Oropharyngeal candidiasis (OPC, thrush) is the term given to opportunistic oral infection caused by the yeast Candida, and is the most common mycotic complication associated with human immunodeficiency virus (HlV)-infected patients. The ability of Candida isolates to form biofilm on devices and host tissue surfaces is believed to be intricately linked with OPC infections and denture stomatitis. Therefore, understanding the role of biofilms in pathogenesis and host-Cand/cfa interactions is critical. In this application, to gain insight into these interactions, we will use a proteomics-based approach to identify specific proteins that are central to the biofilm forming ability of C. albicans and determine their role in interactions between fungal biofilm and host tissues. We have previously established an in vitro model of C. albicans denture biofilm (Publication 1, Appendix). Using this model, we: 1) identified the developmental phases of candidal biofilm formation (Publication 1, Appendix), 2) investigated the antifungal resistance profile of C. albicans biofilms at different growth phases (Publication 2, Appendix), 3) investigated the multifactorial mechanisms of azole resistance of early and mature biofilms formed by C. albicans (Publication 3, Appendix), 4) used a proteomic approach to identify potential target proteins that are differentially expressed in early phase biofilms (see Publication 4, Appendix), 5) used molecular and biochemical methods to show that one of the identified proteins (alcohol dehydrogenase, Adhlp) is a negative regulator of Candida biofilm (Publication 4, Appendix), and 6) moved our studies closer to the clinical setting by using an engineered human oral mucosa (EHOM), developed recently by Dr. Rouabhia (Co-investigator on this application). This collaborative research between the applicant and Dr. Rouabhia showed that Adhlp plays an important role in both Candida biofilm formation and invasion of host mucosal tissues (Manuscript submitted, Appendix). The overall hypothesis of this application is that C. albicans express specific proteins that are essential for biofilm formation and play critical roles in Candida-host tissue interactions. We will test our hypothesis using the following Specific Aims:
Aim 1. Identify early phase biofilm-specific proteins expressed by C. albicans.
Aim 2. Determine whether the identified proteins are critical to the ability of C. albicans to form biofilms in vitro by disrupting genes encoding them.
Aim 3. Use an in v/Vo-like Engineered Human Oral Mucosa (EHOM) Model to determine whether the identified proteins are essential for Candida biofilm formation and host tissue damage.
Aim 4. Validate the in vitro and EHOM results in vivo using a murine oral model of Candida biofilms. Experiments described in this application will provide insight into the biology of OPC-associated C. albicans biofilms and may suggest potential therapeutic and diagnostic targets for the prevention, treatment, and diagnosis of oral Candida infections. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
1R01DE017486-01A1
Application #
7199318
Study Section
Oral, Dental and Craniofacial Sciences Study Section (ODCS)
Program Officer
Rodriguez-Chavez, Isaac R
Project Start
2007-05-15
Project End
2012-04-30
Budget Start
2007-05-15
Budget End
2008-04-30
Support Year
1
Fiscal Year
2007
Total Cost
$388,382
Indirect Cost
Name
Case Western Reserve University
Department
Dermatology
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Ghannoum, Mahmoud A; Mukherjee, Pranab K; Jurevic, Richard J et al. (2013) Metabolomics reveals differential levels of oral metabolites in HIV-infected patients: toward novel diagnostic targets. OMICS 17:5-15
Rouabhia, Mahmoud; Semlali, Abdelhabib; Chandra, Jyotsna et al. (2012) Disruption of the ECM33 gene in Candida albicans prevents biofilm formation, engineered human oral mucosa tissue damage and gingival cell necrosis/apoptosis. Mediators Inflamm 2012:398207
Nweze, Emeka I; Ghannoum, Adam; Chandra, Jyotsna et al. (2012) Development of a 96-well catheter-based microdilution method to test antifungal susceptibility of Candida biofilms. J Antimicrob Chemother 67:149-53
Ghannoum, M A; Herbert, J; Isham, N (2011) Repeated exposure of Candida spp. to miconazole demonstrates no development of resistance. Mycoses 54:e175-7
Rouabhia, Mahmoud; Mukherjee, Pranab K; Lattif, Ali Abdul et al. (2011) Disruption of sphingolipid biosynthetic gene IPT1 reduces Candida albicans adhesion and prevents activation of human gingival epithelial cell innate immune defense. Med Mycol 49:458-66
Lattif, Ali Abdul; Mukherjee, Pranab K; Chandra, Jyotsna et al. (2011) Lipidomics of Candida albicans biofilms reveals phase-dependent production of phospholipid molecular classes and role for lipid rafts in biofilm formation. Microbiology 157:3232-42
Tuttle, Marie S; Mostow, Eliot; Mukherjee, Pranab et al. (2011) Characterization of bacterial communities in venous insufficiency wounds by use of conventional culture and molecular diagnostic methods. J Clin Microbiol 49:3812-9
Jurevic, R J; Traboulsi, R S; Mukherjee, P K et al. (2011) Identification of gentian violet concentration that does not stain oral mucosa, possesses anti-candidal activity and is well tolerated. Eur J Clin Microbiol Infect Dis 30:629-33
Nweze, E I; Mukherjee, P K; Ghannoum, M A (2010) Agar-based disk diffusion assay for susceptibility testing of dermatophytes. J Clin Microbiol 48:3750-2
Leal Jr, Sixto M; Cowden, Susan; Hsia, Yen-Cheng et al. (2010) Distinct roles for Dectin-1 and TLR4 in the pathogenesis of Aspergillus fumigatus keratitis. PLoS Pathog 6:e1000976

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