MYCO VIRO - Laboratory Diagnosis of Fungi

Wood's Lamp at 365nm

Hair should be examine with this apparatus at ____ nm UV light to check for Microsporum audouinii infections.

Outer edge

Skin should be scraped at what region of a lesion?

70% Isopropyl Alcohol

Skin should be clean with this chemical before sampling

Blood and Bone Marrow

These sample is known to harbor pathogenic and opportunistic fungi.

Wampole, Cranbury, NJ Isolator Tube

Most sensitive method. Uses lysis centrifugation to release microorganisms from WBCs and RBCs.

Becton-Dickinson Diagnostic Systems Septi-Check

Biphasic system. Heparinized bone marrow samples should be plated directly onto media at bedside.

Blood Culture Bottles

This is not recommended for Blood and Bone marrow handling

Cerebrospinal Fluid

Media with antimicrobials is NOT necessary for this sample

Centrifugation

Cerebrospinal Fluid is concentrated by using this method before inoculation.

Latex Agglutination

Use one drop of this in CSF sample for Cryptococcus infections

Grind / Ground

Tissue Samples should be _____ which is necessary for KOH prep.

Purulent and Discolored

Tissue sections with this characteristics should be selected

Sputum
Tracheal aspirate
Pleural Lavage

Commonly Respiratory Specimens

Dacron swab

When dealing with respiratory specimen that is a viscous materials use this type of swab

Sterile pipette

Excessively viscous materials should be inoculated onto media using this equipment

N-acetyl-L-cysteine

Respiratory specimen can be digested with this chemical

Oropharyngeal Candidiasis

This should be direclty smear and culture on a CHROMagar Candida

KOH Preparation

The purpose of this is to detect fungal elements in skin, hair, nails, and tissue.

10% - 20%

Working solution of KOH

Potassium Hydroxide (KOH)

Breaks down keratin and skin layer

Dimethyl Sulfoxide

Chemical that rapidly breakdown of cellular debris; no heating required.

Calcofluor White

Fluorescent dye; fungal elements appear apple-green or blue-white.

Phenol - Kills
Lactic Acid - Preserves
Cotton Blue - Stain

Lactophenol Cotton Blue’s composition and their function

India Ink

Its purpose is to negative stain for Cryptococcus neoformans in CSF.

Periodic Acid-Schiff

Tissue stains that stains polysaccharides

Fontana-Masson

Tissue stains that detects melanin

Gomori Methenamine-Silver Nitrate

Tissue stains that stains fungal elements

Purplish-red

Color of PAS

Dark brown or Black

Color of Fontana-Masson

Black

Color of GMS

SDA at 37*C

Yeast form of dimorphic fungi and other organisms grow; dermatophytes grow poorly

BHI with blood with 37*C

Yeasts such as Cryptococcus grow well; Histoplasma capsulatum takes up heme pigment, becoming light tan with a grainy, wrinkled texture.

SDA at 22*C

Initial isolation of pathogens and saprobes; dimorphic fungi may show mycelial phase.

SDA with antibiotics at 22*C

Saprobes inhibited; dermatophytes and primary pathogens grow.

BHI at 22*C

Initial isolation of pathogens and saprobes

BHI with antibiotics at 22*C

Recovery of pathogenic fungi; dermatophytes not usually recovered.

Inhibitory Mold Agar at 22*C

Initial isolation of pathogens except dermatophytes

Cycloheximide at 22*C

Primary recovery of dermatophytes.

Trichophyton mentagrophytes

Positive Organism in Hair Perforation Test which shows perpendicular penetration pegs within hair shafts

5-Day Urease Test

Differentiate Trichophyton mentagrophytes from T. rubrum

Christensen Agar

Culture medium of 5-Day Urease Test

5 days at RT

Incubation of sample in 5-Day urease test

Positive - T. mentagrophytes
Negative - T. Rubrum

Result of 5-day Urease Test

Growth on Rice Grains

Its purpose is to differentiate Microsporum canis from M. audouinii

10 days at RT

Incubation of sample in Growth on Rice Grains

Microsporum Canis

Growth on Rice Grains result = Abundant growth + conidia formation.

M. audouinii

Growth on Rice Grains result = No growth; turns rice brown

Germ Tube Production

Most important test, easiest test for yeast identification

Carbohydrate Assimilation

Identifies which carbohydrates a yeast can use aerobicall

CHROMagar Candida

Presumptively identifies C. albicans, C. tropicalis, C. krusei

Cornmeal Agar

Recognizes yeast morphologies: blastoconidia, chlamydoconidia, pseudohyphae, arthroconidia.

C. albicans
C. Tropicalis
C. Krusei

CHROMagar candida presumptiviley identifies these species

Candida tropicalis

Color blue colony in CHROMagar Candida

Candida albicans

Color green colony in CHROMagar Candida

Candida krusei

Color violet/pink colony in CHROMagar Candida

Slide culture (Riddell technique)

This method’s purpose is to aid in the identification of molds by microscopic appearance

Potato dextrose agar

Medium used in Slide culture

C. albicans

This fungus is used as positive control in germ tube

Cryptococcus spp

This fungus is used as negative control in germ tube