Basic Mycology


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Chapter 47 Basic Mycology Medical Microbiology Mycology

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Medical Microbiology Bacteriology Virology Mycology Protozoalogy Helminthology Parasitology Immunology Medical Microbiology The study of fungi causing diseases in humans

What is Mycology?:

What is Mycology? Mycology is the study of fungi Fungi play a major role in the environment, industry and our life: Bread Cheese Alcoholic beverages (ethanol) Mushrooms Environmental toxins Biodegradation Disease

Definition of some Terms:

Definition of some Terms Yeasts - are unicellular fungi which reproduce asexually by blastoconidia formation (budding) or fission Dimorphism - is the condition where by a fungus can exhibit either the yeast form or the hyphal form, depending on growth conditions Hypha, Hyphae - are multi-cellular fungi which reproduce asexually and/or sexually Pseudohyphae - elongation of buds forming an appearance of hyphae It can be hard to distinguish hyphae from pseudohyphae. Pseudohyphae form indentiations at each new juncture of a new cell. True hyphae do not generally form indentations at septations.

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Arthroconidia - barrel shaped cells formed from hyphae Chlamydoconidia - round thick walled cells formed from hyphae or pseudohyphae A mass of hyphal elements is termed the mycelium (synonymous with mold ). Aerial hyphae often produce asexual reproduction propagules termed conidia (synonymous with spores ).


Con … Relatively large and complex conidia are termed macroconidia while the smaller and more simple conidia are termed microconidia . When the conidia are enclosed in a sac (the sporangium), they are called endospores . The presence/absence of conidia and their size, shape and location are major features used in the laboratory to identify the species of fungus in clinical specimens.

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A: Blastoconidia and pseudohyphae ( Candida ) B: Chlamydospores ( Candida ). C: Arthrospores ( Coccidioides ). D: Sporangia and sporangiospores ( Mucor ) E: Microconidia ( Aspergillus ). F: Microconidia and macroconidia ( Microsporum ).




Conidia usually free of any surrounding membranes that hold them in one loctions

Sporangiospores :

Sporangiospores usually contained in a sac


STRUCTURE & GROWTH Two fungal cell structures are important medically: 1- The fungal cell wall consists primarily of chitin thus, fungi are insensitive to antibiotics, such as penicillin, that inhibit peptidoglycan synthesis. - Chitin is a polysaccharide composed of long chains of N -acetylglucosamine . The fungal cell wall contains other polysaccharides as well, the most important of which is Beta -glucan , “a long polymer of D-glucose”. The medical importance of Beta -glucan is that it is the site of action of the antifungal drug caspofungin .

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2- The fungal cell membrane contains ergosterol , in contrast to the human cell membrane, which contains cholesterol. The selective action of amphotericin B and azole drugs , such as fluconazole and ketoconazole , on fungi is based on this difference in membrane sterols Con…

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Table 47 – 1. Comparison of Fungi and Bacteria. Feature Fungi Bacteria Diameter Approximately 4 µ ( Candida ) Approximately 1 µ ( Staphylococcus ) Nucleus Eukaryotic Prokaryotic Cytoplasm Mitochondria and endoplasmic reticulum present Mitochondria and endoplasmic reticulum absent Cell membrane Sterols present Sterols absent (except Mycoplasma ) Cell wall content Chitin Peptidoglycan Spores Sexual and asexual spores for reproduction Endospores for survival, not for reproduction Thermal dimorphism Yes (some) No Metabolism Require organic carbon; no obligate anaerobes Many do not require organic carbon; many obligate anaerobes

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Yeasts grow as single cells that reproduce by asexual budding. Yeasts and Molds

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Molds grow as long filaments (hyphae) and form a mat ( mycelium ). Some hyphae form transverse walls ( septate hyphae ), whereas others do not ( nonseptate hyphae ). Nonseptate hyphae are multinucleated (coencytic). Several medically important fungi are thermally dimorphic; i.e., they form different structures at different temperatures. They exist as molds in the environment at ambient temperature and as yeasts (or other structures) in human tissues at body temperature.

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Most fungi are obligate aerobes; some are facultative anaerobes ; but none are obligate anaerobes. All fungi require a preformed organic source of carbon—hence their frequent association with decaying matter. The natural habitat of most fungi is, therefore, the environment. An important exception is Candida albicans , which is part of the normal human flora. Con…

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Some fungi reproduce sexually by mating and forming sexual spores, e.g., zygospores, are single large spores with thick walls ascospores, are formed in a sac called ascus basidiospores are formed externally on the tip of a pedestal called a basidium. The classification of these fungi is based on their sexual spores. Fungi that do not form sexual spores are termed "imperfect" and are classified as fungi imperfecti. Con…

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Most fungi of medical interest propagate asexually by forming conidia (asexual spores) from the sides or ends of specialized structures. The shape, color, and arrangement of conidia aid in the identification of fungi. Some important conidia are (1) arthrospores, 1 which arise by fragmentation of the ends of hyphae and are the mode of transmission of Coccidioides immitis; (2) chlamydospores, which are rounded, thick-walled, and quite resistant (the terminal chlamydospores of C. albicans aid in its identification); (3) blastospores, which are formed by the budding process by which yeasts reproduce asexually (some yeasts, e.g., C. albicans, can form multiple buds that do not detach, thus producing sausagelike chains called pseudohyphae, which can be used for identification); and (4) sporangiospores, which are formed within a sac (sporangium) on a stalk by molds such as Rhizopus and Mucor. Con…

Arthrospores :


Chlamydospores :


Blastoconidia and pseudohyphae :

Blastoconidia and pseudohyphae

Sporangia and sporangiospores :

Sporangia and sporangiospores

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Although this lecture focuses on the fungi that are human pathogens, it should be remembered that fungi are used in the production of important foods , e.g., bread, cheese, wine, and beer. Fungi are also responsible for the spoilage of certain foods. Because molds can grow in a drier, more acidic, and higher-osmotic-pressure environment than bacteria, they tend to be involved in the spoilage of fruits, grains, and vegetables.


PATHOGENESIS The response to infection with many fungi is the formation of granulomas. Granulomas are produced in the major systemic fungal diseases, e.g., coccidioidomycosis, histoplasmosis, and blastomycosis, as well as several others. The cell-mediated immune response is involved in granuloma formation. Acute suppuration( pyogenic response ), characterized by the presence of neutrophils in the exudate, also occurs in certain fungal diseases such as aspergillosis and sporotrichosis. Fungi do not have endotoxin in their cell walls and do not produce bacterial- type exotoxins .

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Activation of the cell-mediated immune system results in a delayed hypersensitivity skin test response to certain fungal antigens injected intradermally. A positive skin test indicates exposure to the fungal antigen. It does not imply current infection, because the exposure may have occurred in the past. A negative skin test makes the diagnosis unlikely unless the patient is immunocompromised. Because most people carry Candida as part of the normal flora, skin testing with Candida antigens can be used to determine whether cell-mediated immunity is normal.

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Intact skin is an effective host defense against certain fungi (e.g., Candida, dermatophytes), but if the skin is damaged, organisms can become established. Fatty acids in the skin inhibit dermatophyte growth, and hormone-associated skin changes at puberty limit ringworm of the scalp caused by Trichophyton. The normal flora of the skin and mucous membranes suppress fungi. When the normal flora is inhibited, e.g., by antibiotics, overgrowth of fungi such as C. albicans can occur.

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In the respiratory tract, the important host defenses are the mucous membranes of the nasopharynx, which trap inhaled fungal spores, and alveolar macrophages. Circulating IgG and IgM are produced in response to fungal infection, but their role in protection from disease is uncertain. The cell-mediated immune response is protective; its suppression can lead to reactivation and dissemination of asymptomatic fungal infections and to disease caused by opportunistic fungi.


FUNGAL TOXINS & ALLERGIES Gwo other kinds of fungal disease: (1) Mycotoxicoses, caused by ingested toxins (2) allergies to fungal spores. The best-known mycotoxicosis occurs after eating Amanita mushrooms. These fungi produce five toxins, two of which—amanitin and phalloidin—are among the most potent hepatotoxins. The toxicity of amanitin is based on its ability to inhibit cellular RNA polymerase , which prevents mRNA synthesis.

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Another mycotoxicosis, ergotism , is caused by the mold Claviceps purpura , which infects grains and produces alkaloids (e.g., ergotamine and lysergic acid diethylamide [LSD]) that cause pronounced vascular and neurologic effects.

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Other ingested toxins, aflatoxins, are coumarin derivatives produced by Aspergillus flavus that cause liver damage and tumors in animals and are suspected of causing hepatic carcinoma in humans. Aflatoxins are ingested with spoiled grains and peanuts and are metabolized by the liver to the epoxide , a potent carcinogen. Aflatoxin B1 induces a mutation in the p53 tumor suppressor gene, leading to a loss of p53 protein and a consequent loss of growth control in the hepatocyte. aflatoxins

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Allergies to fungal spores, particularly those of Aspergillus, are manifested primarily by an asthmatic reaction ( rapid bronchoconstriction mediated by IgE ), eosinophilia, and a "wheal and flare" skin test reaction. T hese clinical findings are caused by an immediate hypersensitivity response to the fungal spores. Allergies


LABORATORY DIAGNOSIS There are four approaches to the laboratory diagnosis of fungal diseases: (1) direct microscopic examination, (2) culture of the organism, (3) DNA probe tests, and (4) serologic tests.

KOH and other stains:

KOH and other stains Direct microscopic examination of clinical specimens such as sputum, lung biopsy material, and skin scrapings depends on finding characteristic asexual spores, hyphae, or yeasts in the light microscope. The specimen is either treated with 10% KOH to dissolve tissue material , leaving the alkali-resistant fungi intact, or stained with special fungal stains.

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Some examples of diagnostically important findings made by direct examination are the spherules of C. immitis and (2) the wide capsule of Cryptococcus neoformans seen in India ink preparations of spinal fluid. Calcofluor white is a fluorescent dye that binds to fungal cell walls and is useful in the identification of fungi in tissue specimens. Methenamine-silver stain is also useful in the microscopic diagnosis of fungi in tissue.

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Fungi are frequently cultured on Sabouraud's agar , which facilitates the appearance of the slow-growing fungi by inhibiting the growth of bacteria in the specimen. Inhibition of bacterial growth is due to : low pH of the medium chloramphenicol cycloheximide . The appearance of the mycelium and the nature of the asexual spores are frequently sufficient to identify the organism. Sabouraud's agar

DNA Techniques:

DNA Techniques Tests involving DNA probes can identify colonies growing in culture at an earlier stage of growth than can tests based on visual detection of the colonies. As a result, the diagnosis can be made more rapidly. At present, DNA probe tests are available for Coccidioides, Histoplasma, Blastomyces, Cryptococcus .

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A significant rise in the antibody titer must be observed to confirm a diagnosis. The complement fixation test is most frequently used in suspected cases of coccidioidomycosis, histoplasmosis, and blastomycosis. In cryptococcal meningitis, the presence of the polysaccharide capsular antigens of C. neoformans in the spinal fluid can be detected by the latex agglutination test . Serology


ANTIFUNGAL THERAPY The most effective antifungal drugs, amphotericin B and the various azoles , exploit the presence of ergosterol in fungal cell membranes that is not found in bacterial or human cell membranes. Amphotericin B disrupts fungal cell membranes at the site of ergosterol and azole drugs inhibit the synthesis of ergosterol , which is an essential component of fungal membranes. Another antifungal drug, caspofungin (Cancidas), inhibits the synthesis of -glucan , which is found in fungal cell walls but not in bacterial cell walls. Human cells do not have a cell wall.

Systemic use (intravenous, oral) :

Systemic use (intravenous, oral) Name of Drug Mechanism of Action Important Adverse Reactions Amphotericin B Binds to ergosterol and disrupts fungal cell membranes Renal toxicity, fever, and chills; monitor kidney function; use test dose; liposomal preparation reduces toxicity Azoles such as fluconazole , ketoconazole , itraconazole , voriconazole , posaconazole Inhibits ergosterol synthesis Ketoconazole inhibits human cytochrome P450; this decreases synthesis of gonadal steroids resulting in gynecomastia Echinocandins such as caspofungin, mycafungin Inhibits synthesis of D-glucan, a component of fungal cell wall Well tolerated Flucytosine (FC) Inhibits DNA synthesis; FC converted to fluorouracil, which inhibits thymidine synthetase Bone marrow toxicity Griseofulvin Disrupts mitotic spindle by binding to tubulin Liver toxicity

Topical use (skin only); too toxic for systemic use :

Topical use (skin only); too toxic for systemic use Name of Drug Mechanism of Action Important Adverse Reactions Azoles such as cotrimazole, miconazole Inhibits ergosterol synthesis Well tolerated on skin Terbinafine Inhibits ergosterol synthesis Well tolerated on skin Tolnoftate Inhibits ergosterol synthesis Well tolerated on skin Nystatin Binds to ergosterol and disrupts fungal cell membranes Well tolerated on skin


Summary Morphologically fungi are unicellular (yeasts) or multicellular (hyphae). Some fungi can alternate between the two forms (dimorphic fungi). Hyphae are branching, threadlike, tubular filaments that either lack cross walls (coenocytic) or have cross walls (septate). Hyphae reproduce asexually via the formation of spores termed microconidia or macroconidia. India ink may be used as a negative stain to emphasize the capsule of yeast. Fungi are most commonly cultured on Sabouraud's agar or Mycosel agar. Antifungal agents are classified according to their chemical structure as macrolides, azoles, allylamines, and pyrimidine analogs. The polyene antifungals are amphotericin B and nystatin which bind to ergosterol in the plasma membrane, thus disrupting it.

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