Thesis (MSc)--University of Stellenbosch, 2006. / ENGLISH ABSTRACT: Fish infections caused by pathogenic Flavobacterium spp. are a major
problem in the aquaculture industry worldwide, often leading to large
economic losses. Thirty-two Flavobacterium spp. isolates, obtained from
various diseased fish species and biofilm growth, were characterised
genetically using 16S rRNA gene sequencing, 16S rRNA gene PCR
restriction fragment length polymorphism (RFLP), randomly amplified
polymorphic DNA (RAPD) PCR, repetitive extragenic palindromic (REP)
element PCR, plasmid profiling, whole cell protein (WCP) and outer
membrane protein (OMP) analyses. The biofilm-forming capability of five
genetically heterogeneous Flavobacterium spp. study isolates was
investigated using a modified microtiter-plate adherence assay, as well as
flow cell studies. Experimental infection studies with Mozambique tilapia
(Oreochromis mossambicus) were carried out in order to determine the
virulence of the Flavobacterium spp. study isolates. 16S rRNA gene
sequence analysis showed the Flavobacterium spp. study isolates were
closely related, and 97% sequence similarity was shared with published F.
johnsoniae sequences. A high degree of genetic heterogeneity was
displayed by the Flavobacterium spp. study isolates following RAPD-PCR,
REP-PCR and OMP analysis, however, based on the results obtained by
plasmid profiling and WCP analysis, the isolates appeared genetically very
homogeneous. The biofilm phenotype was displayed by all five
Flavobacterium spp. isolates tested and varied from weakly to strongly
adherent. No specific correlation was observed between the RAPD, REP
and/or OMP profiles and degree of adherence displayed by Flavobacterium
spp. isolates. However, a specific WCP profile (profile B), exhibited by 48% of
the Flavobacterium spp. isolates, was linked to strong adherence.
Experimental infection studies showed that Flavobacterium spp. isolates
displayed variable levels of virulence, which could not be linked to biofilm
formation, nor specific genotypes. This is the first reported isolation and
characterisation of Flavobacterium spp. isolated from diseased fish in
Southern Africa, and there appears to be significant diversity amongst the
isolates which is not geographically linked nor host related. / AFRIKAANSE OPSOMMING: Visinfeksies veroorsaak deur Flavobacterium spp. is problematies in die
akwakultuur industrie wêreldwyd en lei tot groot ekonomiese verliese. Twee
en dertig Flavobacterium spp. isolate, geïsoleer vanaf verskye geïnfekteerde
visspesies en biofilm groei, was geneties gekarakteriseer met behulp van 16S
rRNS geenvolgorde, 16S rRNS geen PKR restriksie fragment lengte
polimorfisme (RFLP), toevallig geamplifiseerde polimorfiese DNS (TGPD)
PKR, herhaalde ekstrageniese palindromiese (HEP) element PKR, plasmied
profilering, heelsel protein (HSP) en buite membraan protein (BMP) analise.
Die vermoë van vyf geneties heterogene Flavobacterium spp. isolate om
biofilms te vorm was ondersoek met behulp van ‘n gemodifiseerde
mikrotiterplaat vashegtings toets asook vloei-sel studies. Eksperimentele
infeksie studies was uitgevoer op bloukurpers (Oreochromis mossambicus)
om die virulensie van die Flavobacterium spp. studie isolate te toets. 16S
rRNS geenvolgorde analise het getoon dat die Flavobacterium spp. studie
isolate naby verwant was, en het 97% ooreenstemming getoon met
gepubliseerde F. johnsoniae volgordes. TGPD-PKR, HEP-PKR en BMP
analise het ‘n hoë graad van heterogeniteit tussen die Flavobacterium spp.
studie isolate aangetoon, egter, op grond van plasmied profilering en HSP
analise, was die studie isolate geneties baie homogeen. Die biofilm fenotipe
was getoon deur al die getoetsde Flavobacterium spp. isolate en het
gevarieer van swak tot sterk vashegting. Geen spesifieke korrelasie was
waargeneem tussen die TGPD, HEP en/of BMP profiele en graad van
vashegting vertoon deur Flavobacterium spp. isolate nie, maar ‘n spesifieke
HSP profiel (profiel B), getoon deur 48% van die Flavobacterium spp. isolate,
was verbind met sterk vashegting. Eksperimentele infeksie studies het
getoon dat Flavobacterium spp. isolate varierende grade van virulensie
vertoon het en wat met biofilm formasie of spesifieke genotipes geassosieer
kon word nie. Hierdie is die eerste gedokumenteerde isolasie en
karakterisering van Flavobacterium spp. geïsoleer van geïnfekteerde vis in
Suider Afrika, en daar is beduidende diversiteit tussen die isolate wat nie
geografies of gasheer geassosieerd is nie.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/17351 |
| Date | 04 1900 |
| Creators | Flemming, Leonard (Leonard Arnold) |
| Contributors | Chenia, H.Y., Rawlings, D.E., University of Stellenbosch. Faculty of Science. Dept. of Microbiology. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
| Format | xiv, 192 leaves : ill. |
| Rights | University of Stellenbosch |
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