around Lake Chad and the Komadougou river (South-East) were also collected in the Niger River area. Farmers' naming and ecological classification of rice varieties was generally consistent. Three major phenotypic groups were found during the field trials, and the overall phenotypic variability of the collection (as measured by the Shannon-Weaver Diversity Index) was relatively high. There was no significant difference in diversity between the main eco-geographical zones of collection, as well as between the identified phenotypic groups, suggesting a high level of germplasm exchange between the regions in Niger.
From the collection, 264 accessions were genotyped from the collection using 18 well distributed SSR markers and two main genetic compartments were detected, comprising O. sativa subsp. indica varieties and O. glaberrima and its wild relative O. barthii and O. longistaminata. The O. sativa group in Niger was divided into irrigated and floating rice, bound by lowland rice. The wild progenitor O. barthii was widespread but without any clear genetic differentiation from O. glaberrima, probably due to the presence of admixtures within the collected samples of O. barthii. Allelic diversity was relatively high, despite the geographical distance from the centre of domestication of African rice, and the points of entry of Asian rice to Africa. The findings reflect the underuse of Niger's rice landraces genetic potential for rice breeding, given that all the "improved" varieties released during the last 25 years in Niger were clustered together on the dendrogram. The response of a set of the rice collected from Niger and some accessions from Mali to inoculation by RYMV was evaluated using five different virus isolates from Niger (3), Benin (1) and Burkina Faso (1). All rice varieties were susceptible to the disease. However, depending on the virus strain, a few O. glaberrima accessions displayed partial resistance, similar to the highly resistant TOG5681. Allelic research based on primers derived from the RYMV1 gene revealed one accession with allele rymv1-3, and two accessions with allele rymv1-4, and one accession with a different resistance gene. The implications of the finding were discussed and a strategy proposed for breeding varieties with a comprehensive resistance to RYMV.
After three generations of backcrossing, the major resistance gene of the variety Gigante was successfully introgressed into five elite rice varieties of West Africa by Marker-Assisted Backcross (MABC). The newly developed BC3F3 progenies were screened for resistance to RYMV in farmers' fields in Guinea and Mali and also under controlled conditions in a screenhouse in Benin. As shown by low virus content and level of disease incidence, low tiller number and plant height reduction, the transferred gene was fully functional in the new genetic background. Moreover, some lines also displayed a high level of resistance to rice blast (Pyricularia oryzae) and stem borer infestation in Guinea. Four of those lines are in the second year of multi-location trial in seven West African countries. Therefore, effective deployment of the newly developed varieties, coupled with good cultural practices, should reduce the damaging effects of RYMV in lowland and irrigated rice cropping systems and thereby increase the income of small scale farmers from rice cultivation.Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.
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Abalo, Grace. "A comparative analysis of conventional and marker assisted selection methods in screening for resistance to maize (Zea mays L.) streak virus disease." Thesis, 2006. http://hdl.handle.net/10413/3495.
Full textAbstract:
Maize (Zea mays L.) streak virus disease (MSD) is the most important virus disease in Africa but farmers are unaware of its status. A project was initiated to assess the current status of MSD and to breed for its resistance. Four populations comprised of two BC1F1 and two F2 progenies developed by backcrossing and selfing the F1 progenies of two crosses between a donor line (CMl 202) and two susceptible lines (CMl 321 and CMl 384) were developed. Conventional and molecular marker assisted selection (MAS) methods were used to screen for resistance to MSD in each of the four populations. To facilitate unbiased comparison, separate screening nurseries were established for MAS and conventional screening. The objectives of the study were five-fold; 1) to assess the status of
MSD in Uganda and understand farmers' preferences and varietal selection criteria for maize using a participatory rural appraisal (PRA), 2) to screen for MSD resistance in early generations of segregating maize populations using conventional method, 3) to screen for resistance to MSD using SSR marker assisted selection , 4) to compare the effectiveness of marker assisted selection and conventional methods for selection for resistance to MSD,
and 5) to compare costs associated with MAS and conventional selection methods.
Results of PRA showed that unreliable rainfall and insect pests were the dominant constraints to maize productivity in Uganda. Diseases were ranked fifth among the production constraints . Maize streak virus disease was considered the most important disease constraint. Farmers showed common preference for high yielding and early maturing cultivars. However, farmers had other special preferences which were diverse and included large, white and high test density kernels for marketing, and sweet taste,
particularly for home consumption. Farmers' research priorities included tolerance to drought, resistance to insect pests and diseases, sweetness, prolificacy, resistance to lodging, and drooping leaves because theyt cover the soil fast and prevent weed growth.
Conventional screening for resistance to MSD showed that backcross and selfing populations segregated in 1:1 and 3:1 Mendelian ratios confirming the presence of one major gene with simple inheritance . Severity and incidence of disease were positively correlated suggesting a non-reference by the insects. In the selfing populations, the presence of complete esistance against MSD was suggested because frequency distribution patterns were highly skewed in favour of resistance. There was a decrease in disease severities with selection from BC1F1 to BC2F1 and from F2 to F3 generations indicating that high response to selection was achieved. On the other hand, one marker, umc1917, consistently polymorphic and eo-dominant was selected and used in MAS protocol. Results showed that the observed outcomes fitted the expected ratio of 1:2:1 for a F2 population and 1:1 for a BC1F1 population (X2 not significant). Evaluation of F3 and
BC2F1 progeny selected using markers showed low disease severity suggesting that marker assisted selection was effective. However, the study showed that the presence of the O'Tl, was not consistent with symptom expression in the field.
Evaluation of lines in three-way crosses identified ten potential lines that were high yielding, highly resistant to MSD and stable across three locations. Both MAS and conventional selection were equally effective in identifying high yielding lines although resistance was higher under MAS.
Costs of MAS and conventional method varied depending on the units for
comparison. The total costs of conventional method were higher than that of MAS in both first and second selection cycles. Comparing costs per row for conventional and costs per plant or data point for MAS showed that conventional selection was 2.4 times more expensive than costs per sample for MAS. However, costs per plant for MAS were 6.6 times
higher than for conventional selection.Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2006
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Sivparsad, Benice. "The development of transgenic sweet potato (Ipomoea batatas L.) with broad virus resistance in South Africa." Thesis, 2013. http://hdl.handle.net/10413/10038.
Full textAbstract:
Sweet potato (Ipomoea batatas Lam.) is ranked as the seventh most important food crop in the world and its large biomass and nutrient production give it a unique role in famine relief. However, multiple virus infection is the main disease limiting factor in sweet potato production worldwide. The main objective of this research project was to develop a transgenic sweet potato cultivar with broad virus resistance in South Africa (SA).
A review of current literature assembled background information pertaining to the origin, distribution and importance of the sweet potato crop; viruses and complexes infecting sweet potato; and the strategies used in sweet potato virus detection and control.
A survey to determine the occurrence and distribution of viruses infecting sweet potato (Ipomoea batatas Lam.) was conducted in major sweet potato-growing areas in KwaZulu-Natal (KZN). A total of 84 symptomatic vine samples were collected and graft inoculated onto universal indicator plants, Ipomoea setosa Ker. and Ipomoea nil Lam. Six weeks post inoculation, typical sweet potato virus-like symptoms of chlorotic flecking, severe leaf deformation, stunting, chlorotic mosaic, and distinct interveinal chlorotic patterns were observed on indicator plants. Under the transmission electron microscope (TEM), negatively stained preparations of crude leaf sap and ultra-thin sections from symptomatic grafted I.setosa plants revealed the presence of elongated flexuous particles and pinwheel type inclusions bodies‟ that are characteristic to the cytopathology of Potyviruses. Symptomatic leaf samples from graft-inoculated I. setosa and I. nil were assayed for Sweet potato feathery mottle virus (SPFMV), Sweet potato mild mottle virus (SPMMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato virus G (SPVG), Sweet potato mild speckling virus (SPMSV), Sweet potato caulimo-like virus (SPCaLV), Sweet potato latent virus (SPLV), Cucumber mosaic virus (CMV), and Sweet potato C-6 virus (C-6) using the nitrocellulose membrane enzyme-linked immunosorbent assay (NCM-ELISA). The majority of leaf samples (52%) tested positive for virus disease and showed the
occurrence of SPFMV, SPMMV, SPCSV, SPCFV, SPVG, SPMSV, and SPCaLV. Of these 7 viruses, the most frequently detected were SPFMV (39%), SPVG (30%), followed by SPCSV (13%) and SPMMV (12%). SPCaLV and SPCFV at 10% and SPMSV at 7% were found exclusively in samples collected from one area. SPFMV, SPVG, SPCSV, and SPMMV were identified as the most prevalent viruses infecting sweet potato in KZN.
The genetic variability of the three major viruses infecting sweet potato (Ipomoea batatas Lam.) in KZN was determined in this study. A total of 16 virus isolates originating from three different locations (Umbumbulu, Umfume and Umphambanyomi River) in KZN were analyzed. These comprised of 10 isolates of Sweet potato feathery mottle virus (SPFMV), five isolates of Sweet potato virus G (SPVG) and one isolate of Sweet potato chlorotic stunt virus (SPCSV). The phylogenetic relationships of the SPFMV, SPVG and SPCSV isolates from KZN relative to isolates occurring in SA and different parts of the world were assessed. The division of SPFMV into four genetic groups (strains) according to the phylogenetic analysis of coat protein encoding sequences revealed mixed infections of the O (ordinary) and C (common) strains in sweet potato crops from KZN. All SPFMV isolates showed close lineage with isolates from South America, East Asia and Africa. The SPVG isolates showed high relatedness to each other and close lineage with other isolates, especially those from China and Egypt. Analysis of the partial sequence of the Heat shock protein 70 homologue (Hsp70h) gene indicated that the SPCSV isolate from KZN belongs to the West African (WA) strain group of SPCSV and showed close relatedness to an isolate from Argentina. The knowledge of specific viral diversity is essential in developing effective control measures against sweet potato viruses in KZN.
Multiple virus infections of Sweet potato feathery mottle virus (SPFMV), Sweet potato chlorotic stunt virus (SPCSV), Sweet potato virus G (SPVG) and Sweet potato mild mottle virus (SPMMV) cause a devastating synergistic disease complex of sweet potato (Ipomoea batatas Lam.) in KZN. In order to address the problem of the multiplicity and synergism of sweet potato viruses in KZN, this study aimed to develop transgenic sweet
potato cv. Blesbok with broad virus resistance. An efficient and reproducible plant regeneration protocol for sweet potato (Ipomoea batatas Lam.) cultivar Blesbok was also developed in this study. The effect of different hormone combinations and type of explants on shoot regeneration was evaluated in order to optimize the regeneration protocol. Coat protein (CP) gene segments of SPFMV, SPCSV, SPVG and SPMMV were fused to a silencer DNA, the middle half of the nucleocapsid (N) gene of Tomato spotted wilt virus (TSWV) and used as a chimeric transgene in a sense orientation to induce gene silencing in the transgenic sweet potato. Transformation of apical tips of sweet potato cv. Blesbok was achieved by using Agrobacterium tumefaciens strain LBA4404 harboring a modified binary vector pGA482G carrying the plant expressible neomycin phosphotransferase ll gene (nptll), the bacterial gentamycin-(3)-N-acetyl-transferase gene and the expression cassette. A total of 24 putative transgenic plants were produced from the transformed apical tips via de novo organogenesis and regeneration into plants under 50mg/L kanamycin and 200 mg/L carbenicillin selection. Polymerase chain reaction (PCR) and Southern blot analyses showed that six of the 24 putative transgenic plants were transgenic with two insertion loci and that all plants were derived from the same transgenic event. The six transgenic sweet potato plants were challenged by graft inoculation with SPFMV, SPCSV, SPVG and SPMMV- infected Ipomoea setosa Ker. Although virus presence was detected using NCM-ELISA, all transgenic plants displayed delayed and milder symptoms, of chlorosis and mottle of lower leaves when compared to the untransformed control plants. These results warrant further investigation under field conditions.Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.
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Moodley, Vaneson. "Development of a pepper (Capsicum annuum L.) hybrid variety with resistance to potato virus Y (PVY) using molecular breeding." Thesis, 2013. http://hdl.handle.net/10413/10829.
Full textAbstract:
Pepper (Capsicum annuum L.) is an important vegetable crop grown and consumed worldwide. Potato virus Y (PVY) is a globally economically important pathogen which significantly reduces the yield and quality of cultivated pepper. The virus is considered as a major limiting factor to the economic production of pepper in the province of KwaZulu-Natal (KZN) in the Republic of South Africa (RSA). Many applied practices to control the spread of PVY are ineffective to mitigate the losses incurred by many farming communities across the KZN province. Therefore, the objectives of this study was to determine the full genome sequence of a PVY isolate from KZN, to identify resistance alleles in commercially available pepper varieties in KZN and to develop a pepper hybrid variety with resistance to PVY using a molecular breeding strategy
The first part of the study was conducted to determine the first full genome sequence of a PVY isolate (JVW-186) infecting pepper from KZN. The complete genome sequence of JVW-186 was assembled from overlapping RT-PCR clones using MEGA 5 software. Individual ORFs were identified using the nucleotide data base NCBI and aligned using CLUSTALW. RDP4 software was used to identify recombination junctions in the sequence alignment of JVW-186. CLC Main Workbench 6 software was used to determine the nucleotide sequence similarity of recombinant and non-recombinant fragments of JVW-186 in conjunction with ten PVY parental isolates. Based on sequence data, virus morphology and the coat protein size as determined by SDS-PAGE analysis, the identity of the isolate JVW-186 was confirmed as PVY. Phylogenetic trees were constructed from all recombinant and non-recombinant segments of the sequence by the maximum likelihood method using MEGA 5 software. The full length sequence of JVW-186 consisted of 9700bp. Two ORF’s were identified at position 186 and 2915 of the sequence alignment encoding the viral polyprotein and the frameshift translated protein P3N-PIPO, respectively. RDP4 software confirmed two recombination breakpoints at position 343 and 9308 of the sequence resulting in four segments of the genome. At each recombination event, a 1021-bp fragment at the 5’
end in the region of the P1/HC-Pro protein and a 392-bp fragment in the region of the coat protein shared a high sequence similarity of 91.8 % and 98.89 % to the potato borne PVYC parental isolate PRI-509 and the PVYO parental isolate SASA-110 respectively. The non-recombinant fragment 1 clustered within the C clade of PVY isolates; however the large 7942-bp fragment 3 did not cluster within any of the clades although it shared > 80% nucleotide sequence similarity to other PVY isolates used in this study. Our results suggest that isolate JVW-186 is a novel recombinant strain of PVY that could have evolved due to the dynamics of selection.
The second part of the study aimed to evaluate different pepper lines for resistance to PVY. Two recessive alleles (pvr21 and pvr22) located on the pvr2-elF4E locus are known to confer resistance to the virus. To this end, six pepper lines were challenged with PVY infected Nicotiana tabacum cv. Xanthi leaf material using mechanical inoculation under greenhouse conditions. Each line was assessed for resistance to PVY by visual screening for disease severity and quantitative enzyme linked immunosorbent assay (ELISA) for virus load. Pepper lines were further characterized using tetra-primer ARMS-PCR (amplification refractory mutation system polymerase chain reaction) to identify and differentiate the presence of homozygous/heterozygous resistance alleles that confer PVY resistance. Evaluations revealed two resistant pepper lines (Double Up and Cecelia) and varying levels of susceptibility in the other four pepper lines challenged with PVY. The most susceptible pepper line was Benno, although high levels of susceptibility were observed in three other lines (IP, Mantenga and Excellence). The pvr2+ allele was positively identified in all the susceptible pepper lines using the T200A tetra-primer which confirms that the presence of this allele is dominant for PVY susceptibility. Double Up and Cecelia were genotyped homozygous pvr21/pvr21 and pvr22/pvr22 respectively, and remained asymptomatic throughout the trial which indicates that these alleles confer resistance to the isolate of PVY used in this study. The information generated in this study can be incorporated into breeding programs intended to control PVY on pepper in KZN.
The final part of the study focused on the development of resistant varieties as the best alternative to manage PVY diseases on pepper. Homozygous F2 pepper lines were
developed from local germplasm carrying PVY resistance genes (pvr21 and pvr22) using marker assisted selection (MAS). The F1 progeny was obtained by crossing a homozygous pvr21 (resistant) ‘Double Up’ cultivar with a heterozygous susceptible (pvr2+/pvr22) ‘Benno’ cultivar. F1 and F2 generations were assessed for the presence of PVY resistance/susceptibility alleles (pvr2+/pvr21/pvr22) at the pvr2-elF4e locus using the tetra primer amplification refractory mutation system – polymerase chain reaction (ARMS-PCR) procedure. Negative selection was carried out using the tetra-primer T200A marker to detect the pvr2+ (susceptible) allele. All F1 progeny displaying the pvr2+ allele were eliminated from further study. All 302 plants belonging to 29 F2 families expressing homozygous recessive traits were tested via mechanical inoculation for their response to PVY infection and resistance to PVY was confirmed in all selected families based on symptomatology in greenhouse house screens using double antibody sandwich enzyme linked immunosorbent assay (DAS-ELISA). These results show that ARMS-PCR can be used to successfully screen pepper genotypes for alleles that confer PVY resistance thereby contributing to the improvement of pepper production using molecular breeding approaches.Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.