Transcript Mycology – Introduction Student Lab Division of Medical Technology

Mycology –
Introduction
Student Lab
Division of Medical Technology
Carol Larson MSEd, MT(ASCP)
Mycoses
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Superficial
Subcutaneous
Systemic
Opportunistic
Characteristics of fungi
• Eukaryotic
• Growth requirements
• Forms
– Mold
– Yeast
Hyphae
• Septate
• Aseptate
Hyphae
• Hyaline
• Dematiaceous
Mycelium
• Mass of branching intertwined hyphae
– Vegetative
– Aerial
– Fertile
Vegetative types
• Favic chandeliers
• Nodular organs
• Racquet hyphae
• Spiral hyphae
Reproduction
• Identify fungi by:
– Morphology of reproductive structures
– Spores from vegetative mycelium or
aerial fruiting bodies
Asexual Reproduction
• Conidia
• Conidiophore
• Arthroconidia
Asexual Reproduction
• Blastoconidia
• Pseudohyphae
• Chlamydoconidia
• Chlamydospores
Asexual Reproduction
• Macroconidia
• Microconidia
• Phialoconidia
• Phialide
Asexual Reproduction
• Annelloconidia
• Annellide
• Sporangiospores
• Sporangium
• Sporangiophore
Sexual Reproduction
• Perfect Fungi – has a sexual stage
• Fungi Imperfecti – no know sexual
stage
• Spores
Sexual Reproduction
• Ascospores
– Ascus
– Ascocarp
• Basidiospores
• Zygospores
In review …
• Mycoses – fungal diseases
• Characteristics of fungi
– Growth requirements
– Forms (mold, yeast)
– Structures
– Reproduction
• Asexual
• Sexual
Fungal Culture Process
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Specimen collection and transportation
Direct examination of specimen
Selection and inoculation of media
Evaluation of fungal growth
Serological testing
Antifungal susceptibility testing
Specimen Collection
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Specimen types
Collect from area most likely infected
Use sterile technique
Keep specimen moist
Label container properly
Transport right away
Process right away
Direct Examination
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Provides preliminary report
Guides MD in treatment of patient
Observe yeast phase of dimorphic
Gives clues to id causative agent
Inoculate special media
May require more than one direct
examination method
Direct Examination
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Saline wet mount
Lactophenol cotton blue wet mount
10% KOH preparation
Gram stain
Acid fast stain
India ink stain
Direct Examination
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Calcofluor white stain
Wright’s stain
Gomori Methenamine Silver stain
Periodic Acid Schiff stain
Specimen Processing
• Safety
– Tube media preferred over plate media
– Work in safety hood
– Wear gloves and lab coat
– Autoclave specimens and media
– Disinfect work area daily
Specimen Processing
• Primary isolation media
– Goal: isolate potential pathogens
– Use non-selective and selective media
– Proper ingredients
– Incubation temperature
– Incubation time
– Incubation atmosphere
Non-selective Media
• Sabouraud dextrose agar
• Brain heart infusion (BHI) with/without
5% blood and 1% glucose
Selective Media
• Mycosel agar
• Inhibitory mold agar
• Dermatophyte test medium
Subculture / Identification Media
• Neutral Sabouraud dextrose agar
(Emmon’s)
• Cornmeal-Tween 80 agar
• Niger seed agar (Birdseed agar)
• Tween 80 / Oxgall / caffeic acid agar
• Potato dextrose agar
Examination of Culture Growth
• Potential pathogens
– Slow growers
– Growth on Mycosel
– Color: dull buff, brown, mousy gray
– Dimorphic
Examination of Culture Growth
• Growth rate
– Rapid growers: 1-5 days
– Intermediate growers: 6-10 days
– Slow growers: >10 days
Colony Morphology –
Appearance
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Rugose
Umbonate
Verrucose
Flat
Colony Morphology –
Texture
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Cottony
Glabrous
Granular
Velvety
Colony Morphology –
Pigmentation
• Surface
• Reverse
Microscopic Morphology
• Definitive means of identification
• Evaluate:
– Shape
– Method of production
– Arrangement of conidia/spores
– Size and color of hyphae
Microscopic Techniques
• Tease mount
• Scotch tape preparation
• Slide culture
Serological Diagnosis
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Immunodiffusion
Complement fixation
EIA
Latex agglutination
Antifungal Susceptibility
• Determine appropriateness
– Standardization of testing
– Methods
– Predictability in vivo
• Antifungal agents
In Summary …
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Specimen collection and transport
Specimen processing and culture
Direct examination of specimen
Examination of culture
Serological testing
Antifungal susceptibility