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Microbiology, virusology, immunology department
Medical Mycology
as. O.B. Kuchmak
Medical Mycology
TheseLecture
Students
The Sea of Knowledge
Medical Mycology Outline
1. Introduction, Actinomycetes2. Yeasts, Dermatophytes3. Filamentous Fungi, Dimorphic Fungi4. Dimorphic Fungi5. Opportunistic Fungi
OBJECTIVES
–To impart sufficient basic science of the medically important fungi to assist you in diagnosing mycotic diseases.
–To impart sufficient clinical knowledge
to raise your index of suspicion for mycotic diseases.
What is a Fungus ?
• Eukaryotic – a true nucleus
• Do not contain chlorophyll
• Have cell walls
• Produce filamentous structures
• Produce spores
KINGDOM CHARACTERISTIC EXAMPLE
Monera Prokaryocyte BacteriaActinomyces
Protista Eukaryocyte Protozoa
Fungi Eukaryocyte * Fungi
Plants Eukaryocyte PlantsMoss
Animals Eukaryocyte * ArthropodsMammals
Man
KINGDOM CHARACTERISTIC EXAMPLE
Monera Prokaryocyte BacteriaActinomyces
Protista Eukaryocyte Protozoa
Fungi Eukaryocyte * Fungi
Plants Eukaryocyte PlantsMoss
Animals Eukaryocyte * ArthropodsMammals
Man
ALL EUKARYOTIC CELLS CONTAIN STEROLS
•Mammalian cells – cholesterol
•Fungal cells - ergosterol
Cocci 0.8 u
Bacilli 4-6 u
Spirochetes 8 - 10 u
Viruses 0.08 u
Protozoa 15 u
Nematodes 10 mm
Fungi 10 – 15 u
SIZE COMPARISON OF PATHOGENS
Species of Fungi
• 100,000 – 200,000 species
• About 300 pathogenic for man
The taxonomy of the Kingdom Fungi is evolving and is controversial. Formerly based on gross and light microscopic morphology, studies of ultra structure, biochemistry and molecular biology provide new evidence on which to base taxonomic positions.
Medically important fungi are in four phyla:
1.Ascomycota - Sexual reproduction in a sack called an ascus with the production of ascopspores
2.Basidiomycota -Sexual reproduction in a sack called a basidium with the production of basidiospores
3.Zygomycota - sexual reproduction by gametes and asexual reproduction with the formation of zygospores
4.Mitosporic Fungi (Fungi Imperfecti) - no recognizable form of sexual reproduction. Includes most pathogenic fungi.
Classification
◘ ASCOMYCOTA Trichophyton Microsporum Blastomyces◘ BASIDIOMYCOTA Cryptococcus neoformans◘ DEUTEROMYCOTA Epidermophyton Sporothrix Candida species ◘ ZYGOMYCOTA Rhizopus nigrans
Actinomyces(True Bacteria)
• Tradition• Clinical infection resembles mycoses• Actinomyces grow on mycotic media• Actinomyces grow slowly (24-48 h)• Gross colonies resemble fungi
– (rough,heaped, short aerial filaments)
• Resemble mycelia microscopically, with branched mycelia in tissue and smears.
What is Medical Mycology ?
PATHOGENIC FUNGI
• NORMAL HOST• Systemic pathogens - 25 species• Cutaneous pathogens - 33 species• Subcutaneous pathogens - 10 species
• IMMUNOCOMPROMISED HOSTOpportunistic fungi - 300 species
MYCOTIC DISEASES(Four Types)
1. Hypersensitivity– Allergy
2. Mycotoxicosis– Production of toxin
3. Mycetismus (mushroom poisoning)– Pre-formed toxin
4. Infection
Hypersensitivity
Farmer’s lung Moldy hay
Malt worker’s disease Moldy barley
Cheese washer’s lung Moldy cheese
Wood trimmer’s disease Moldy wood
PARASITIC STATE
1. Increased metabolic state
2. Modified metabolic pathways
3. Modified cell wall structure– Carbohydrate content– Lipid structure– RNA aggregates
PATHOGENICITY OF FUNGI
1. Thermotolerance
2. Ability to survive in tissue environment
3. Ability to withstand host defenses
REVIVED INTEREST IN MYCOLOGY
• Increased frequency of mycotic diseases• Increased awareness by physicians• Better trained laboratory personnel• More invasive procedures used on patients• Increased use of immunosuppressive drugs• Increase in immunosuppressive disease
7. Better laboratory diagnostic tools
EYE
SKIN
UROGENITAL TRACT
ANUS
MOUTHRESPIRATORY
TRACT
PORTAL OF ENTRY
•SKIN
•HAIR
•NAILS
•RESPIRATORY TRACT
•GASTROINTES-TINAL TRACT
•URINARY TRACT
EYE
SKIN
UROGENITAL TRACT
ANUS
MOUTHRESPIRATORY
TRACT
COLONIZATION
Multiplication of an organism at a given site without harm to the host
EYE
SKIN
UROGENITAL TRACT
ANUS
MOUTHRESPIRATORY
TRACT
INFECTION
Invasion and multiplication of organisms in body tissue resulting in local cellular injury..
GEOGRAPHIC DISTRIBUTION
The present ease and frequency of world-wide travel make it more likely that physicians in the United States will be confronted with a variety of unfamiliar mycoses acquired in distant parts of the country or of the world.
MORPHOLOGY
Fungi exist in three morphologic form:
1. Yeasts 2. Molds. 3. Dimorphs.
• Pathogenic fungi can exist as yeasts or as hyphae (figure 4). A mass of hyphae is called mycelia. Yeasts are unicellular organisms and mycelia are multicellular filamentous structures, constituted by tubular cells with cell walls. The yeasts reproduce by budding. The mycelial forms branch and the pattern of branching is an aid to the morphological identification. If the mycelia do not have SEPTA, they are called coenocytic (nonseptate). The terms "hypha" and "mycelium" are frequently used interchangeably. Some fungi occur in both the yeast and mycelial forms. These are called dimorphic fungi.
Clinical Classification of Mycoses
CutaneousSubcutaneousSystemicOpportunistic
Cutaneous Mycoses
Skin, hair and nails
Rarely invade deeper tissue
Dermatophytes
Subcutaneous Mycoses
• Confined to subcutaneous tissue and rarely spread systemically.
• The causative agents are soil organisms introduced into the extremities by trauma
Systemic Mycoses
• Involve skin and deep viscera
• May become widely disseminated
• Predilection for specific organs
OPPORTUNISTIC FUNGI
Ubiquitous saprophytes and occasional pathogens that invade the tissues of those patients who have:
•Predisposing diseases: Diabetes, cancer, leukemia, etc.
•Predisposing conditions: Agammaglobulinemia, steroid or
antibiotic therapy.
MOST MYCOTIC AGENTS
ARE SOIL SAPRPHYTES
EPIDEMIOLOGY
Mycotic Diseases Are NOT
Contagious
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
DIAGNOSIS
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
DIRECT MICROSCOPIC OBSERVATION
• 10 % KOH
• Gentle Heat
KOH Wet Mount
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
SKIN TESTING(DERMAL HYPERSENSTIVITY)
Use is limited to :
– Determine cellular defense mechanisms– Epidemiologic studies
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
FUNGI ARE POOR ANTIGENS
FUNGAL SEROLOGYANTIBODIES
• Latex Agglutination IgM
• Immunodiffusion IgG
• Complement Fixation IgG
Most serological tests for fungi measure antibody. Newer tests to measure antigen are now being developed
ANTIGEN DETECTION PRESENTLY AVAILABLE
Cryptococcosis Aspergillosis Histoplasmosis
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
DIRECT FLUORESCENT ANTIBODY
CAN BE APPLIED TO
1. HISTOLOGIC SECTIONS
2. CULTURE
• Viable organisms• Non-viable organisms
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probe
INFLAMMATORY REACTION
• Normal host–Pyogenic
–Granulomatous
• Immunodeficient host–Necrosis
Polymorphic Nuclear Leukocytes
GMS
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
ISOLATION MEDIA
SABOURAUD DEXTROSE AGAR
(pH ~ 5.6)
•Plain
•With antibiotics
•With cycloheximide
INCUBATION TEMPERATURE
• 37 C - Body temperature
• 25 C - Room temperature
Diagnosis
1. Wet Mount2. Skin test3. Serology4. Fluorescent antibody5. Biopsy and histopathology6. Culture7. DNA probes
DNA Probes
• Rapid (1 Hour)
• Species specific
• Expensive
TREATMENT
THERAPY
Because they are eukaryotic, fungi are biochemically similar to the human host. Therefore it is difficult to develop chemotherapeutic agents that will destroy the invading fungus without harming the patient.
A BASIC TENET OF PATHOLGY IS:
A CAUSE OF IRREVERSIBLE CELL INJURY IS CELL MEMBRANE DAMAGE.
IN FUNGAL THERAPY
We attempt to induce cell injury by causing the cell membrane of the fungus to become permeable.
PROBLEM
Finding an agent that will selectively injure fungal cell walls without damaging the host cell.
Cryptococcosis
A sub-acute or chronic infection which may affect the lungs or skin but most commonly manifests as a meningitis
Cryptococcus neoformans
• Spherical
• 5-10 microns
• Narrow- base
• Surrounded by polysaccharide capsule
Cryptococcus neoformans
Virulence factors
1. Anti-phagocytic polysaccharide capsule
2. Antioxidant melanin
3. Ability to grow at 37 C
Ecological Niche
Cryptococcus neoformans
• Pigeon droppings
• Chicken droppings
Cryptococcusportal of entry
• Inhalation
• inoculation
Typical clinical course• Inhalation
• Mild pneumonia
• Vision, lethargy, headache
• Delirium, nuchal rigidity, coma
• Over a period of months
• Death if not treated
Clinical Specimens
Cryptococcosis
•CSF•Sputum•Pus•Biopsy material•urine
Specimen of choice
Cerebrospinal fluid
Serological Tests
Cryptococcosis
• Indirect fluorescent antibody• Tube Agglutination• Latex Agglutination (measures antigen)
Drugs of Choice
Cryptococcosis
•Amphotericin B plus 5-Flucytosine
OR
•Amphotericin B plus Fluconazole
Yeasts
Unicellular forms that are
spherical or ovoid in shape.
Round to oval, single cells
which reproduce by budding
8-12 MICRONS
Pseudomycelium
Clinical SpecimensCandida
• Sputum
• Scrapings from lesions
• Blood smears
• Vaginal discharge
• Urine
• Feces
• Nail clippings
SEROLOGY• There have been several antibody
detection test, but none are significant.
• Beta-Glucan measurement is now showing promise
Therapy
• Nystatin– Vaginitis– Cutaneous
• Fluconazole *
• Itraconazole
Cryptococcosis
A sub-acute or chronic infection which may affect the lungs or skin but most commonly manifests as a meningitis
Yeasts
Unicellular forms that are spherical or ovoid in shape
• Hyphae (filamentous fungi, mycelium)– Septate– Coenocytic (non-septate)
• Dimorphic– Yeast– Mycelium
Yeasts
Unicellular forms that are spherical or ovoid in shape
• Hyphae (filamentous fungi, mycelium)– Septate– Coenocytic (non-septate)
• Dimorphic– Yeast– Mycelium
Dimorphic Fungi
• Yeast Form• Parasitic form• Tissue form• Cultured at 37 C
• Mycelial Form• Saprophytic form• Cultured at 25 C
SPORES
• SEXUAL
• ASEXUAL– Arthrospore– Blastospore– Chamydospore– Conidia
• Microconidia• Macroconidia