Mycology
General Characteristics
Eukaryotic
Sterols
Chemoheterotrophs
Aerobes
Neutral pH
Saprophytic
Living on dead/decayed organic matter
Acquire food by absorption
Produce sexual or asexual spores (conidia)
Candida (yeast) Cell Wall
Contains:
Mannoproteins (protein): cell stability
Chitin (polysaccharide): structural integrity
Glucans (polysaccharide): structural integrity
Cryptococcus Cell Wall
Capsule (polysaccharide): virulence factor, protects against and interferes with host immune cells
Chitin: structural integrity
Glucans: Structural integrity
Melanin (phenolic compound): virulence factor, protects against and interferes with host immune cells
Aspergillus Cell Wall
Glactomannan (polysaccharide): released during tissue invasion
Glucans and Chitin: Structural integrity
Molds:
Multicellular
Filamentous form of fungi consisting of thread-like filaments
For fuzzy colonies
Yeast:
Unicellular
Produce circular. Restricted, pasty mucoid colonies
Dimorphic:
Exhibit either yeast or mold phase
Filamentous Fungi
Composed of hyphae: microscopic filaments
Forms mycelium: network of hyphae
Aerial: hold reproductive structures
Vegetative: absorb nutrients
Hyphae
Shape: tubular
Septate- cell wall breaks
Frequent perpendicular cross-walls = septate
Infrequent = sparely septate
Pigmentation
Hyaline (nonpigmented)
Dematiaceous (dark, pigmented hyphae, presence of melanin)
Unicellular Yeast
Capable of reproducing asexually/sexually
Asexual reproduction:
Budding
Binary fission
Fungal Spores (Conidia)
Role:
Dispersal
Survival (low metabolic state)
Dormant state:
Germinate if conditions are favorable
uni/multicellular
Varies in size, shape, color
Spectrum of Disease
Superficial
Cutaneous
Mucosal
Systemic
Fungal Populations in Humans
Oral Activity
Candida
Aspergillus
Malassezia
Cladosporium
Saccharomyces
Fusarium
Penicillium
Pichia
Rhodotorula
Lungs
Aspergillus
Cladosporium
Penicillium
Candida
Urinary Tract
Candida
Saccharomyces
Malassezia
Vagina
Candida
Saccharomyces
Pichia
Cladosporium
Phodotorula
Breast Milk
Malassezia
Davidiella
Sistotrema
Penicillium
Skin
Malassezia
Aspergillus
Candida
Intestine
Candida
Saccharomyces
Aspergillus
Cyberlindnera
Fusarium
Malassezia
Rhodotorula
Wallemia
Cladosporium
Penicilium
Classification
Clinical Microbiology based on mucoses (fungal diseases)
Superficial
Confined to outermost “dead” layer of skin/hair
Cutaneous
Affects keratinized layer of skin, hair, nails
Subcutaneous
Deeper skin layers
Systemic
Affects internal organs/deep tissues of body
Opportunistic
Found in immunocompromised people
Innate Resistance
Body temp = too high for fungi to adapt/mutate for
Exposed to a huge amount of spores
Mode of Transmission
superficial/cutaneous
Person to person
Animal too human
Subcutaneous
Through the skin (via injury)
Deep mycoses
Opportunistic growth in immunocompromised
Inhalation of spores/conidia
Specimen Collection/Transport
Selection based on
Information obtained from clinical examination/radiographic studies
Consideration of most likely fungal pathogens
Collect aseptically
Collect adequate amount of material
Submit sterile leak-proof container
Store in 4C
Specimen Handling
Process under biological safety cabinet
ID methods:
Direct microscopy
Biochemical tests
Susceptibility testing
Automated identification systems
Histopathological methods
Immunologic methods
Molecular methods
Culture Time/Conditions
Time
Yeast: Growth detected 48-72 hours
Mold: 4-6 weeks
Temp
Incubation for fungals = 25-30C
Dimorphic fungi
Mold temp (30C)
Yeast phase (37C for 7-14 days)
Fungal Culture Media
Battery of media should use:
Enriched media
Ensure growth of fastidious thermally dimorphic fungi
General purpose media
Ensure growth of variety of molds and yeast
No antimicrobial agents
Antibacterial
Chloramphenicol
Gentamicin
antibacterial/antifungal
Cycloheximide
May inhibit Cryptococcus neoformans, Mucorales, Candida sp. Aspergillus sp, and Histoplasma
Colonial Morphology
Texture: height of aerial hyphae
woolly/cottony (High/dense)
Velvety (low)
granular/powdery (flat/dense)
Color: Surface and reverse
Topography: designs of kills/valleys
Microscopy
Stains/reagents
Lactophenol cotton blue
Kills any organisms
Lactic acid acts as clearing agent
Preserves fungal structures
Stains chitin in fungal cell walls
Potassium hydroxide
10 to 20% solution dissolves keratin
Calcofluor white
Fluorescent stain
Binds to cellulose and chitin in fungal walls
Gram stain
Stain gram positive
ID
Observe asexual reproductive structures (sporangiospores/conidia)
Antifungal Agent
Fungistatic (slows down growth) or Fungicidal (kills fungus)
Systemic active agents
Polyenes
Weakens cell membrane
Kills cell due to loss of cell integrity
Bindings to ergosterol produce little holes in the membrane
Problem: binds to cholesterol
Can kill human cells
Broad spectrum of activity
Fungicidal
Main adverse effect: nephrotoxicity (kidney damage)
Azoles
Attacks cell membrane
Prevents ergosterol to be produced
Weakens cell membrane
Fungistatic to yeast
Fungicidal to mold
Echinocandins
Attacks cell wall
Inhibits glucans, affecting the cell’s integrity
Low toxicity
Fungicidal (Candid) and fungistatic (Aspergillus)
Flucytosine
Prevents DNA, RNA, protein synthesis
Prevents fungal cell wall from forming
Fungistatic
Limited spectrum
Candida, Cryptococcus select dematiaceous molds
Used in combination with another antifungal (due to resistance)
Allylamines
Inhibit squalene
Decreases ergosterol and increases squalene in cell (toxic to cell)
Broad spectrum
Fungicidal
Topical Active Agent
Creams
Lotions
Ointments
Powders
Sprays
Treats superficial, cutaneous, mucosal infections
Topical vs. Systemic depends on:
Status of host
type/extent of infection
Antifungal Resistance
Resistance develops slowly
Involves alteration of cellular structures/functions
Mechs:
Biofilm formation (reduces penetration of antifungal)
Efflux pumps (reduces accumulation of medicine)
Target alterations/mutations
Overexpression of target (overproduction of enzyme)