MICROSCOPIC STUDY OF FUNGI Flashcards
It is used to observe clinical specimens for the presence of
fungal elements or to identify the fungus following cultur
Microscopy
What is the stain of choice that we use in microscopy?
LACTOPHENOL
COTTON BLUE
It is a simple cover slipped preparation
and mostly good for yeast usually or commonly used
procedure that we performed
Wet preparation/ Wet mount
What objective is a wet mount observed?
LPO
This dissolves keratin
KOH
This dissolves keratin
KOH
What prevents thr crystalization?
Glycerol
spores are seen outside the hair
Ectothrix
spores are inside the hair
Endothrix
Gives more rapid maceration and clearing than KOH
however, preparations does not last because specimen
dissolve
. Not for hair or thin scales of skin
. Specimen may dissolve
Dimethyl sulfoxide (DMSO)
This is a fluorescent dye, which binds selectively to chitin of
the fungal cell wall. The specimen then can be observed
under fluorescent microscope.
Calcofluor white
This is used for encapsulated yeasts
India Ink/ nigrosin
This is used for rapid staining of blood and bone marrow fungi. And is used for histoplasma capsulatum.
Wright’s Giemsa
- Pink to pinkish blue
- Useful for staining tissue sections
- Common stain for tissue processing
H & E (HEMTOXYLIN AND EOSIN)
- It will stain the polysaccharide in the cell wall of the
fungi - For the demonstration of glycogen
and neutral mucins. - Red/ purple against green
Periodic Acid Schiff
What is the color of PAS positive material, Nuclei, PAS digest material?
- PAS positive material: MAGENTA
- Nuclei: BLUE
- PAS digest material: COLORLESS
What is the mechanism of PAS?
Certain tissue elements are oxidized by PAS
- For tissue section
- Black with lavender-gray areas
because of silver precipitate
deposits
METHENAMINE SILVER (MS)
What is the mechanism of methenamine silver?
Chromic acid treated fungi possess aldehydes which will reduce the
hexamine-silver mixture to produce a black deposit.
What originally contains meyer’s mucicarmine?
Aluminum hydroxide
Gives more consistent results for the
detection of Cryptococcus neoformans
MEYER’S MUCICARMINE (MM)
What is the color of MEYER’S MUCICARMINE (MM) under the microscope?
An evident rose to red with nucleus
May be used to detect fungal antigen
in clinical material such a pus, blood,
CSF, tissue impression smears and in
paraffin sections of formalin fixed
tissue.
FLUORESCENT ANTIBODY STAINING
Enumerate the method under wet preparation used in microscopic study of fungi
KOH
DIMETHYL SULFOXIDE (DMSO)
CALCOFLUOR WHITE
INDIA INK/ NIGROSIN
WRIGHT’S GIEMSA
Enumerate the special stains used
H & E (HEMTOXYLIN AND EOSIN)
PERIODIC ACID SCHIFF (PAS)
METHENAMINE SILVER (MS)
MEYER’S MUCICARMINE (MM)
FLUORESCENT ANTIBODY STAINING
This is the most common stain used for microscopy of fungi
Lactophenol Cotton Blue
Describe lactic acid, phenol and cotton blue.
- Lactic acid: fixative
- Phenol: fungicide
- Cotton Blue: stain
Enumerate the stains we used in examination of results
LACTOPHENOL COTTON BLUE (LPCB)
POLYVINYL ALCOHOL (PVA-CB)
- Very common and rapid method
- Disadvantage: Distorts colonial morphology
Tease mount
Substitute for tease mount
Morphology is preserved
Cannot be preserved for long periods
The tape isn’t sterile
May cause widespread dispersion of conidia since
the procedure is somewhat messy
Cellulose tape swab
What would cellulose tape swab cause to widespread since it is messy?
Conidia
Needs tease-mount first (get small amount of
sample from the culture)
Usually perform or done when negative to conidia
The macroscopic characteristics are observed
Used for fungi with “delicate linkages”
Between conidium and conidiophore
MICROSLIDE/ AGAR BLOCK
What is the characteristic of fungi if used in MICROSLIDE/ AGAR BLOCK?
Delicate linkages
MICROSLIDE/ AGAR BLOCK is usually performed if the conidia is?
Negative
More preferred than the media since it is lesser chance that
the spores will be release in the environment
Less chance for dehydration
and it is easier to store also
Germ test tube
What is the characteristic of germ test tube?
Germ tube is a short outgrowth, non-septate germinating
hypha.
From what microorganism is yeast differentiated to using the germ test tube?
C. Albicans
An initial filamentous outgrowth from a conidia or spore
An asexual reproductive structure
Germ tube
Enumerate the preparation of microscopic examination
Tease mount
Cellulose tape swab
Microslide agar/ block
Germ test tube
Germ tube
How many drops of formaldehyde are absorbed by cotton then placed in a tube?
10 drops
In sealing with parafilm, how long is the interval of culture?
6-12 months
How long is the interval of mineral oil?
24
How long is the interval of freezing?
@ –20°C to –70°C - 1 to 2 years interval
In preserving using sterile water, how many mL of distilled water in culture?
2mL
Monoclonal antibody labeled with
fluorescent dyes can be used to detect several fungi in the
clinical specimens. It is the most commonly used
Immunofluorescence
Detection of anti-fungal antibody is helpful in diagnosis of
sub-cutaneous and systemic mycoses, prognosis and
response to anti-fungal drugs
Serology
Enumerate the different serologic techniques
- Agglutination
- Immunodiffusion
- Counter-immunoelectrophoresis
- Complement fixation test
- Immunofluorescence
- RIA (Radioimmunoassay)
- ELISA (Enzyme-Linked Immunosorbent Assay)
