Browsing by Author "Roos, Wiets Gideon"

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    An assessment of the diversity and pathogenicity of Potato leafroll virus in South Africa
    (Stellenbosch : Stellenbosch University, 2018-03) Roos, Wiets Gideon; Bellstedt, D. U.; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: Potato production in South Africa has steadily intensified through improved pivot irrigation. Since potatoes are vegetatively propagated the potato industry is continuously threatened by Potato leafroll virus (PLRV) which is responsible for increasing yield losses in South Africa. Effective management of PLRV is dependent on its accurate detection in seed potato stocks. PLRV is a small spherical plant virus consisting of a protein capsid and an RNA genome of approximately 5900 bases. This virus is phloem restricted and is vectored by, most notably, by the green peach aphid. Plant RNA viruses pose a threat due to their high mutation rate and the ability to adapt rapidly to a changing environment. To effectively manage PLRV infection, detection of virus infection in seed potatoes is paramount. In this study, five field trials were carried out in potato production fields, to compare the commonly used DAS-ELISA with RT-PCR for PLRV detection. From the results obtained it was concluded that DAS-ELISA detection greatly underestimates the number of infected samples when compared to RT-PCR. The hotter climate of the Sandveld region appears to inhibit PLRV accumulation in infected plants and these infections then remain undetected by DAS-ELISA. PLRV isolates were sequenced and phylogenetic and bioinformatic analyses were performed to identify and characterise local variants of PLRV. PLRV isolates found in the Sandveld region were closely related to PLRV isolates from Australia. Some of these isolates had recombined with variants commonly found in Europe, Asia and the Americas as well as with those similar to PLRV isolates from Peru and Germany. Three locally produced cultivars were graft-inoculated with two PLRV isolates that represent the two main variant groups found to assess symptom development and yield reduction. Symptom development in locally produced cultivars was typical for PLRV. A yield loss resulted from this infection with no difference between the Australian type and the European/Australian recombinant type. The proteins produced by the newly sequenced isolates were further analysed in comparison to other isolates found worldwide. The variation in the proteins produced by the newly sequenced isolates was mainly due to recombination between distinct groups of PLRV in the 5’ half of the genome and through mutation in the 3’region of the genome. A differential RT-PCR was designed to distinguish, in a single reaction, between the Australian type and the European/Australian recombinant type of PLRV. This revealed that simultaneous infections with both types occurred commonly, and could explain why recombination has occurred.
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    An investigation of prevalance and the detection and race identification of South African potato viruses
    (Stellenbosch : Stellenbosch University, 2013-03) Roos, Wiets Gideon; Bellstedt, D. U.; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: Infection of potatoes by viral pathogens causes reduced crop yield and subsequent economic loss. In South Africa Potato virus Y (PVY) and Potato leafroll virus (PLRV) are the two most destructive viruses infecting potatoes. Several other viral pathogens exist, including Potato virus X (PVX), Potato virus M (PVM), Potato virus A (PVA), Potato virus S (PVS), Potato mop-top virus (PMTV), Tomato spotted wilt virus (TSWV) and Potato spindle tuber viroid (PSTVd). Although the aforementioned pathogens are found infecting potatoes around the world, there are no published information pertaining to the prevalence of these viral agents in South Africa. Currently, the occurrence of PLRV infection in potatoes of South Africa has reached epidemic proportions. A previous phylogenetic investigation undertaken in our laboratory of South African PLRV isolates, using coat protein (CP) gene sequences, found large variation between native South African PLRV isolates and most other isolates from elsewhere in the world; with their nearest relatives being single isolates from Australia and North America. In this study the incidence of PVX, PVM, PVA, PVS, PMTV, TSWV and PSTVd was investigated. A large number of potato plant and tuber samples was collected and infected samples were identified with reverse transcriptase polymerase chain reaction (RT-PCR) amplification of the CP gene or the whole genome in the case of PSTVd. The amplified nucleic acid segments were sequenced, aligned with international reference sequences and analysed phylogenetically to determine their relative relationships with these reference sequences. The CP genes of PLRV isolates were sequenced and phylogenetically investigated to determine how these new isolates compared relative to the previous findings from our laboratory. In addition, the complete genomes of two PLRV isolates were sequenced and phylogenetically investigated as a preliminary study to investigate the apparent increase of pathogenicity of certain variants of South African PLRV. Results obtained showed that only PVX and PVS were present in the samples collected and the incidences of these viruses were very low (2.0 and 1.1% respectively). The phylogenetic analyses of the CP genes, indicated that the PVX and PVS variants isolated in this study, were part of the dominant types of variants found worldwide. From the analyses of the PLRV CP and whole genome sequences, it was determined that many of the PLRV variants found in South Africa, are genetically distinctly different from those around the world. This warrants further investigation into the increased pathogenicity experienced with South African PLRV.