Mycology – Yeast Student Lab Division of Medical Technology Carol Larson MSEd, MT(ASCP)

Basic Characteristics Unicellular Generally normal flora

Basic Characteristics Asexual reproduction –Blastoconidia –Pseudohyphae –Arthroconidia

Direct Examination Observe for reproductive structures Gram stain –Gram positive India ink stain –Capsules

Growth Requirements Growth in 2-3 days Temperature –25-37ºC –Best at 30ºC

Colony Morphology Appearance –Similar to bacteria colony Texture –Glabrous –May be filamentous Pigmentation Confuse with Staphylococcus

Yeast ID Methods Germ Tube Production –First test performed when yeast –Procedure –Interpretation

Yeast ID Methods Cornmeal-Tween 80 agar –Procedure –Observe microscopically for Chlamydospores Blastoconidia Pseudohyphae Arthrospores

Yeast ID Methods Niger Seed Agar (Birdseed agar) –Procedure –Observe for brown pigment caused by phenol oxidase activity –Cryptococcus neoformans

Yeast ID Methods Urease –Procedure –Positive: Cryptococcus sp. (rapid) Trichosporon sp. Rhodotorula sp. –Negative: Candida sp. Geotrichum sp.

Yeast ID Methods Carbohydrate assimilation –Utilize carbohydrate as sole source of carbon in the presence of oxygen –Positive = growth Carbohydrate fermentation –Utilize a carbohydrate anaerobically –Positive = gas

Yeast ID Methods Commercial identification kits –Manual methods –Automated methods

Candida species Normal flora Leading opportunistic fungal infection in man Candidiasis –Mucocutaneous –Cutaneous –Systemic

Factors predisposing to infection Immunocompromised Malignancy Prolonged antibiotic therapy Lupus, Diabetes Pregnancy and use of birth control pills Age Damaged skin barrier

Candida albicans #1 fungal isolate in laboratory Germ tube: positive CMT: clustered blastoconidia at septa of pseudohyphae, terminal chlamydospores CHO: sucrose positive

Candida albicans

Candida stellatoidea Germ tube: positive CMT: clustered blastoconidia at septa of pseudohyphae, terminal chlamydospores CHO: sucrose negative

Candida tropicalis Germ tube: negative CMT: sparse single or short-chained blastoconidia anywhere along pseudohyphae, rare chlamydospores CHO: sucrose positive

Candida glabrata Causes UTI Germ tube: negative CMT: only blastoconidia, no pseudohyphae CHO: only glucose and trehalose positive

Candida krusei Germ tube: negative CMT: tree-like branching of abundant blastoconidia from the septa of elongated pseudohyphae (“cross- matchsticks” appearance)

Candida pseudotropicalis Germ tube: negative CMT: branched pseudohyphae with chains of elongated blastoconidia – logs in a stream arrangement of broken up blastoconidia positioned parallel to each other

Candida parapsilosis Germ tube: negative CMT: few single or small clustered blastoconidia at or between septa of thin curved pseudohyphae. Sometimes giant pseudohyphae may be observed

Cryptococcus neoformans Cryptococcosis –Pulmonary –Meningitis Pigeon droppings

Cryptococcus neoformans Gram stain India ink stain

Cryptococcus neoformans Germ tube: negative CMT: large, round blastoconidia, no pseudohyphae Urease: positive (within 3 hours)

Cryptococcus neoformans Niger seed agar: brown pigment

Cryptococcus neoformans Cryptococcal antigen test

Geotrichum candidum Geotrichosis –Mimics Candida infections –Infection in debilitated people Considered a mold Germ tube: negative CMT: arthroconidia and true hyphae Urease: negative

Trichosporon beigelii Causes White Piedra Germ tube: negative CMT: arthroconidia, true hyphae, rare blastoconidia Urease: positive

Rhodotorula species Saprophytic yeast found in soil Rarely a pathogen Often encapsulated Urease: positive Colony pigmentation: reddish-orange

Saccharomyces species Rarely a pathogen Ascospores (acid fast positive)

Malassezia furfur Causes Tinea Versicolor Septicemia in patients receiving intravenous lipid therapy KOH: clusters of thick-walled, round budding yeast and short, straight or angular mycelial fragments (spaghetti and meatball appearance) Lipophilic

In Summary … Key characteristics of yeast Identification methods –Germ tube –CMT –Biochemical tests Clinically significant yeast –Candida albicans –Cryptococcus neoformans