Thesis (MScAgric)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Botrytis cinerea Pers. Fr. [teleomorph Botryotinia fuckeliana (de Bary) Whetzel] causes
serious losses of over 200 crops worldwide, including rooibos seedlings and pears. This
pathogen is characterized by morphological, physiological and genetic diversity. The genetic
diversity and population structure have not been investigated for B. cinerea populations in
South Africa. Botrytis cinerea collected from rooibos seedlings and in pear orchards in the
Western Cape of South Africa were investigated in the present study. The study was done
with the aid of microsatellite markers, the amplification of mating type alleles MAT1-1 and
MAT1-2 and determination of resistance towards various fungicides. Population dynamics
was inferred and a similar picture emerged in both production systems.
Botrytis cinerea annually causes severe losses of rooibos seedlings (Aspalathus
linearis) in nurseries situated in the Clanwilliam region. Sampling was done in five nurseries
and the cryptic species status of the isolates obtained was determined through restriction
enzyme digestion of the Bc-hch gene. All but one (206 out of 207) of the isolates belonged to
Group II or B. cinerea ‘sensu stricto’. Analysis of the B. cinerea Group II population, using
seven microsatellite loci, was performed to assess the genetic population structure. Total
gene diversity (H) was high, with a mean of 0.67. Two of the nurseries populations’ sample
sizes were severely limited after clone correction, yet 100 genotypes were discerned among
the 206 isolates genotyped. The percentage of maximal genotypic diversity (G) ranged
between 16 and 68 for the five populations, with a total value of 17 for the 100 genotypes.
One genotype, represented by 27 clones, was isolated from four nurseries. Relatively low but
significant population differentiation was observed in total between nurseries (mean FST =
0.030, P = 0.001). The distribution of mating types MAT1-1 and MAT1-2 differed significantly
from the ratio of 1:1 for the total population plus two of the nurseries’ populations. Three
nursery populations had an equal mating type distribution. The index of association (IA)
analyses suggests that the populations are asexually reproducing. Analysis of molecular
variance (AMOVA) indicated that 97% of the total genetic variation is distributed within
subpopulations. Fungicide resistance frequency against iprodione for 198 of the genotyped
isolates displayed highly varying levels of resistance amongst the five nurseries. The mean
total incidence of resistance towards iprodione was 43%, ranging from 0% to 81% for the five
nurseries. Baseline sensitivity towards pyrimethanil yielded an average EC50 value of 0.096
mg/L.
Botrytis cinerea isolates were collected from pear blossoms (Pyrus communis) in four
orchards. Two orchards in the Ceres area and two in the Grabouw area were sampled from.
A total of 181 isolates were collected from the four orchards. Incidence of blossom infection
in the orchards ranged from 3% to 17%. Overall, there was a high incidence of isolates that
had only the Boty transposable element (74%) compared to those harbouring both (Boty and
Flipper), simultaneously (transposa, 24%). One isolate examined had the Flipper element only. Cryptic species status according to restriction enzyme digestion of the Bc-hch gene
indicated that all the isolates belonged to Group II or B. cinerea ‘sensu stricto’. Analysis of
the Group II population, through the use of seven microsatellite loci, was performed to
assess the genetic population structure. Total gene diversity (H) was high, with a mean of
0.69 across all populations. Although two of the subpopulations displayed a high clonal
proportion, overall 91 genotypes were discerned among the 181 isolates. The percentage of
maximal genotypic diversity (G) ranged between 18 and 33 for the four populations, with a
total value of 14 for the 91 genotypes. One genotype, represented by 27 clones, was isolated
from all orchards. Moderate, but significant population differentiation was present in total
among orchards (mean FST = 0.118, P = 0.001). The distribution of the mating types, MAT1-1
and MAT1-2, did not differ significantly from a 1:1 ratio for the total population as well as the
subpopulations. Index of association (IA) analyses, on the other hand, suggests that the
populations reproduce asexually. Analysis of molecular variance (AMOVA) indicated that
88% of the total genetic variation is distributed within subpopulations, 9% between
subpopulations and only 3% between production areas. Fungicide resistance frequency
against fenhexamid, iprodione and benomyl varied, with the highest levels of resistance
present against benomyl and low levels of resistance seen towards iprodione and
fenhexamid.
In conclusion, this study has shown that there exist within the studied populations of
B. cinerea, obtained from rooibos nurseries and pear orchards, an adaptive capacity to
overcome current means of control. The use of population genetics to further our
understanding of how plant pathogens interact and spread throughout a given environment is
of cardinal importance in aiding the development of sustainable and integrated management
strategies. Knowledge of the dispersal of B. cinerea in the two studied cropping systems has
shed light on the inherent risk that it poses, and this together with knowledge of the levels of
resistance that occurs should serve as an early warning to help divert possible loss of control
in future. / AFRIKAANSE OPSOMMING: Botrytis cinerea Pers. Fr. [teleomorf Botryotinia fuckeliana (de Bary) Whetzel] veroorsaak
ernstige verliese van meer as 200 gewasse wêreldwyd, insluitende rooibossaailinge en pere.
Hierdie patogeen word deur morfologiese, fisiologiese, asook genetiese diversiteit
gekenmerk. Die genetiese diversiteit en populasie-struktuur van B. cinerea populasies wat in
Suid-Afrika voorkom, is nog nie ondersoek nie. Botrytis cinerea verkryg vanaf
rooibossaailinge en in peerboorde in die Wes-Kaap van Suid-Afrika is ondersoek. Hierdie
studie is met behulp van mikrosatellietmerkers, amplifikasie van die twee paringstipe gene
(MAT1-1 en MAT1-2), asook die bepaling van weerstandsvlakke teenoor verskeie
swamdoders, uitgevoer. Populasie-dinamika is afgelei en ‘n soortgelyke tendens is in beide
produksie-sisteme waargeneem.
Botrytis cinerea veroorsaak jaarliks ernstige verliese van rooibossaailinge
(Aspalathus linearis) in kwekerye in die Clanwilliam-area. Monsters is in vyf kwekerye
versamel en die kriptiese spesiestatus van die verkrygde isolate is deur restriksie-ensiemvertering
van die Bc-hch geen bepaal. Almal behalwe een (206 uit 207) isolaat het aan
Groep II of B. cinerea ‘sensu stricto’ behoort. Analise van die B. cinerea Groep II populasie,
deur middel van sewe mikrosatellietmerkers, is uitgevoer om die genetiese populasiestruktuur
te bepaal. Totale geendiversiteit (H) was hoog, met ‘n gemiddelde van 0.67.
Alhoewel twee van die kwekerye se monstergrootte erg ingeperk is ná kloonverwydering, is
daar nogtans 100 genotipes onder die 206 isolate wat geïsoleer is, waargeneem. Die
persentasie van maksimale genotipiese diversiteit (G) het tussen 16 en 68, vir die vyf
populasies, gewissel, met ‘n totaal van 17 vir die 100 genotipes. Een genotipe,
verteenwoordig deur 27 klone, is uit vier kwekerye geïsoleer. Relatief lae dog
noemenswaardige populasie-differensiasie is in totaal tussen kwekerye waargeneem (gem.
FST = 0.030, P = 0.001). Die verspreiding van die twee paringstipes (MAT1-1 en MAT1-2) het
beduidend verskil van ‘n 1:1 verhouding vir die totale populasie, asook twee van die
kwekerye se populasies. Die drie oorblywende kwekerye se populasies het egter ‘n gelyke
verdeling van die twee paringstipes getoon. Die indeks van assosiasie (IA) analises toon dat
die populasies ongeslagtelik voortplant. Analise van molekulêre variasie (AMOVA) het
aangedui dat 97% van die totale genetiese variasie binne die subpopulasies versprei is.
Hoogs variërende vlakke van weerstand tussen die vyf kwekerye teenoor die swamdoder
iprodioon, is vir die 198 isolate wat getoets is, gevind. Die totale gemiddelde frekwensie van
weerstand teenoor iprodioon was 43%, wat tussen 0% en 81% vir die vyf kwekerye gevarieer
het. Fondasie-vlak-sensitiwiteit vir pyrimethanil het ‘n gemiddelde EC50 waarde van 0.096
mg/L opgelewer.
Botrytis cinerea isolate is ook vanuit peerbloeisels (Pyrus communis L.) vanuit vier
boorde versamel, twee uit elk van die Ceres- en Grabouw-areas. In totaal is 181 isolate vanuit die vier boorde versamel. Die frekwensie van bloeiselinfeksie het tussen 3% en 17%
gewissel. Oor die algemeen was daar ‘n hoë frekwensie van isolate wat slegs die Boty
transponeerbare element teenwoordig gehad het (74%) in vergelyking met dié wat
tegelykertyd beide (Boty en Flipper) teenwoordig gehad het. Een isolaat het slegs die Flipper
element gehad. Bepaling van die kriptiese spesiestatus met behulp van restriksie-ensiemvertering
van die Bc-hch geen het aangedui dat alle versamelde isolate tot Groep II of B.
cinerea ‘sensu stricto’ behoort het. Analise van die Groep II populasie, deur middel van sewe
mikrosatellietmerkers, is uitgevoer om genetiese populasie-struktuur te bepaal. Totale
geendiversiteit (H) was hoog, met ‘n gemiddelde van 0.69 oor alle populasies. Alhoewel twee
subpopulasies ‘n hoë klonale fraksie getoon het, is 91 genotipes tussen die 181 isolate wat
verkry is, onderskei. Die persentasie van maksimale genotipiese diversiteit (G) het tussen 18
en 33 vir die vier populasies gewissel, met ‘n totale waarde van 14 vir die 91 genotipes. Een
genotipe, verteenwoordig deur 27 klone, was in al vier boorde teenwoordig. Gematigde dog
beduidende populasie differensiasie was in totaal tussen boorde teenwoordig (gem. FST =
0.118, P = 0.001). Die verspreiding van die paringstipes (MAT1-1 en MAT1-2) het nie
betekenisvol van ‘n 1:1 verhouding vir die totale populasie, insluitende die subpopulasies,
verskil nie. Indeks van assosiasie (IA) analises het egter aangedui dat die populasies
ongeslagtelik voortplant. Analise van molekulêre variasie (AMOVA) het aangedui dat 88%
van die totale genetiese variasie in subpopulasies te vinde was, 9% tussen subpopulasies en
slegs 3% tussen produksie-areas. Frekwensie van swamdoder weerstandbiedendheid vir
fenhexamid, iprodioon en benomyl het gewissel, met die hoogste vlakke teenoor benomyl
waargeneem, maar baie lae vlakke teenoor fenhexamid en iprodioon.
Samevattend het hierdie studie getoon dat die populasies van B. cinerea wat in
hierdie twee produksie-sisteme, op rooibossaailinge en in peer boorde, ondersoek is, ‘n
aanpasbaarheid toon om huidige metodes van beheer te oorkom. Die gebruik van populasiegenetika
as ‘n hulpmiddel om ons kennis van patogeen-interaksies en -verspreiding te
verbreed, is van kardinale belang in die ontwikkeling van geïntegreerde en volhoubare
beheermaatreëls. Kennis van die verspreiding van B. cinerea in die bestudeerde
gewasproduksiestelsels, werp lig op die inherente risiko wat dié patogeen inhou. Dít, tesame
met kennis van die weerstandsvlakke wat voorkom, kan as ‘n vroegtydige waarskuwing dien
ten einde moontlike verlies van beheer in die toekoms te help teenwerk.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/20070 |
| Date | 03 1900 |
| Creators | Wessels, Andries Bernardus |
| Contributors | Mostert, Lizel, Stellenbosch University. Faculty of AgriSciences. Dept. of Plant Pathology. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
| Format | 95 p. : ill. |
| Rights | Stellenbosch University |
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