Laboratory Diagnosis Of Fungi Flashcards
Collection of specimens:
Skin specimens - clean with _________ to remove dirt, oil and surface saprophytes.
Nails - cleaned with ________. It is Usually ______; need to be _______ before inoculating to media.
Hair - obtained from _______________ ; hair can be obtained by ______,______ , or with a _______. A ________ can be helpful in locating infected areas.
Body fluids - normal sterile collection procedures.
70% alcohol
70% alcohol ; clipped; finely minced
edge of infected area of scalp; plucking, brushing; sticky tape; Wood’s lamp
Preparation of specimens for transport:
Hair & nails sent in a _________ , inside proper container.
Other specimens are usually sent _____ or ______
dry envelope
frozen; on dry ice.
Packaging of specimens - must meet _____________.
Cultures must be on _____ media (not ____).
Inside labeling information: ______,_____,_______
Outside labeling - _______________
biohazard regulations
tubed; plates
patient ID, specimen source, suspected organism.
WARNING: POTENTIAL PATHOGEN
Processing of specimen to recover fungus:
Body fluids -
CSF - should be ______ ; examine _____ _____scopically, then ________.
Pleural fluid, sputum, and bronchial aspiration - specimen must be _______ , as _____ would overgrow pathogens such as ________. The Specimens may be refrigerated up to _________
Gastric washings - same as for ______ fluids.
centrifuged; sediment; micro; inoculate media
fresh; saprophytes; H. capsulatum
2 hours.
pleural
Direct examination of specimens:
Direct exam is required on any biological material sent to lab for fungus culture.
Examine for ____,______,_____,____,____
spores, hyphae, mycelial elements, budding yeast, mycotic granules.
Direct examination of specimens:
Wet mount - good for ______; examination is done in _____ environment, so that ________ is minimal.
KOH - done on _____,_____,_____,_____,_____ specimens. KOH _____________ so fungal elements may be seen.
yeast
natural; loss of fragile structure
skin scrapings, hail, nails, sputum, vaginal
clears tissue cells
Processing of specimen to recover fungus:
Body fluids - (continued)
Genito-urinary specimens - _______ specimen preferred; should be _____ and _______ on media.
Blood/bone marrow - _________ to _____ and _____
Wound abscess or drainage - culture _____ if actinomycosis is suspected.
Tissue specimens - examine for _____,______, or ______; mince _____cally, inoculate on media.
first morning ; centrifuged & inoculated
inoculate to BHI broth and BHI slant.
anaerobically
pus, caseous material or granules
asepti
Staining
______________________ - very popular for quick evaluation of fungal structures; stains chitin in cell walls of fungi.
____________________ - stains polysaccharide in the cell wall of fungi.
_____________________ Stain - silver nitrate outlines fungi in black due to the silver precipitating on the fungi cell wall.
Lactophenol Cotton Blue
Periodic Acid - Schiff Stain (PAS)
Gomori Methenamine Silver
With Periodic Acid - Schiff Stain (PAS), Fungi stain ______ with ____ nuclei.
pink-red
blue
Staining
Stains used: (continued)
___________ Stain - extremely specific method of detecting fungi in tissues or fluids.
_________ Stain - good for initial differentiation of dimorphic fungi. Works well on sputum smears.
Fluorescent Antibody
Papanicolaou
In Gridley Stain - Hyphae and yeast stain _______ or ____ . Tissues stain ________ and background is ______.
dark blue or rose
deep blue ; yellow
Mayer Mucicarmine Stain - will stain capsules of Cryptococcus neoformans _________
deep rose.
Gram Stain - generally fungi are gram _______; but ______ and _____ are gram variable.
positive
Actinomyces and Nocardia
Staining
Stains used: (continued)
Acid-Fast Stain - used to differentiate the __________ from other ________
Giemsa Stain - used for _____ and ______ specimens.
India Ink - demonstrates the _______ of ________ in _____ specimens.
acid-fast Nocardia; aerobic Actinomyces.
blood and bone marrow
capsule of Cryptococcus neoformans; CSF
Fungal Culturing:
Culture
______ media is used rather than _____ media.
Tubed
plated
Fungal Culturing:
Culture
Tubed media is used rather than plated media.
Why
Less chance for _______
Less chance for ______.
Easier _______.
spore release into the environment.
dehydration
storage
The agar in a tube is inoculated in a ________.
Preliminary identification is based on __________ on various media.
straight line
differential growth patterns
Fungi culture media
__________ agar - classic medium, recommended for most studies.
Sabouraud’s dextrose
Sabouraud’s dextrose agar with ______ which inhibits bacterial growth
chloramphenicol
______ agar - a commercially produced agar containing chloramphenicol to inhibit bacterial growth, and _______ to inhibit saprophytic fungi and some yeasts (including C. neoformans).
Mycosel
cycloheximide
________ and _______ are opportunistic pathogens. Cycloheximide will prevent their growth. Bacteria-like fungi (such as _________ ) are inhibited by chloramphenicol.
Aspergillus and Scopulariopsis
Actinomycetes
BHI- ______
Brain heart infusion
Which is more enriched
BHI or Sab-Dex?
Brain heart infusion slant (BHI) is more enriched than Sab-Dex.
BHI is used For recovery of H. capsulatum.
T/F
T
______ and _________ agar are used in slide cultures to induce spore formation, which aids in identification.
Potato-dextrose agar (PDA) and Corn-meal
Special media
Caffeic Acid Agar - Cryptococcus neoformans will produce ______ resulting in _____ colonies (must protect media from ______).
Birdseed Agar - used to _________ from contaminated cultures.
melanin ; black; light
isolate Cryptococcus neoformans
Special media
KT Medium & Kelley Agar - used to convert dimorphic fungus ________ from _____ to _____ form.
Blastomyces dermatitidis
mycelial to yeast
Special media
Modified Converse Liquid Medium (Levine’s) - used to promote ______ production by _________
spherule
Coccidioides immitis.
Incubation
Fungal growth requirements -
Temperature -____ temperature (__-__C.) for most fungi.
Nocardia sp. and some dimorphic organisms grow best at ___ degrees C.
room; 25-30
37
Any fungus capable of growing at 37 C, should be considered potentially __________.
pathogenic
Incubation Atmosphere
True fungi is generally _______ but there are some ______ among the _____ fungi
Aerobes
Anaerobes; bacteria- like
Incubation time:
Generally, cultures are held for ______
4 weeks
Some yeasts grow overnight
T/F
T
Saprophytes are slow growers( several days)
T/F
F
Fast growers (several days)
Incubation time
Paracoccidioides brasiliensis may require ______ weeks
4-5
Incubation time
____ weeks are recommended if Histoplasma capsulatum is suspected.
10
Colony Morphology
Surface topography - some fungal colonies may be ____ growing, covering the _________; others may grow in a ______ manner.
free; entire surface of agar
restricted
Colony morphology
Surface texture - cottony or wooly (floccose), granular, chalky, velvety, powdery, suede, silky, glabrous (smooth, creamy), or waxy.
T/F
T
Colony Morphology (continued):
Pigmentation - Fungi may be colorless or brightly colored. Color may be on ______ itself, on its ______ apparatus, on the _____, or _______of the colony (reverse pigmentation).
fungus
sporulating
agar; on the bottom
Mycelium -
Vegetative mycelium - provides _________.
Aerial mycelium –_________.
nutrition
reproductive
Biochemical studies - used to ID yeast and yeast- like organisms.
Carbohydrate fermentation -
____________ of a carbohydrate under ———— conditions as determined by _____ and _____ production.
Growth and utilization
anaerobic
acid and gas
Biochemical studies - used to ID yeast and yeast- like organisms.
Carbohydrate fermentation -
Specimen is inoculated ________.
__________ is the indicator.
Acid production turns _____ to ______.
Gas is detected by ____________ trapped in the fermentation tube.
Observe every _______ for ———-
beneath the broth
Bromcresol purple
purple to yellow.
appearance of bubbles
48 hours for 14 days.
Biochemical test
Carbohydrate assimilation -
Determines ability to ______________
utilize a carbohydrate as sole source of carbon.
Biochemical test
Carbohydrate assimilation -
Bromcresol purple indicator turns from _____ to ______
Tubes unchanged (as determined by __________) after _____ days are negative.
purple to yellow.
comparing to a blank tube; 10
API;_________ for yeast identification
Auxocolor panels
Biochemical studies - (continued)
Nitrogen assimilation -
Utilizes ____ tubes with differing sources of nitrogen.
___________ is the indicator
Color change of _____ to _____ is positive
3
Bromthymol blue
blue to yellow
Growth on specific agars:
______________ agar - Urea is hydrolyzed by some yeast to form ammonia (pH increases) which turns media from _______ to _______
________ medium (must protect media from ____) - Production of melanin by Cryptococcus neoformans resulting in black colonies.
Christensen’s urea
yellow to dark pink.
Caffeic acid; light
Germ tube -_______ and ______ produce germ tubes when incubated in a protein medium.
Candidia albicans & Candidia stellatoidea
Demonstration of chlamydospores - Yeast is inoculated by jabbing appropriate agar (___________ with _______) and observed every 24 hours for 3 days for chlamydospore production.
Cornmeal with tween 80
Serology
Hypersensitivity
Skin tests - demonstrates _________ to a fungus.
Serological tests - demonstrates __________ to a fungus; sera should be drawn in pairs (______ and _____).
T-cell immunity (cellular)
B-cell immunity (humoral)
acute and convalescent
Serology
Hypersensitivity: ( _____ tests and _____ tests)
skin
serological
Which is faster,
Culture or serology
Serology is Faster compared to culture
