Browsing by Subject "Candida albicans"
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Item Biochemical characterization of cell surface components of Candida albicans(Texas Tech University, 1982-12) Mendel, Susan Mari NegaardExtracellular material (ECM) was isolated from media in which yeast phase Candida albicans of unknown serotype was grown. Cold 95% ethanol precipitated the ECM and upon Sephadex G-50 chromatography, high and low molecular weight components were identified. They consisted of ECM 1 (30,000 Mr), ECM 2A (4,500-6,000 Mr), and ECM 2B (3,800 Mr). The ECM 2B was separated into two components upon ion exchange chromatography. Glucose and mannose are its carbohydrate moieties and proline, lysine and glutamic acid are its major amino acids. It appears to have more protein content than ECM 2B2 as evidenced by absorbance at 2 80 nm. ECM 2B2 contains only mannose and some protein composed mostly of proline, threonine and alanine. The components are neither strain-specific, nor produced only during yeast phase growth. The ECM were compared to material released from the cell surface upon treatment of whole cells with the nonspecific protease papain, and the disulfide bond reducer, dithiothreitol. An ECM 2A is produced upon both treatments. ECM 2B1 closely resembles a low molecular weight component produced upon DTT treatment, and ECM 2B2 is similar to the papain released molecule. The ECM appears to be present either on or within the cell surface of Candida albicans. The ECM did not appear to be strongly antigenic, although unpurified media did induce an antibody response.Item Characterization of cobalmin-independent methionine synthase from Candida albicans and Saccharomyces cerevisiae(2006) Suliman, Huda Sirageldin; Robertus, Jon D.Item In Vitro Evolutionary Dynamics of C. albicans during Adaptation to Fluocnazole(2012-10-19) Huang, MianMany drug-resistant mechanisms in Candida albicans (C. albicans), a clinical important fungal pathogen, have been well characterized. However, few studies investigated the emergence of drug resistance from the evolutionary perspective and little is known about the evolutionary trajectories during the adaptation to the drug. Here, we examined the evolutionary dynamics of C. albicans both in the presence and absence of fluconazole, a first line drug, using the visualizing evolution in real-time (VERT) method. Evolutionary dynamics of replicate C. albicans populations, either in the presence or absence of fluconazole, were determined and adaptive mutants arose in the populations were systematically isolated using the VERT method. Drug susceptibility assays were performed to measure the fluconazole minimum inhibitory concentration (MIC) for the adaptive isolates from drug-exposed populations. Analysis of the evolutionary dynamics revealed that mutations arose more frequently in the presence of the drug compared to the absence of the drug and the drug-resistant mutations occurred in independent lineages, suggesting a heterogeneous nature of the populations during the adaptation. In addition, fitness effects were evaluated for each adaptive mutant both in the presence and absence of drug and we found most of them gained significant increase in the drug resistance without a fitness cost in the absence of the drug. Interestingly, the aneuploidy and gross chromosomal rearrangements, common drug-resistant mechanisms, were not responsible for the increased resistance to fluconazole of most adaptive isolates, suggesting single-nucleotide polymorphisms (SNPs) or other stable unknown chromosomal rearrangements may contribute to the increased drug resistance.Item Influence of inhibitors of sterol biosynthesis on growth and sterol metabolism in Candida albicans(Texas Tech University, 2003-08) Nguyen, Thi Thuy MinhCandida albicans was treated with seven antifungal agents designed to inhibit ergosterol biosynthesis. Two types of inhibitors were studied based on their hypothetical mode of action: mechanism-based inactivators, containing either a sulfur or bromine in the sterol side chain and transition state mimics, which contain a nitrogen in the sterol side chain or in the nucleus. The sterol composition of cells treated with the various compounds was determined using three chromatographic techniques, thin-layer chromatography, reverse-phase high-performance liquid chromatography and gas chromatography-mass spectrometry. Additional characterization of the sterol identities was by proton nuclear magnetic resonance spectroscopy and ultra-violet spectroscopy. Ergosterol was found to be the major sterol in control cultures (69%). In treated cells, the dominant effect on sterol biosynthesis was a significant decrease in the amount of ergosterol. The influence of containing a heteroatom in the side chain on sterol methyl transferase activity was evident in aberrant cell morphology of treated cells and a reduction in ergosterol production. Disturbance of the 24-alkyl to 24-desaikyi ratio by as little as 10 % is sufficient to elicit a fimgistatic response. A second class of inhibitor represented by 3p-aminoianosteroi, designed to inhibit the C-4 demethyiation pathway, was found to inhibit growth by impairing lanosterol metabolism. The relationship between ergosterol biosynthesis inhibition action on fungal growth and ergosterol homeostasis is discussed.Item Peroxidase-mediated oxygenation and microbicidal activity(Texas Tech University, 1999-05) Vigerust, David JohnIt is well documented that a peroxidase, H2O2, and a halide form a "cytotoxic triad." As a result of the interactions of the components of the triad, reactive oxygen intermediates (ROI) are formed and these ROI help to destroy various invading pathogens including Candida. The present study was undertaken to determine if equivalent units of peroxidase activity also induced equivalent macrophage-mediated killing of Candida. Peritoneal macrophages were obtained from age matched C57BL/6J mice and exposed to various concentrations of eosinophil peroxidase (EPO), myeloperoxidase (MPO), and horseradish peroxidase (HRP). Equivalent units of peroxidase as determined by oxidation of guaiacol, did not induce equivalent production of ROI. Luminol-dependent chemiluminescence studies indicated that 10 units of EPO induced more ROI than either HRP or MPO. Candidicidal activity and phagocytosis of M(t) was measured using a fluorescence acridine orange phagocytosis assay. The following pattern EPO>MPO>HRP emerged for both assays. Therefore, enzymatic activity does not directly correlate with candidicidal activity. These data indicate a distinct order of peroxidases relative to their ability to stimulate chemiluminescence and macrophage-mediated killing.Item Phagocytic and candidacidal activity of murine peritoneal macrophages(Texas Tech University, 1995-05) Gelderman, Monique PascaleMacrophages (Mo) are involved in host defenses against opportunistic pathogens. Previous studies reported that Mo exposed to enzymatically active myeloperoxidase (MYPO), exhibited both increased phagocytosis and killing of Candida albicans (C. albicans). The purpose of tiiis study was to detamine if enzymatically inactive MMR ligands could fimction similarly. Resident murine peritoneal Mo were exposed to tiie MMR ligands, mannosylated bovine serum albumin (mBSA) and enzymatically inactive myeloperoxidase (iMYPO), followed by exposure to opsonized C. albicans. Both mBSA and LMYPO enhanced phagocytosis and killing of C albicans over that of the control. The production of reactive oxygen intermediates (ROI) was detected using chemiluminescence. After employment of ROI scavaigers, a decrease in candidicidal activity was observed. The data suggest that MMR-ligand interaction enhances both phagocytosis and candidicidal activity of Mo and that MYPO and iMYPO released at a site of inflammation mduce Momediated candidicidal activity.Item Synthesis and testing of sterol biosynthesis inhibitors with Candida albicans(Texas Tech University, 2002-12) Liu, JialinA series of lanosterol analogs which contain a heteroatom substituted for C-24 or C-25 or otherwise modified in the double bond character in the sterol side chain at C-24 (25) have been prepared and assayed with Candida albicans as antifungal agents. 25- Azalanosterol and 24(R, S), 25- piminolanosterol were shown to be potent inhibitors of C. albicans growth, whereas 24-bromolanosterol, 24(28)-methylene lanosterol, 26,27- dehydrolanosterol, 4,4,14a-trimethylcholesta-8,24,26-trien-3P-ol, and 4,4,14atrimethylcholesta- 8, 24-dien-26-yne-3P-ol had no effect on growth. In the case of the ammonium-containing 25-azalanosterol and 24(i?, S), 25-epiminolanosterol, the IC50 was determined to be ca. 2.5 i^M. Eleven sterols were isolated from growth-arrested cells cultured aerobically. The identities of these compounds were established by a combination of gas chromatography; mass spectroscopy; proton nuclear magnetic resonance spectroscopy and ultra-violet spectroscopy. Ergosterol was found to be the major sterol in control cultures (63%). The sterol composition of the treated cells was found to accumulate 24-desalkyl sterol with a corresponding decrease in ergosterol. The total sterol of treated cells was increased by a factor of two compared to the wild-type control cultures. The results are interpreted to imply that the shift in the type and amount of sterol produced by treated cultures disturbs carbon flux and ergosterol homeostasis generating cell death.