Pathogenic Fungi: Structural Biology and Taxonomy
Chapter Abstracts

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Chapter 1. The Structure and Composition of the Fungal Cell Wall
Rafael Sentandreu, M. Victoria Elorza, Eulogio Valentín, and José Ruiz-Herrera

Abstract
The cell wall is probably one of the most characteristic structures of fungi and can be sued to define this group of organisms. The cell wall has many functions including the protection of the fungal cell from the harsh physical, chemical and biological conditions. It is also responsible for the shape of the organism. But, the cell wall also is the structure responsible for all of the interactions of fungi with their environment. Cell surface biomolecules are responsible for attachment of fungi to several types of surfaces, including host cells in the case of pathogenic fungi. The cell wall also acts as an immunogen, cause immunosuppression, or act as a virulence factor. Enzymes present in the cell wall degrade large molecules, such as proteins, polysaccharides, and lipids, to provide nutrients for cell growth. Chemically the fungal cell wall is mostly composed of polysaccharides. Proteins are less abundant, followed by lipids, and other minor components such as pigments and inorganic salts. These components are held together as a coherent structure by non-covalent bonds, mainly hydrophobic, hydrogen bonds, and by covalent bonds. Thus, the wall is a three-dimensional structure that serves its multiple functions. Structurally, the fungal cell wall appears as a biphasic system consisting of two kinds of components, structural polysaccharides, such as chitin and glucans, that are organized as microfibril bundles that provide rigidity for the cell, and an amorphous component, primarily glycoproteins and other polysaccharides that cover the inner, microfibrillar layer. The amorphous layer contains the adherence glycoproteins and other functional biomolecules associated with protection against the environment.


Chapter 2: Biosynthesis of the Fungal Cell Wall
José Ruiz-Herrera, M. Victoria Elorza, Peggy E. Alvarez, and Rafael Sentandreu

Abstract
The biosynthesis of the fungal cell wall is an extremely complex phenomenon. The wall is made of different components, some of which are synthesized intracellularly, while others are polymerized outside the permeability barrier, but all of them must come together at a precise time and place, and in the precise amounts, to react and be organized into a coherent structure. Accordingly, there must exist precise mechanisms that regulate these processes. We know that synthesis of the cell wall is a polarized process that depends on the transport of components and biosynthetic enzymes to the growing sites. This is a function that depends on the cytoskeleton that carries all these components inside secretory vesicles and microvesicles to sites where wall expansion takes place. This process is tightly associated with the functioning of the secretory pathway. During its operation, proteins are synthesized, and placed into the lumen of the endomembrane system, where they become glycosylated and associated in part with some other polysaccharides. This pathway is also operative for the enzymes responsible for the biosynthesis of structural polysaccharides. By means of specific vesicles, the chitosomes, the enzymes responsible for chitin synthesis are carried to the cell surface where synthesis of the polysaccharide takes place. Similar mechanisms must be operative for glucan synthesis. Once at the cell surface, association among the different components takes place to produce a three-dimensional structure maintained and organized by non-covalent and covalent bonds that join them together. These processes are themselves subjected to regulatory mechanisms that permit changes in the wall, both qualitative and quantitative, in response to internal or external effectors. These changes permit the fungus to alter its morphology during its cell and life cycles, when interacting with other organisms, to better exploit the environment, compensate reduction in the synthesis of wall components, or adapt to different types of stress.


Chapter 3: Cell Cycle of Fungal Pathogens
Judith Berman and Neil A.R. Gow

Abstract
The cell cycle coordinates morphogenesis so that mitosis and growth are coupled to the doubling of all cellular components. Recent work with certain fungal pathogens has revealed modifications of the basic cell cycle pattern as exhibited in model yeast species that tailors the needs of the pathogen to its growth in the host environment. Many fungal pathogens such as Candida albicans are dimorphic or pleomorphic, and so the cell cycle must be regulated to enable cell shape to be modulated while the nuclear cycle is maintained. Filamentous pathogens such as Aspergillus fumigatus maintain a nuclear cycle but uncouple cell division from the growth cycle. Therefore fungal pathogens provide interesting examples of how cell cycle events are both conserved and modified to facilitate replication in the context of invasion of human tissues.


Chapter 4: Morphogenesis in Candida albicans
Tamaki Cho

Abstract
The human pathogen Candida albicans displays polymorphic growth. That is, it grows as unicellular yeast but also converts reversibly to filamentous growth (hyphae) by germination of yeast cells, a process that is referred to as morphogenesis and can form an intermediate type of growth or pseudohyphae. This polymorphic growth is regulated by a number of environmental conditions and is considered to be one of virulence factors of the organism. A number of groups have studied the factors that influence morphogenesis in this organism. More recent approaches that emphasize molecular genetics have rapidly appeared in the literature. Functional analysis of genes associated with morphogenesis in C. albicans has included the identification of regulatory systems with the objective of constructing protein networks that regulate growth patterns. In this chapter, the literature will be summarized on several aspects of morphogenesis, including, 1) the identification of growth form-specific proteins and morphogenesis signaling pathways, 2) recent studies that examine transcriptional profiling of morphogenesis through microarray approaches, and 3) gene analysis of the early stages of morphological conversion of yeast to hyphae.


Chapter 5: Morphogenesis in Other Agents of Systemic Mycoses
Gioconda San-Blas and Gustavo Niño-Vega

Abstract
Important dimorphic pathogenic fungi like Histoplasma capsulatum, Blastomyces dermatitidis, Sporothrix schenkii, Paracoccidioides brasiliensis, Penicillium marneffei, Coccidioides immitis, and Wangiella dermatitidis have not been studied as systematically as Candida albicans or Aspergillus species from the point of view of molecular mechanisms controlling the dimorphic process. There is only fragmentary information that is available on assorted mechanisms probably involved in the dimorphic transition. The recent focus of study on dimorphism has been of genes associated with cell wall and membrane lipid synthesis, structure, cytoskeleton, and metabolic pathways. Research is needed in both signal transduction pathways and on the molecular level of the genome and the supramolecular level of the cell in order to really understand the morphogenetic processes carried out during the life cycles of many of these fungi.


Chapter 6: Regulation of Morphogenesis by Conserved Developmental Pathways in Pathogenic Fungi
Idit Hazan and Haoping Liu

Abstract
The highly conserved pheromone response MAP kinase and nutrient-sensing cAMP/PKA signal pathways are critical for filamentation, mating and virulence in many pathogenic fungi. A comparison of their functions in two human pathogens Candida albicans and Cryptococcus neoformans, and in two plant pathogens, Magnaporthe grisea and Ustilago maydis, shows that virulence is tightly associated with filamentation and mating in these fungi, suggesting an evolutionary link between pathogenesis and cellular development.


Chapter 7: Beyond Molecular Biology: Fungal Morphology as a Mathematical, Biophysical and Computational Subject
Gioconda San-Blas and Juan Murgich

Abstract
Morphogenesis at the level of the cell requires details of molecular biology, but that is not sufficient to determine spatial patterns that exist at a scale that is larger than three to five orders of magnitude. Thus, the molecular paradigm, while not incorrect, is incomplete. Therefore, it follows that principles of spatial organization must exist at the level of the cell that influence the morphogenetic process. Physical forces and fields, applied to biological systems, must be at work to explain morphogenesis, once the molecules have been synthesized. Biophysical, mathematical, and computational approaches have been developed in order to explain morphogenetic events, in an attempt to raise the molecular data to a higher level of understanding. In this chapter we summarize a mathematical morphological index, the steady-state, biomechanical and vesicle-supply centre models, and molecular modelling of cell wall glucans.


Chapter 8: The Use of Phylogenetic Analysis to Investigate Uncultivated Microbes in Medical Mycology
Leonel Mendoza and Victor Silva

Abstract
The classical way to investigate the relationship between organisms includes, but is not limited to, taxonomic macro- and microscopic morphological changes within individuals of the same group. One drawback to this approach has been that the uncultured microbes are difficult to study in fine detail in the absence of cultures. With the advent of phylogenetic analysis, however, several taxonomic mysteries, including the phylogenetic connections of uncultivated microbes, have been revisited. In medical mycology in particular, two enigmatic microbes that resisted cultivation were investigated: Lacazia loboi and Rhinosporidium seeberi. Despite ultra-structural, serological and chemical studies, the relationship of these two pathogens remained a mystery for more that 100 years. Using 18S SSU rDNA, ITS sequences and chitin synthase genes, however, we found that L. loboi formed a sister clade to Paracoccidioides brasiliensis in the dimorphic Onygenales, and that R. seeberi was a protistal Mesomycetozoea located at the point where the fungi first diverged from animals about 1.6 billion year ago. This article will discuss in detail some of the aspects that made these studies possible and future directions to investigate other traits in these unusual microbes.


Chapter 9: Gene Genealogical Analyses of Human Fungal Pathogens
Jianping Xu

Abstract
The last several years have seen increasing use of DNA sequence data in population genetic analyses. In this chapter, I summarized the principles of gene genealogical analyses and how such analyses have been used to address evolutionary and population genetic issues of three human fungal pathogens Cryptococcus neoformans, Histoplasma capsulatum and Coccidioides immitis. Within each of the species, gene genealogical analyses provided unambiguous evidence for significant divergence, recent dispersion, clonality and recombination. In addition, hybridization was detected in both C. neoformans and H. capsulatum. While gene genealogical analyses so far have been focused on haploid fungal species, the approach described here should be applicable to diploid organisms such as C. albicans.


Chapter 10: Polysaccharides F1SS: Taxonomic and Evolutionary Characters for Ascomycetes
A. Prieto, O. Ahrazem, M. Bernabé, and J.A. Leal


Abstract

The alkali-extractable and water-soluble F1SS polysaccharides from the fungal cell wall seem to be a reliable taxonomic and evolutionary character. The number of polysaccharidic structures that may be formed by a combination of different sugars and linkages can be used for the delimitation of genera, grouping of taxa in higher ranks, establishment of connections among meiotic and strictly mitotic genera, and the elaboration of fungal evolutionary schemes. In addition, the immunogenic potential of fungal cell wall and cell surface polysaccharides may result in the establishment of sensitive immunological techniques for the rapid identification of pathogenic fungi.

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