Mycology lab final exam

Types of tinea infections

Tinea barbae: beard

Tinea capitis: scalp

Tinea corporis: body

Tinea cruris: jock itch

Tinea pedis: foot

Tinea onychomycosis: nail

Tinea versicolor: skin, M. furfur

"Tinea" (worm) + location of infection

Dermatophyte morphology (most)

cottony (fluffy)

Granular, powdery

*white front, yellow reverse

Sab-Dextrose

non-selective fungal agar

Inhibitory mold agar (IMA)

chloramphenicol: inhibits many bacteria

Mycosel agar

Chloramphenicol and cycloheximide

Inhibits: zygomycetes, aspergillus, Cryptococcus, some Candida

Dermatophyte test medium (DTM)

chloramphenicol and cycloheximide

also a differential media

Dermatophyte test medium coloration

Phenol red: red color appears in presence of alkaline metabolites

--chloramphenicol and cycloheximide inhibit other organisms

Ectothrix

spores and hyphae form sheath around hair

Spp: Microsporum spp, Trichophyton mentagrphytes

Endotrhix

spores and hyphae inside hair shaft

Spp: Trichophyton tonsurans and Trichophyton violaceum

Wood's UV lamp

for detection of ectothrix hair infection

KOH preps of skin/nails

degrade keratinized tissue to free fungal elements

--may add calcofluor white or LPCB to help visualize

Conidia

asexual spore structures formed from septate hyphae

MACROconidia: hyphal elements convert to multi-cellular structure

aseptate hyphae

septate hyphae

spiral hyphae

nodular organ

favic chandeliers/antler hyphae

Malassezia furfur

tinea VERSICOLOR

--lipophilic: needs lipid overlay to grow

Conidia and hyphae: spaghetti and meatballs

--normal flora in 90% of adults

*yeast-like fungus

Exophiala dermatitidis

colony: mucoid changing to velvet

--dark brown-black

--reverse (same)

Young culture: yeast-like cells

Mature colonies: pointed annelides branch from hyphae

--conidia cluster at tipe (cylindrical-oval)

septate hyphae

Disease:

Cutaneous: phaehyphomycosis granular cysts (immunocompetent)

Systemic: affinity for CNS, eyes (immmunocompromised)

*yeast-like fungus

Epidermophyton floccosum

Colony morph:

--felty, cottony tufts in older colony, white-yellow-khaki

--Reverse: light to medium brown

Microconidia: NO

Macroconidia: 'beaver tail', one end broader, smooth cell wall (2-5 perpendicular division, clustered at tapered end)

septate hyphae

Conditions: Tinea corporis and cruris, onchomycosis, DOES NOT INFECT HAIR

Microsporum canis

Colony: downy, white-yellow

Reverse: orange-yellow

Microconidia: rare

Macroconidia: fusiform spindle, septate macroconidia, thick cell wall, echinulate, 3-15 cells

septate hyphae

Conditions: scalp and skin;

--dogs, cats, farm animals

Nannizzia gypsea (Microsporum gypseum)

Colony: powdery, granular, yellow-beige

Reverse: same or red-brown

Microconidia: claviform (club-shaped)

Macroconidia: fusiform spindle, septate, thin-moderate cell wall, < 6 cells in spindle, spindle ends rounded

septate hyphae

Conditions: scalp and skin

animal cases more common

Trichophyton mentagrophytes (interdigitale) complex

Colony: velvety-white cream

Reverse: yellow or red-brown

Microconidia: pyriform to round

Macroconidia: claviform (club-shaped), may be absent; septate

septate hyphae

Disease: skin, hair, nails all common

--zoophilic

Trichophyton rubrum

Colony: powdery-downy, granular, white-pink

Reverse: red or yellow-brown

Microconidia: claviform or pyriform "birds on a wire"

Macroconidia: cigar shaped, often absent; septate and may be at end of hyphae

septate hyphae

Conditions: skin and nails common

Rare: beard/hair/scalp

Trichophyton tonsurans

Colony: powdery-downy, white-yellow-beige

Reverse: brown or red-brown

Microconidia: teardrop, club, balloon shaped, numerous

Macroconidia: club-shaped/curved, may be absent; septate

septate hyphae

Disease: tinea capitis (#1 cause in US)

--Skin, nails also possible

--anthropophilic

Trichophyton violaceum

Colony: glabrous, red violet brown or cream

Reverse: same, diffuses

Microconidia: usually absent

Macroconidia: typically absent

Hyphae: septate, irregular width

--may form chlamydospores

Disease: tinea capitis (#1 in Africa, Asia)

--skin, nails

Actinomycetoma

caused by actinomycetes (fungus-like bacteria)

--slow-growing, will grow on fungal media

Eumycotic mycetoma

true fungal infection

What do eumycotic mycetoma and actinomycetoma cause?

tumerous masses in subq tissue

--painless

--weeping sinuses

--granular discharge

--feet most commonly infected

Nocardia spp characteristics

Branching, filamentous AEROBES, grow on many media types

--Beaded GPB

--partially acid fast (some mycolic acid in cell wall)

--colony and micro morpho can resemble fungus

--slower growing

Soil organisms:

--Immunocompro: at higher risk for pulmonary--> brain lesion

--immunocompet: cutaneous, feet

Nocardia spp gram stain

beaded GPB

Acid fast: partially

Streptomyces spp

branching, filamentous aerobes

--common cause of actinomycetomas

--rare systemic infections

--used for antimicrobials

Actinomyces spp

Branching, filamentous ANAEROBES (many NOT obligate), fastidious

Colonies may be pigmented

A. israelii: molar tooth colonies

Actinomyces israelii molar tooth colonies

Actinomyces spp disease locations

mouth, pharynx, GU tract, distal esophagus

Mycobacterium spp

Acid fast: large amounts of mycolic acid in cell wall; AFB +

Skin infections: M. marinum, M. ulcerans, M. leprae

Pulmonary infections: M. tuberculosis, M. avium-intracellulare complex "MAC"

Mycobacterium spp in the lab

Acid fast:

--Kinyoun and Ziehl-Neelson stain

OR

Auramine rhodamine stain (fluorescent)

Growth media: Middlebrook 7H10, 7H11

Lowenstein-Jensen

Urea test

Nocardia spp POSITIVE

pinK: Klebsiella

P: Proteus

U: Ureaplasma

N: Nocardia

C: Cryptococcus

H: Helicobacter

Temperature tolerance testing

some actinomycetes can tolerate high temp

-score growth as 0 to 4+

--incubate at various temps (ex: 25, 37, 42 C)

Dimorphic fungi

yeast phase: body temp

Mold phase: room temp

Risk factors for dimorphic fungal infection

immunocompromised and immunocompetent

--certain geographic locations

Prepping dimorphic mold for microscopy

BSL3

--Coccoides spp has the highest risk for aerosolization and lab-acquired infection

Blastomyces dermatitidis

Blastomycosis: all ages vulnerable, immunocompromised more severe infection

Infection: inhalation of conidia, expore to lake/stream soil or disturbed soil

Infection types: skin lesions or pneumonia

Blastomyces dermatitidis mold form

B. dermatitidis

Coccioides imnitis

Coccoidiodomycosis/ Valley fever

--hot and dry conditions increase incidence

--inhalation of conidia

--all at risk, esp elderly/immunocompromised

Histoplasma capsulatum

Histoplasmosis

--inhaled microconidia

Risk factors: disturbed soil, bird bat dropping exposure, immunocompromised

Target tissue: pulmonary

Paracoccidioides brasiliensis

Paracoccidioidomycosis

Infection: inhalation of conidia (disturbed soil)

--direct handling of plants

Sites of infection: pulmonary, lymphatics, mucous membranes

Estrogen protects against infection

"mariner's wheel"

Sporothrix schenckii

"rose gardener's disease"

--cutaneous lesions on people that work with plants

--pulmonary or disseminated disease (joints) in the immunocompromised

Opportunists

very common in the environment, but can cause infections in certain scenarios (ex: pt is immunocompromised or in sterile sites)

--common culture contaminants

Opportunistic mold characteristics

fast growers; 2-3 days to mature

Not a contaminant when: isolated repeatedly from same patient/different sites, predominant organism in culture or pt is immunocompromised

Aspergillus spp prevalence

(in order)

1. Aspergillus fumigatus

2. Aspergillus flavus

3. Aspergillus niger

4. Aspergillus terreus

Media to Grow and ID the Opportunists

SAB-DEX

IMA (+ chloramphenicol)

Mycosel (+ chloramphenicol and cycloheximide; MANY Aspergillus spp inhibited)

Mucormycosis

a serious, acute fungal infection of the nasal sinuses+ eyes (rhino-orbital); leads to meningitis and can be rapidly fatal

Associated with: lymphoma, diabetes, AIDS, post-transplant

Suspect if: wooly, gray-white rapid grower; "lid-lifter"

Most common cause: Rhizopus spp

Mucormycete hyphae

broad and ribbon-like; ASEPTATE (sparse septa possible, but rare, unlike SEPTATE)

Alternaria spp

2-3 days

Light gray, wooly

Reverse: black

Microscopic: dark hyphae, chained, club-shaped poroconidia with tapered ends

Conditions: keratomycosis, skin, osteomyelitis, pulmonary disease, nasal septum

Bipolaris spp

2-3 days

Velety, wooly, gary-brown

Reverse: black

Microscopic: septate, dark hyphae

--cylindrical-shaped poroconidia with 4-5 cells inside

Conditions: keratomycosis and fungal sinusitis

Cladophialophora/ Cladosporium spp

Moderately slow; 7 days to mature

Powdery, velvet, dark gray, green; heaped and folded

Reverse: black

Microscopic: septate hyphae, short chains of dark blastoconidia

Conditions: keratomycosis/allergies

Curvularia spp

Rapid; 2-3 days

Cottony, white, light pink, orange or green

Reverse: brown

Microscopic: septate hyphae with dark poroconidia with 4-5 cells inside; boomerang shape

Conditions: keratomycosis, mycetoma, endocarditis, pulmonary, allergies, nasal septum

Exserohilum spp

2-3 days; rapid

Light gray, wooly, becomes gray-black

Reverse: black

Microscopic: septate hyphae with long cylindrical poroconidia with 6-14 cells. has a scar on the conidium

Conditions: Fungal sinusitis

Epicoccum spp

2-3 days; rapid

Rings of yellow, orange, brown pigment that diffuses into agar

Reverse: black

Microscopic: thick clusters of poroconidia

Conditions: allergies

Nigrospora spp

rapid; 2-3 days

White, wooly

Reverse: black

Microscopic: short, fat conidiophores; black conidia at tips

Conditions: keratomycosis

Poroconidium

a conidium developed through a channel or pore in hyphae

Acremonium spp

rapid; 2-3 days

Wrinkled, white, gray or rose

Reverse: colorless, pale yellow, pink

Microscopic: septate hyphae; tapering conidiophores that support balls of of conidia; can be confused with Sporothrix

Conditions: keratomycosis

Aspergillus spp

Rapid; 2-3 days

Rugose, velvety, blue, green, yellow, black or white

Reverse: colorless

Microscopic: septate, unbranched conidiophores with a foot cell and large vesicle at tip. phialides will form a single or double row. around vesicle

A. fumigatus will grow at 45 C

Conditions: disseminated aspergillosis, pulmonary, allergies, keratomycosis, otomycosis, nasal sinusitis

**most common opportunist pathogen

Chrysosporium spp

rapid; 2-3 days

Heaped, velvety, buff color

Reverse: white, yellow or brown

Microscopic: single, clubbed conidia: may be confused with blasto or histo

Conditions: n/a usually a contaminant

Fusarium spp

Rapid

White, wooly, turns lavender with age

Reverse: colorless

Microscopic: macrophialoconidia banana shaped

Conditions: keratomycosis (most common cause)

Gliocladium spp

Rapid

White, fluffy, turns green

Reverse: white

Microscopic: phialoconidia held in a ball

Conditions: contaminant; n/a

Paecilomyces spp

rapid

Powdery, velvety-white, becomes olive tan or pink

Reverse: colorless

Microscopic: elongated phialides have chains of oval conidia

Conditions: skin lesions and endocarditis

Penicillium spp

Rapid; 2-3 days

Velvety-white, becomes powdery, blue green

Reverse: colorless

Microscopic: chains of phialoconidia--paint brush

Conditions: contaminant; also causes keratomycosis, penicillinosis, otomycosis, onychomycosis

Scopulariopsis spp

Rapid; 2-3 days

Velvety, rugose, white then tan-brown

Reverse: brown

Microscopic: penicillium-like anelides support large, lemon-shaped anelloconidia that have spines (with age)

Conditions: keratomycosis

Sepedomium spp

Rapid; 2-3 days

Waxy, white, velvety lemon color with age

Reverse: white

Microscopic: thick walled, round macroconidia; differentiate from Histoplasma by lack of yeast phase (37 C)

Conditions: n/s, contaminant

Uniserate phialides

1 level of phialides and then conidia

biserate phialides

two levels of phialides, then conidia

Rhizopus spp

Mucor spp

Cunninghamella spp

Aspergillus niger

Aspergillus niger microscopic

Aspergillus flavus

Aspergillus flavus microscopic

Aspergillus fumigatus

Aspergillus fumigatus microscopic