Overview of fungal identification methods and strategies (Week 2)

Corneal Scrapings or Vitreous Humor

Clinical specimen for Eye infection

Respiratory Sample

Most common samples collected for fungal culture

Cycloheximide

Prevents overgrowth by rapidly growing molds

Room temp

Respiratory sample can be stored in _____ if processed within 2 hours

Ref temp

Respiratory samples should be _____ if delayed

CSF

Should be filtered using membrane filter

0.45um

Size of membrane filter

Daily

When should CSF cultures should be checked

False

CSF media should contain antibacterial or antifungal elements

False

CSF sample can be refrigerated

Room temp

If delayed, CSF samples should be placed in _____ (temp)

Blood

BACTEC, BacT/ALERT, VersaTREK is used in?

Malassezia

Exception for BACTEC, BacT/ALERT, VersaTREK

Isolator

Should be used for optimal recovery in blood samples

RBC and WBC w/ microorganisms are lysed

Isolator function

4 days

Most fungi in blood samples are detected in?

10 to 14 days

Days required for H. capsulatum recovery

21 days

Suggested incubation time for blood fungal culture

Corneal Scrapings

Type of sample that should be placed directly onto microscopic slides

Non-inhibitory media

Corneal scraping should inoculated to

X or C

Pattern used for corneal scraping inoculation?

Vitreous Humor

Should be concentrated by centrifugation

Sediments

Part used to smear and culture vitreous humor sample

Media containing Cycloheximide

Should be avoided for eye sample

Hair, Skin, and Nail scrapings

Submitted for Dermatophyte culture

Leading edge

Part of lesions used for Skin sample

True

Hair, Skin, and Nail scraping should NOT be refrigerated

Mycosel Agar

Contains Chloramphenicol and Cycloheximide

Olive oil infused paper disk

Used for Malassezia furfur infections

First

Which quadrant should M. furfur be placed

Minimum of 21 days at 30C

Days for incubation of cutaneous samples

Vaginal

Should be transported to the laboratory within 24 hours

2 swabs

Number of swabs collected for vaginal swab

Both Selective and inhibitory agar

Vaginal swab should be plated in

7 days at 30C

Vaginal culture incubation period

Urine

Should be processed as soon after collection as possible
Media containing antibacterial agent must be used

Unacceptable

Acceptable or Unacceptable: 24 hour urine culture

72 hours

If urine transport system is used, how long can urine stay in RT?

Mincing

All tissue should be processed by ____ before culture

21 Days at 30C

Tissue sample incubation period

Bone Marrow

Collected using heparinized syringe

Sterile body fluids

What specimen should be centrifuged at least 1mL of sample before culturing

Bone marrow, Sterile fluids, and blood

Isolator tube can be used in

Acid fast and Auramine-rhodamine stain

Used for detection of mycobacteria and Nocardia spp.

Apple-green

Color of Calcofluor white stain

India Ink stain

Used for detection of Cryptococcus spp in CSF

Clear Halo

India ink appearance after 1 min

Lactophenol cotton or Aniline Blue wet mount

Most widely used method of staining fungi

Potassium hydroxide

Used for rapid detection of fungal elements

Masson-Fontana stain

Used for Examination of melanin pigment

Methenamine silver stain

Best stain for fungal elements
Used for Histologic section

Saline wet mount

Quickly performed and Cost-effective method

Wright's stain

Detects H. capsulatum & Cryptococcus spp.

Primary recovery media

Include general isolation media supplemented w/ nutrients and antibiotics
Require a Carbon source to grow

Chromogenic agar

Isolation and presumptive identification of yeast and filamentous fungi

Cornmeal agar with Tween 80 and Trypan blue

Identification of Candida albicans

Cottonseed conversion agar

Conversion of dimorphic fungus Blastomyces from mold to yeast form

Czapek's agar

Differential identification of Aspergillus spp.

Niger Seed (birdseed agar)

Identification of C. neoformans & C. gattii

Rice medium

Identification of M. audouinii

30C

Optimal temperature for incubation

Aerobic

Aerobic or Anaerobic: Most fungi are?

Paraflim/Masking tape

All plates must be sealed with

2-4 weeks

Fungal culture are incubated for

4 weeks

Minimum time needed before reported as negative

Slow growers

11 to 21 days

Immediate growers

6 to 10 days

Rapid growers

5 days or less

Verrucose

Furrowed or convoluted

Umbonate

Center slightly raised

Rugose

furrows radiate out from the center

Texture

Determined by observing a cross section of the colony

Cottony

loose, high aerial mycelium

Velvety

Low aerial mycelium

Glabrous

Smooth surface w/ no aerial mycelium

Granular

Dense, powdery, granular

Wooly

high aerial mycelium that is slightly matted down

Tease mount

Drop of LPCB in slide and spread mycelium using needle

Slide culture (Microculture)

Best method to preserve fungal morphology
Slide is put on a bent rod inside a petri dish

MALDI-TOF

Serologic testing molecular methods

Neoformans, albicans, parapsilosis, tropicalis, Coccidiodes spp., capsulatum

Recovered from CSF specimens