Candida albicans and C. dubliniensis are two closely related pathogenic yeast
species, sharing many phenotypic characteristics which make it difficult to
differentiate them, especially in clinical samples. As a result of the similarities
between these species, identification techniques, based on phenotypic
characteristics, have been developed. In this study some of these techniques
and virulence factors were used to characterise strains belonging to these
species and to select phenotypically dissimilar strains for further study. This
was performed to evaluate if the effect of arachidonic acid (20:4) on these
strains were the same, even though they are phenotypically different. Candida
albicans and C. dubliniensis can form biofilms which play an important role
during infection. During C. albicans infection, 20:4, a long-chain
polyunsaturated fatty acid (PUFA), derived from the phospholipids (PLs) of the
infected host cell membrane, serves as carbon source and precursor for
eicosanoid production. Conflicting results are presented in literature regarding
the effect of 20:4 on morphogenesis in C. albicans. In addition, the effect of
20:4 on growth and morphology of C. dubliniensis is unknown. Microscopic
examination and enzyme activity assay indicated that 1 mM 20:4 had little to
no effect on growth and metabolic activity of planktonic cells and biofilms, as
well as on the morphology and viability of the cells in the biofilms. The uptake
of PUFAs by yeasts is necessary for utilisation as metabolic fuels, cellular
building blocks and the production of signalling molecules. However, there are
no definitive studies regarding the uptake and cellular metabolism of 20:4 by
these pathogenic yeasts. The uptake and incorporation of 20:4 by planktonic
cells and biofilms of selected strains of C. albicans and C. dubliniensis were
evaluated by subjecting residual and cellular lipids from planktonic cells and
biofilms, grown in the presence and absence of 20:4, to gas chromatography
and gas chromatography-mass spectrometry. Strain specific variation in 20:4
uptake and incorporation into different lipid fractions of planktonic cells and
biofilms were found. In addition, eicosanoids produced by biofilms in the
presence of 20:4 were extracted and it was found that biofilms of these strains
were capable of producing 3-hydroxy fatty acids from 20:4. Arachidonic acid
can be incorporated into the PLs of yeasts, influencing saturation in cell
membranes. It is suggested that the effectiveness of antifungals may depend upon the level of unsaturation and ergosterol content of the membrane,
therefore the effect of 20:4 on the cell membrane and susceptibility of C.
albicans and C. dubliniensis biofilms towards amphotericin B and clotrimazole
were also determined. This was performed by confocal laser scanning
microscopy, determination of mitochondrial metabolism, unsaturation index of
the PL fractions and ergosterol content of the membranes of biofilms grown in
the presence and absence of 20:4. The results indicated that 20:4 influences
PL unsaturation and ergosterol content of both C. albicans and C. dubliniensis
type strains, increasing susceptibility to the antifungals. Pre-treatment of
biofilms with PUFAs may result in the reduction in antifungal dose needed to
inhibit biofilms.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ufs/oai:etd.uovs.ac.za:etd-10122009-095911 |
| Date | 12 October 2009 |
| Creators | Ells, Ruan |
| Contributors | Prof JLF Kock, Dr CH Pohl |
| Publisher | University of the Free State |
| Source Sets | South African National ETD Portal |
| Language | en-uk |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.uovs.ac.za//theses/available/etd-10122009-095911/restricted/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University Free State or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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