Prof. Mahmoud Ghannoum, Director of Center for Medical Mycology, has been awarded the prestigious Kuwait Prize by the Kuwait Foundation for Advancement of Sciences for his contribution to scientific research in the field of fungal infections, pathogenesis, antifungal drug discovery, and preclinical evaluation of drugs.  Notification of this award was reported by newspapers including The Plain Dealer, and and the Hudson Hub Times. 

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A brief Autobiographical Sketch of Dr. Ghannoum follows below:

Major Research Interests:  fungal pathogenesis, antifungal drug discovery and preclinical evaluation

            In 1973, I received my Bachelor of Science degree from the American University of Beirut. Subsequently, I was awarded Master of Science (1974) and Doctor of Philosophy degrees (1978) from Loughborough University of Technology, England.  Prior to moving to the United States, most of my academic career was spent at the Department of Botany and Microbiology, Kuwait University, where I held a full-time faculty appointment from 1980 until 1991.  The invasion of Kuwait forced me to relocate to the United States, where I joined the Infectious Diseases Division, Department of Medicine, University of California at Los Angeles.  In 1996, I joined the Dermatology Department, Case Western Reserve University, Cleveland, Ohio, as an Associate Professor and Director for the Center for Medical Mycology, University Hospitals of Cleveland. I was promoted to a full Professor in July 2000.  Over the last 10 years, my laboratory has been devoted to the study of medically important fungi, focusing on pathogenesis, as well as development of in vitro and in vivo preclinical assays.

1. Fungal pathogenesis:

In this area, I have focused on the study of Candida albicans, a fungus that causes the important systemic infection, candidiasis.  I have studied this organism because it is the most important emerging clinical fungal pathogen, accounting for 8% of all hospital-acquired bloodstream infections, and the fourth most common cause of septicemia.  Even with ideal antifungal treatment therapy, mortality due to candidiasis still persists at an unacceptably high range between 38-50%.  New innovative treatment approaches, based on an understanding of the biology and virulence of this organism, are needed.  In this regard, our efforts focused mainly on the study of candidal virulence factors (e.g. adherence, phospholipases). More recently, I initiated research into the new area of fungal biofilms, because these complexes likely represent the predominant mode of fungal infection.  The following is a highlight of some of the work my group undertook in the last decade.

a) Candidal adherence studies:

            My interest in the study of C. albicans adherence to mammalian cells, believed to be the first step in host invasion, dates to my tenure at Kuwait University.  These studies established a correlation between proteinase production, adherence, and pathogenicity of various strains of C. albicans.  Additionally, we discovered that certain lipid classes (phospholipids and sterols) play an important role in candidal adherence to host cells.  Upon arriving in the United States, I resumed my work on candidal adherence, studying the anti-adherence activity of antifungals and natural proteins.  These experiments revealed that fluconazole and platelet microbicidal protein inhibit Candida adherence to platelets.  We also showed that, in addition to their direct effects on the growth of Candida, fluconazole and amphotericin B may decrease the ability of the fungus to disseminate hematogenously by inhibiting the organism’s capacity to adhere to and injure endothelial cells.  To understand candidal adherence at the molecular level, we cloned and characterized CAD1/AAF1, a gene from C. albicans capable of inducing adherence to endothelial cells after expression in normally non-adherent Saccharomyces cerevisiae.  This work culminated in the publication of a number of original research articles and a CRC book entitled “Candida Adherence to Epithelial Cells” (Boca Raton, Florida).

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            b) Contribution of extracellular phospholipases to candidal virulence: 

Since earlier work provided experimental evidence for the role of phospholipids in Candida adherence, and due to our extensive experience in fungal lipid biochemistry acquired while in Kuwait, experiments were initiated to study the role of secreted phospholipase in Candida virulence.  The “Molecular Koch’s Postulates” criteria were followed to prove that phospholipase B is a virulence factor for C. albicans.  Towards this end, the gene (caPLB1) encoding this enzyme was cloned, and an isogenic strain pair of C. albicans, differing only in their ability to secrete PLB1, was constructed.  The pathogenicity of the constructed strains was examined in two murine models of candidiasis (intravenous, and oral-intragastric).  Our data showed that, unlike the parental strain, the virulence of mutants deleted for caPLB1 were significantly attenuated.  Finally, using immuno-electron microscopy and western blot analyses, it was demonstrated that caPLB1 is expressed and the encoded protein is detectable in host tissues during the infectious process.  These findings provided unequivocal evidence that phospholipase B is a virulence determinant in C. albicans.  This work was funded by an RO1 from the National Institutes of Health (NIH), USA.

            Because we showed that phospholipase B is an important pathogenicity determinant in C. albicans, the NIH renewed my RO1 grant to study the mechanisms by which this enzyme augments candidal virulence.  We used both in vitro tissue culture systems and in vivo animal models to compare the ability of the isogenic strain pair in host cell penetration and transmigration across the gastrointestinal tract of mice.   Taken together, our findings provided evidence that the mechanism by which phospholipase B contributes to the virulence of C. albicans involves direct damage of host cell membranes.  As destruction of the submucosa progresses, the fungus transmigrates across the gastrointestinal tract, penetrates the vascular tissue, and disseminates hematogenously. The article published in Clinical Microbiology Reviews summarizes of our efforts to understand the role of extracellular phospholipase in fungal pathogenesis.

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c) Lipid and sterol biochemistry:

Fungal lipids and sterols received our attention due to their importance in the understanding of C. albicans pathogenicity, and as antifungal targets. Specific areas investigated included the role of lipids in candidal morphology, adherence and biofilm formation. More recently, we investigated the effect of voriconazole (a third generation azole approved by the FDA in 2002) on sterol synthesis of resistant and susceptible Candida species.  I have published extensively in this area, including a CRC book entitled “Lipids of Pathogenic Fungi” (Boca Raton, Florida, USA).

d) Biology and drug resistance of Candida Biofilms:

            Over the last three years, my group started work on the new area of biofilms.  Biofilms can serve as a nidus for disease and are often associated with high-level antimicrobial resistance.  My interest in the study of Candida biofilm stems from the fact that candidiasis associated with intravenous lines and indwelling medical devices is especially problematic, associated with mortality rates that approach 40%, and, because of our expertise in fungal pathogenesis, biochemistry, and antifungal susceptibility, we are ideally situated to study multiple aspects of fungal biology and drug resistance.  Importantly, in contrast to the extensive literature describing bacterial biofilms, little attention has been paid to medically relevant fungal biofilms.

            To investigate fungal biofilms, we developed two biofilm models representing invasive and superficial forms of candidiasis.  In the invasive model, we used silicone elastomer (utilized extensively in indwelling medical devices such as catheters) as substrate, while in the superficial model, we employed polymethylmethacrylate, which is used in construction of dentures.   Next, we used these models to study the development, architecture, and drug resistance of C. albicans biofilms.  Our data show that candidal biofilm formation proceeds through three distinct developmental phases, resulting in a highly heterogeneous architecture of well-defined cellular communities encased in a polysaccharide matrix.  Comparison of biofilms formed in our in vitro model and those found on an infected central venous catheter from a patient revealed that the biofilm formed on the infected catheter was similar in structure to biofilms grown using our silicone elastomer system.  This data indicated that our in vitro model is analogous to in vivo events and may be clinically relevant.  These studies formed the basis for investigations into the molecular mechanisms of Candida biofilm biology and antifungal resistance.

            Next, we examined the antifungal susceptibilities of C. albicans and C. parapsilosis (the latter species is associated with central lines and parenteral nutrition, particularly in critically ill neonates) biofilms to conventional and new antifungals.  These studies took advantage of my expertise in developing methods to measure the susceptibility of fungi to antifungals.  Susceptibility testing of fluconazole, nystatin, chlorhexidine, terbinafine, amphotericin B, and triazoles (voriconazole and ravuconazole), revealed resistance by all Candida isolates examined, when grown as biofilms, compared to planktonic forms.  In contrast, lipid formulations of amphotericin B and echinocandins showed activity against Candida biofilms.  If these results can be extended to animal models, our discovery could represent a breakthrough in the treatment of invasive Candida infections.  We are currently developing a guinea pig model of catheter-associated Candida biofilm to determine whether our in vitro findings could be reproduced in vivo.   

            After showing that Candida growing in biofilms is highly resistant to most antifungals, we initiated studies examining the mechanisms underlying this resistance at the genetic and biochemical levels, exploiting our expertise in Candida molecular biology and sterol biochemistry.  Our data demonstrated that antifungal resistance in biofilms is a multifactorial phenomenon, with one mechanism operational in the early phase [drug efflux pumps such as CDR (a member of the ABC super-family) and MDR (a member of the major facilitator super-family)], while others (alterations in membrane sterol composition) are functional in mature biofilms.  I recently received an RO1 grant from the National Institutes of Health to continue our studies into the biology and drug resistance of Candida biofilms, using state-of-the-art genomic and proteomic approaches.  These studies will allow us to identify genes and proteins involved in the formation of biofilms, as well as development of drug resistance.  Based on our expertise and publications, the American Society for Microbiology Press has commissioned Dr. O’Toole (Dartmouth College) and me to edit a book on microbial biofilms.  This book should be completed by September 2003. 

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2. Development of in vitro and in vivo assays:

            Since beginning my academic career, I have had an interest in developing: a) in vitro antifungal susceptibility testing that has utility in drug discovery and patient management, and b) in vivo animal models for evaluating virulence factors. To this end, over the last 9 years, and as a member of the National Committee for Clinical Laboratory Standards, Subcommittee on Antifungal Susceptibility Testing (NCCLS), we developed an in vitro test system for determining the antifungal susceptibility of C. neoformans (a fungus that causes meningitis in AIDS patients) that is specific, standardized, and reproducible.  Importantly, because MIC data generated using this method can predict treatment success for patients with acute AIDS-associated cryptococcal meningitis, this method has clinical utility.  Based on these findings, the NCCLS has adopted this technique as a reference method for determining the antifungal susceptibility of C. neoformans.  More recently, we developed a new method for determining the antifungal susceptibility of dermatophytes.  This method is currently being tested in other laboratories nationally as a prelude to adopting it as a reference method.

For the past decade, through collaborative efforts with academic laboratories and the pharmaceutical industry, my group has developed and used a variety of experimental animal models including 1) murine, guinea pig, and rabbit, 2) normal and immunocompromised animals, 3) localized and disseminated infection, and 4) treatment vs. prophylaxis models. We used these models to evaluate the antifungal activity of a broad range of chemical entities, including: microbicidal proteins derived from host neutrophils and platelets (defensins and platelet microbicidal proteins), azoles (fluconazole, voriconazole and ravuconazole), allylamines (terbinafine), polyenes and their liposomal antifungal preparations (AmBisome, ABELCET and Nyotran), immunomodulator (interferon-g), antineoplastic agents, and various types of other synthetic molecules.  The Center for Medical Mycology, which I direct, has in place a facility directed at pre-clinical evaluation of candidate antifungal agents against different fungal infections, including those caused by Candida, Cryptococcus, Fusarium, and Aspergillus.  Based on our expertise, we are among few laboratories nationally, and internationally, that are contacted by the pharmaceutical and biotechnology industry to evaluate potential antifungal agents.  The candidiasis experimental animal models have been employed to determine the role of specific genes in fungal virulence by assessing the pathogenicity of genetically altered mutants.

In conclusion, over the last decade, I have established a medical mycology center of excellence, funded by industry and government agencies, where multidisciplinary investigations into the bases of fungal diseases and translational research are undertaken.  The center attracts scientists/physicians aiming to consolidate their expertise in medically important fungi.  My scientific contribution has been acknowledged by my peers at national and international levels, as illustrated by invitations to serve as a member of: the National committee for Clinical Laboratory Standards, editorial boards of highly prestigious scientific microbiology journals, organizing committees for scientific conferences, as well as grant review committees for the VA, NCCLS, and NIH.

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