Magister Chirurgiae Dentium (MChD) / The rate at which the population is ageing is much faster than in the past. An increase in age results in an increase in oral diseases. One of the most common types of oral diseases in the elderly are fungal infections caused by Candida albicans. It has been noted that drug resistance to fungal pathogens is developing into a serious threat to public health and healthcare systems worldwide. This has consequently led to the need to develop effective and innocuous treatment modalities. The purpose of this study was to explore the antimicrobial and cytotoxic potential of silver nanoparticles (AgNPs) synthesised from Berzelia lanuginose, Helichrysum cymosum, and Searsia crenata.

Objectives: Candida glabrata is the second most common cause of oral candidiasis, second only to C. albicans. Incidence of antifungal resistance has shown a steady increase for C. glabrata. Iron has shown to modulate C. albicans pathogenesis and affect drug-susceptibility. Here, we assess the effect of iron on the growth, antifungal-susceptibility, biofilms, and cell wall of C. glabrata.Methods: Growth, minimal inhibitory concentration (MIC), and biofilm experiments were conducted using 96-well polystyrene plates. Yeast Nitrogen Base medium was used for growth experiments. Cultures of C. glabrata and C. albicans were grown over two nights in respective media containing varying iron concentrations. Rosewell Park Memorial Institute medium was used for MIC and biofilm experiments. Serial dilution was performed to obtain desired concentrations of antifungal drugs. For all experiments, growth was assessed at OD600nm over 24 hours using BioTek Synergy Multi Mode Reader. Paraformaldehyde treated cells and specific stains were used for cell wall studies. Results: Growth of C. glabrata declined significantly below 5μM iron, while C. albicans continued to grow at decreasing iron concentrations, up to 0.5μM. MIC experiments revealed 1.562μM, 1.562μM, and 4μM, as the MIC for Deferasirox, Nystatin, and Fluconazole, respectively. Drug synergy experiments revealed a 128-fold reduction in the amount of Nystatin and Fluconazole needed, with the addition of 1/8th of Deferasirox concentration. The biofilm experiments were inconclusive and the cell wall studies showed decreased levels of mannan, chitin, and an increased β-glucan exposure in high iron conditions. Conclusion: C. glabrata is more sensitive to alterations in environmental iron when compared to C. albicans. Drug synergy experiments underscore the importance of Deferasirox in lowering the MICs of Nystatin or Fluconazole. This can allow use of classical antifungals at lower doses, thereby limiting their side effects. Cell wall studies discuss the effect of iron on the virulence of the C. glabrata. / Oral Biology

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Thesis (MSc)--Stellenbosch University, 2002. / ENGLISH ABSTRACT: The aim of this study at the Brooklyn Chest Hospital (BCH) was to assess the change in the oral carriage of Candida species in twenty-nine patients co-infected with the Human Immunodeficiency Virus (HIV) and Tuberculosis (TB), before and after anti-fungal treatment. Each patient accepted onto the study underwent a comprehensive oral and peri-oral examination where the presence, site and clinical features of all oral and peri-oral lesions were recorded. The purpose of the examination was to provide a clinical diagnosis of oral candidasis. Each patient was also asked to provide a sample of oral fluid for laboratory analysis. This was collected using an oral rinse. The results of a variety of laboratory investigations were used to identify the species of Candida obtained from the oral rinse. Both the oral and peri-oral examination and the oral rinse procedure were repeated after one month and at three months. A sample from each oral rinse was inoculated on CHROMagar Candida chromogenic medium (CHROMagar Candida, France, Paris). CHROMagar is used for the isolation and presumptive identification of Candida sp. from other yeasts on the basis of strongly contrasted colony colours, which are produced by the reactions of species-specific enzymes with a proprietary chromogenic substrate. After forty-eight hours the CHROMagar plate was examined for growth, when a record of colony morphology and colour was made. A single sample from each different colour-coded colony was taken and streaked onto a Sabouraud plate (Oxoid, Basingstake, England) and then incubated for forty eight hours at thirty-seven degrees centigrade. A variety of laboratory investigations were subsequently carried out on a single colony taken from the Sabouraud agar plate (Oxoid). The results of these tests were used to identify the individual species of Candida isolated from each oral rinse. Oral candidasis was the most prevalent oral lesion observed on admission and at three months. Six different species of Candida were identified during this study, namely Candida albicans, Candida dubliniensis, Candida krusei, Candida glabrata, Candida parapsilosis, and Candida tropicalis. C.albicans was the most commonly identified species in study population. Candida dubliniensis was isolated and identified for the first time in a South African HIV population. Each specimen of Candida sp. identified by laboratory analysis was tested for sensitivity to Nystatin, Amphotericin B and Fluconazole anti-fungal agents. An additional sensitivity test was performed using Ajoene and Allicin (extracts of garlic) to assess the comparative antifungal properties of these compounds. / AFRIKAANSE OPSOMMING: Die doelwit van hierdie studie by die Brooklyn Borshospitaal (BCH) was om die verandering in orale draerstatus van die Kandida spesies in nege-en-twintig HIVfTB koïnfekteerde pasiënte vas te stel, voor- en na antifungale behandeling. Elke pasiënt in die studie het 'n volledige intra- en ekstra-orale ondersoek ondergaan. Die teenwoordigheid, area en kliniese voorkoms van alle letsels is noteer. Die doel van die ondersoek was om 'n kliniese diagnose van orale kandidiase te verkry. 'n Monster orale vloeistof is geneem van elke pasiënt vir laboratorium analise. Die monster is in die vorm van 'n mondspoel geneem. Verskeie toetse is gedoen om die verskillende Kandida spesies in elke monster te identifiseer. Die orale- en ekstra-orale ondersoek sowel as die mondspoelmonster is na 1 en 3 maande herhaal. Elke mondspoelmonster is op CHROMagar Kandida chromogene medium (CHROMagar Candida, France) inokuleer. CHROMagar word gebruik vir die vermoedelike identifikasie en isolasie van Kandida spesies teenoor ander swamme. Dit word gedoen op die basis van kontrasterende koloniekleure, wat teweeggebring word deur spesie-spesifieke ensiemreaksies op 'n chromogene substraat. Die CHROMagar plate is na 48 uur ondersoek vir groei en die kolonie-morfologie en - kleur is noteer. 'n Enkel monster. is geneem van elke verskillende kolonie (geskei op kleur) en is uitgestreep op 'n Saboraud plaat (Oxoid, Basingstoke, England). Dit is dan vir 48 uur inkubeer teen 37°C. Verskeie laboratorium ondersoeke is daarna uitgevoer op 'n enkel kolonie geneem vanaf die Saboraud agar plaat (oxoid). Die resultate van die ondersoeke is gebruik om individuele spesies van Kandida te identifiseer. Orale Kandidiase was die mees algemene orale letsel geïdentifiseer by toelating en 3 maande ondersoeke. Ses verskillende spesies Kandida is identifiseer tydens die studie, naamlik: Kandida albicans, K.dubliniensis, K.Krusei, K.glabrata, K.parapsilosis en K.tropicalis. K.albicans was die mees algemeen identifiseerde spesie in die studiepopulasie. K.dubliniensis is vir die eerste keer in Suid-Afrika in 'n HIV<+lpopulasie isoleer en geïdentifiseer. Elke monster van identifiseerde Kandida spesies is getoets vir sensitiwiteit teenoor Nistatien, Amfotensien B en Flukonasool. Addisioneel is ook getoets vir sensitiwiteit teenoor Ajoene en Allicin (knoffelekstrakte).