Thesis (PhD)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Potato virus Y (PVY) is responsible for considerable yield losses in the South African potato industry.
The incidence of this virus has greatly increased over the past 20 years. In previous studies nonrecombinant
strains of PVY, PVY N and PVY O, were detected in South African potatoes. In a recent study
the occurrence of non-recombinant strains of PVY in South African potatoes was shown to have
decreased while infection by more virulent recombinant strains, PVY NTN and PVY N-W, had increased
dramatically. Infection of potato plants with PVY may cause stunted growth and mosaic or necrotic leaf
symptoms which in turn can lead to a significant reduction in yield. Highly virulent recombinant PVY
isolates as well as some of the non-recombinant strains may cause potato tuber necrotic ringspot disease
(PTNRD) which may result in losses of 10% to total crop failure. For this reason investigation of
infection by local recombinant isolates on local cultivars was important. To this end a representative
number of isolates were selected for whole genome sequencing based on the relative occurrence of the
various isolates in South Africa. A number of these sequenced isolates were subsequently used to infect
local cultivars of potato in order to investigate the influence of genetic variation within the viral genome
on symptom expression. In this study 27 South African isolates of PVY were sequenced through overlapping RT-PCR fragments.
Seven of these isolates, six PVY NTN and one PVY N-W, were used to mechanically infect four local
cultivars of potatoes under greenhouse conditions. The infected plants were monitored to establish the
rate of systemic spread using a highly sensitive qRT-PCR and resulting tubers were visually screened for
PTNRD. Highly variable recombinant isolates appear to be less virulent than the more conserved
recombinant isolates possibly indicating molecular determinants for pathogenicity. For this reason the
amino acid sequences of the South African isolates were compared to those of international isolates and
scrutinized for variation and substitutions. Some South African isolates displayed amino acid
substitutions unique to the specific isolate, making them unlike those found internationally. Substitution
rates throughout the amino acid sequences differed greatly, with some isolates displaying hardly any
changes whilst others varied a great deal from overseas isolates. Certain regions, many of which had
specific functions, were more conserved than others. This study further investigated the recombination events within the PVY genome using reticulate
phylogenetic analysis, molecular dating and network construction techniques. Unlike existing approaches,
the one described in this study neither assumes an underlying strictly bifurcating species tree nor assumes
prior knowledge of processes underlying deviations between individual gene trees. Through the use of the
resulting robust time calibrated phylogeny, the patterns of diversification and recombination in PVY may
be placed in the historical context of human cultivation of potatoes. Through the use of these techniques
the study aimed to test whether diversification of the major strains of PVY and recombination between
them occurred within the time frame of the domestication and modern cultivation of potatoes. From these
analyses it can be deduced that recombinant strains of PVY were imported into South Africa. / AFRIKAANSE OPSOMMING: Aartappel virus Y (PVY) is verantwoordelik vir aansienlike opbrengs verliese in die Suid-Afrikaanse
aartappelbedryf. Die voorkoms van die virus het grootliks toegeneem oor die afgelope 20 jaar. In vorige
studies is nie-rekombinante rasse van PVY, PVY N en PVY O, gedokumenteer in Suid-Afrikaanse
aartappels. 'n Onlangse studie het gevind dat die voorkoms van nie-rekombinante rasse van PVY in Suid-
Afrikaanse aartappels aansienlik gedaal het terwyl infeksie deur virulente rekombinante rasse, PVY NTN en PVY N-W, dramaties toegeneem het. Infeksie van aartappelplante met PVY kan vertraagde groei en
mosaïek- of nekrotiese blaarsimptome veroorsaak wat kan lei tot aansienlike vermindering in opbrengs.
Hoogs virulente rekombinante PVY isolate, sowel as sommige nie-rekombinante rasse, kan aartappel
nekrotiese ring simptome (PTNRD) veroorsaak wat verliese van 10% tot totale misoes tot gevolg kan hê.
Om hierdie rede was die ondersoek van infeksie deur plaaslike rekombinante isolate op plaaslike
kultivare belangrik. Vir hierdie doel is 'n verteenwoordigende aantal isolate gekies, gebaseer op die
relatiewe voorkoms daarvan in Suid-Afrika, vir heelgenoom-volgordebepaling. Van die isolate is
vervolgens gebruik om plaaslike kultivare te besmet ten einde die invloed van genetiese variasie binne die
virale genoom op simptoom uitdrukking te ondersoek.
In hierdie studie is 27 heelgenoomvolgordes van Suid-Afrikaanse PVY isolate bepaal deur oorvleuelende
RT-PCR fragmente. Sewe van hierdie isolate, ses PVY NTN en een PVY N-W, is gebruik om vier plaaslike
aartappel kultivare, gegroei onder kweekhuis kondisies, meganies te infekteer. Die geïnfekteerde plante is
gemonitor om die tempo van sistemiese verspreiding vas te stel deur middel van 'n hoogs sensitiewe qRTPCR
en knolle is visueel inspekteer vir PTNRD. Hoogs variante rekombinante isolate blyk om minder
virulent te wees as die meer bewaarde rekombinante isolate wat dui op molekulêre determinante van
patogenisiteit. Om hierdie rede is die aminosuurvolgordes van die Suid-Afrikaanse isolate vergelyk met
die van internasionale isolate en ondersoek vir variasie en substitusies. Sommige Suid-Afrikaanse isolate
vertoon aminosuur substitusies wat uniek is tot die spesifieke isolaat en maak hul dus anders as
internasionale isolate. Die aantal aminosuursubstitusies in die volgordes verskil grootliks. In vergelyking
met internasionale isolate toon sommige isolate skaars enige veranderinge terwyl ander ‘n aantal verskille
toon. Sekere gebiede, waarvan baie spesifieke funksies het, was meer gekonserveerd as ander.
Hierdie studie ondersoek ook rekombinasie gebeure binne die PVY genoom deur retikulêre filogenetiese
analise, molekulêre datering en netwerk konstruksie tegnieke. In teenstelling met bestaande benaderinge,
aanvaar die tegniek wat hier beskryf word nie ‘n streng bifurkeerende filogenie, wat onderliggende
verdeel, of enige voorafgaande kennis van die prosesse onderliggend aan afwykings tussen individuele
filogenieë nie. ‘n Robuuste, tyd gekalibreer filogenie kan diversifikasie patrone en rekombinasie van PVY
plaas in die historiese konteks van menslike verbouing van aartappels. Deur gebruik te maak van hierdie
tegnieke poog die studie om te toets of diversifikasie en rekombinasie van PVY rasse plaasgevind het
binne die tydsbestek van die inburgering en moderne verbouing van aartappels. Van hierdie ontledinge
word afgelei dat rekombinante rasse van PVY wat in Suid-Afrika voorkom, ingevoer is.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/71637 |
| Date | 12 1900 |
| Creators | Visser, Johan Christiaan |
| Contributors | Bellstedt, D. U., Stellenbosch University. Faculty of Science. Dept. of Biochemistry. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | Unknown |
| Type | Thesis |
| Rights | Stellenbosch University |
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