In Vitro Studies on Candida, Antimycotics and Oral Defences

This dissertation, "In Vitro Studies on Candida, Antimycotics and Oral Defences" by Sukumaran, Anil, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Abstract of thesis entitled In vitro studies on Candida, antimycotics and oral defences Submitted by Sukumaran Anil for the degree of Doctor of Philosophy at The University of Hong Kong in October 2002 Abstract There is scant information on the effect of exposure to low dilutions of antifungal agents on the virulence attributes of the human fungal pathogen Candida, such as its cell surface hydrophobicity, and phospholipase and haemolysin production. Further, the impact of host mucosal antifungal proteins such as lactoferrin and lysozyme on drug-exposed Candida species has been little studied. In order to obtain basic data on the latter aspects of Candida, in vitro evaluation of the post-antifungal effect (PAFE) of two polyenes (nystatin and amphotericin B), two-azoles (fluconazole and ketoconazole) and a single DNA-analogue (5- fluorocytosine) against ten oral isolates each of Candida albicans and Candida tropicalis, all from human immunodeficiency virus infected individuals, was performed. The term PAFE is given to the suppression of fungal growth that persists after limited exposure to a low dilution of an antifungal agent. One-hour exposure to twice the minimum inhibitory concentration of all the drugs, except fluconazole, elicited a consistently high PAFE in both Candida species. Furthermore, the PAFE elicited by the antifungals (except fluconazole) was significantly prolonged for C. tropicalis compared with C. albicans. The speedy recovery of C. albicans isolates exposed to transient, low concentrations of antifungals appears to be a reflection of its higher virulence. This data, while confirming the existence of PAFE in a non-albicans species of Candida, provided further clues for the recalcitrance of C. albicans infections in the face of antifungal therapy. The cell surface hydrophobicity (CSH) of Candida is considered a critical factor contributing to its colonization potential and virulence. Comparison of the CSH of 1the aforementioned Candida isolates following their brief exposure to nystatin, amphotericin B, ketoconazole, fluconazole and 5-fluorocytosine showed that the CSH of C. albicans is reduced to a significant extent when exposed to the antifungals, illustrating another mode of action of antifungals on Candida species that modify their colonization potential. On investigating the in vitro phospholipase activity of Candida exposed to the foregoing antifungals, it was evident that C. albicans continued to produce higher levels of phospholipase than C. tropicalis. Furthermore, pre-exposure of all the isolates to antimycotics led to a significant reduction in the phospholipase activity. Similarly, all studied Candida isolates exposed to nystatin, amphotericin B and fluconazole showed a significant reduction in haemolysin production. This reduced haemolytic activity, evaluated using a novel plate assay, was maximum for nystatin- exposed yeasts followed by amphotericin B- and fluconazole-exposed Candida. The antifungal effect of mucosal defense proteins lysozyme and lactoferrin on drug-exposed Candida species is not known. Following brief exposure to sub- therapeutic concentrations of the polyenes and fluconazole a significant increased yeast-susceptibility to lysozyme, but not to lactoferrin, was noted. Polyenes had a lesser impact on the lysozyme susceptibility of yeasts, compared with the azoles. Taken together, these data shed light on the mechanisms by which