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Mycology

Dental / Optometry Fundamentals II Stephen A. Moser, Ph.D.

10/26/2011

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Epidemiology

Geography Endemic mycoses Worldwide mycoses

Transmission of infection Respiratory inhalation (systemic mycoses) Cutaneous inoculation (sporotrichosis) Systemic invasion by opportunistic normal flora

(candidiasis) Contact with infected hosts (dermatophytoses)

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Epidemiology (Cont.) Risk factors and manifestations of disease

True pathogens versus opportunists Environmental risk factors for systemic fungal

disease• Location and travel• Occupation

Host defenses and susceptibility to systemic fungal disease (CMI most important)• Congenital and acquired T cell deficiencies (including

AIDS)• Immunosuppression (transplants and malignancies)• Diabetes mellitus

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Endemic Distribution for Blastomycosis

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General Characteristics

Aerobic - obligate or facultative Eukaryotic: membrane bound nucleus

and cytoplasmic organelles (may be multinucleate)

Achlorophyllous Morphology (unicellular or multicellular) Saprophytic (heterotrophic)

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Characteristics of Fungal Cells

Cell wall: multilayered polysaccharide Cellulose, glucans, mannans, chitin, polypeptides Absence of teichoic acids, peptidoglycan, LPS

Cell membrane Phospholipid bilayer Ergosterol (relate to chemotherapy)

Cytoplasm - typical eukaryotic organellesNucleus - either uninucleate or multinucleate

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Characteristics of Fungal Cells

Capsule Present in some species (e.G. Cryptococcus

neoformans) Amorphous polysaccharide coating Functions and activities

AntiphagocyticAntigenic

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Growth forms Yeast - unicellular fungi which reproduce by

budding (Cryptococcus) Mold - hyphae (mycelium)

Septate hyphae (Aspergillus)Non-septate, coenocytic hyphae (Mucor)

Pseudohyphae (Candida albicans) Thermal dimorphism

Characteristics of Fungal Cells

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PROPERTY FUNGI BACTERIA

Cell diameter 5-50 microns 1-5 microns

Nucleus Eukaryotic prokaryotic

Cytoplasmic organelles Present absent

Cell membrane sterols present (ergosterol) absent (exceptMycoplasma)

Cell wall chitin, glucans,mannans, peptides

teichoic acids,peptidoglycan, LPS

Metabolism Mainly aerobes, facultativeanaerobes

obligate and facultativeaerobes and anaerobes

Thermal dimorphism Common in many pathogenicspecies

absent

Differences Between Bacteria and Fungi

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Examples of Yeast & Pseudohyphae

Pseudohypha

Blastoconidia

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Blastomyces dermatitidis Thermal Dimorphism

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Example of True Septate Hyphae

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Non-septate Hyphae

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Asexual Reproduction

Conidia (spores) – asexual structures Blastospores – formed by budding yeasts

(Blastomyces) Chlamydospores – terminal or intercalary

cells with thick walls (Candida albicans) Arthrospores – formed by fragmentation of

hyphae (Coccidioides immitis) Sproangiospores – formed in sporangia by

cleavage (Rhizopus)

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Classification Based onSexual Phase

Ascomycetes: Aspergillus, Histoplasma, Blastomyces, Dermatophytes

Basidiomycetes: Cryptococcus, Mushrooms

Zygomycetes: Order Mucorales - Mucor, Rhizopus

Deuteromycetes (Fungi Imperfecti): Sporothrix, Coccidioides, Candida

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Clinical Types of Fungal Infections

TYPE DISEASE ORGANISM

1. Superficial Pityriasis versicolor Malassezia furfur

2. Cutaneous Ringworm (Tinea)Candidiasis

Trichophyton speciesCandida albicans and others

3. Subcutaneous Sporotrichosis Sporothrix schenckii

4. Systemic Pathogenic Fungi Histoplasmosis Histoplasma capsulatumBlastomycosis Blastomyces dermatitidisCoccidioidomycosis Coccidioides immitisParacoccidioidomycosis

Opportunistic Fungi

Paracoccidioides brasiliensis

Aspergillosis Aspergillus fumigatus andothers

Cryptococcosis Cryptococcus neoformansCandidiasis Candida albicans and othersZygomycosis Mucor and Rhizopus species

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Routes of Infection

Inhalation of spores – major factor Inoculation of spores into skin Disease by normal flora in compromised

host (Candida) Hypersensitivity Contact with infected host

(Dermatophytes) Mycotoxins

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Laboratory Diagnosis of Fungal Infections

Microscopic Examination of tissues and body fluids

Gram stain Giemsa India Ink Potassium hydroxide (KOH) wet prep Hematoxylin and Eosin stain Periodic-Acid Schiff stain (PAS) Gomori-Methenamine Silver stain (GMS) Mucicarmine or Alcian Blue stain

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Budding Yeast - Gram Stain

Staphylococcus

Candida

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Encapsulated Yeast - India Ink

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KOH Prep - Broad-base Budding Yeast

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H&E Stain - Budding Yeasts

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GMS Stain - Septate Hyphae

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Mucicarmine Stain - C. neoformans

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Histopathological Response to Fungal Infection

Acute pyogenic abscess (Candida) Chronic granuloma formation

(Histoplasma) Chronic, localized dermal inflammation

(Dermatophytes) Mixed pyogenic and granulomatous

inflammation (Blastomyces) Blood vessel invasion with thrombosis and

infarction (Mucor, Aspergillus) Hypersensitivity without tissue reaction

(allergic bronchopulmonary aspergillosis)

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Fungal Cultures

Utilize Sabouraud agar with antibiotics Identification criteria

Temperature of growth Rate of growth Colonial and microscopic morphology Sporulation pattern Biochemical reactions (yeast)

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Fungal Serology

Generally poor and not as useful as in other pathogens such as viruses and bacteria, with some exceptions.

Cryptococcal antigen by latex agglutination: serum and CSF.

Coccidioides - early IgM response is useful for identification of acute primary disease - CSF IgG prognostic value.

Skin tests for DTH - problems: Cross-reactivity. High positive rate in endemic areas.

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Candidiasis Clinical manifestations

Mucosal• Vaginitis• Esophagitis• Oral thrush

Cutaneous Chronic mucocutaneous Systemic

• Fungemia• Hepato-spleenic• Endophthalmitis• Renal

Urinary tract

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Oral Candidiasis

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Wet Mount - Candidiasis

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Mucocutaneous Candidiasis

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Candida sp. Tissue GMS Stain

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Aspergillosis Clinical manifestations

Pneumonia Aspergilloma Allergic bronchopulmonary Disseminated multiorgan involvement

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Aspergilloma

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Allergic Bronchopulmonary Aspergillosis

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CNS Aspergillosis

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Aspergillus sp – GMS Stain

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Zygomycosis

Clinical manifestations Sinusitis Rhinocerebral Pulmonary Renal

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Rhinocerebral Mucormycosis

in Diabetic Ketoacidosis

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Postmortem – Rhinocerebral Mucormycosis

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Non-septate Branching Hyphae (PAS)

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Histoplasmosis

Clinical manifestations Most cases mild or sub-clinical pulmonary

disease• Dissemination appears to be common

Pneumonia Chronic progressive pulmonary (cavitary) Histoplasmoma Disseminated

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Histoplasmosis – Calcified

Lesions

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Histoplasmosis- GMS

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Histoplasmosis – Bone Marrow

Histiocyte

H. capsulatum

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Histoplasma capsulatumIn vitro In vivo

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Presumed Ocular Histoplasmosis

Thought to be a late stage of primary histoplasmosis.

Causes abnormal blood vessels – scar tissue.

Organism has not been found in eye.

Treated with laser surgery.

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Risk Factors for Endogenous Endophthalmitis

Candidia species Central venous lines, neutropenia, abdominal surgery, intravenous drug abuse, broad-spectrum antibiotics

Aspergillus species Neutropenia, endocarditis, intravenous drug abuse, pulmonary disease being treated with high dose steroids, organ and stem cell transplant.

H. capsulatumC. immitisB. dermatitidisC. neoformans

May accompany disseminated disease

Fusarium species Neutopenia, intravenous drug abuse

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Fungal Keratitis

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Chemotherapy FDA approved

Polyenes (Amphotericin B, lipid encapsulated forms) Azoles (fluconazole, itraconazole, ketoconazole,

voriconazole) Echinocandin (Caspofungin, Micafungin, Anidulafungin) Nucleoside derivatives (5-flurocytosine) Allyamines (Terbinafine) Microtubule disruption (Griseofulvin)

Investigational Nikkomycins (chitin synthase inhibitors) Echinocandin/pnemocandin/lipopeptide class (inhibit glycan

synthesis)

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Antifungal Drugs for Systemic Mycoses - Amphotericin B

Mode of Action Binds to ergosterol, increases membrane

permeability resulting in leakage of cytoplasmic components and cell death – Fungicidal

Spectrum of Activity Candida, Crypto, Aspergillus, Histo, Blasto, Cocci,

etc Limitations

Nephrotoxicity

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Antifungal Drugs for Systemic Mycoses - Fluconazole

Mode of Action Prevents ergosterol synthesis by inhibiting the C-14

demethylation step (cytochrome P-450 rx) Fungistatic

Spectrum of Activity Candida, Crypto, Trichsporonosis, dermatophytes

Limitations Resistance in some Candida sp – krusei and glabrata Not effective for non-dermatophyte moulds.

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Antifungal Drugs for Systemic Mycoses -Echinocandins

Mode of Action Prevents synthesis of beta 1,3-glucan

required for cell wall. Fungistatic

Spectrum of activity Aspergillus, Candida NOT effective against Cryptococcus,

zygomycetes.

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Early Diagnosis of Invasive Fungal Infections

Obstacles• Because of Immunosuppression typical signs

and symptoms of infection are frequently absent• Few clinical features are uniquely specific for

systemic fungal infection• Sputum and blood cultures are frequently

negative• Invasive procedures

– May be necessary for definitive diagnosis– Are often complicated in severely

immunocompromised patient

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Early Diagnosis of Invasive Fungal Infections (Continued)

Benefits Early diagnosis permits selection of a

therapy of maximal effectiveness Early intervention with antifungal

therapy may help decrease the high mortality rate associated with serious systemic mycoses

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Major Areas Covered

How fungi differ from bacteria The major fungal infections The epidemiology of fungal infections Pathology of fungal infections Mechanism of action of antifungal agents