The document describes various culture media and methods used for growing fungi. It lists different types of basal, nutritionally deficient, enriched/selective, and biochemical test media. Specific media such as Sabouraud's dextrose agar, corn meal agar, and bird seed agar are also described. The document outlines procedures for culturing fungi from samples like blood, tissues, swabs, urine, and peritoneal fluid. Incubation typically occurs at 25-30°C initially and sometimes at 35-37°C for dimorphic fungi.
3. Molds and yeasts are widely distributed in air, dust,
fomites and normal flora.
Humans are relatively resistant.
Fungi are relatively nonpathogenic
4. A)BASAL MEDIA
1)SABOURAUDS DEXTROSE AGAR
2)NEUTRAL SABOURAUDS DEXTROSE AGAR
3)SDA WITH ANTIBIOTICS
B)NUTRITIONALLY DEFICIENT MEDIA
1)CORN MEAL AGAR
2)RICE STARCH AGAR
C)ENRICHED/SELECTIVE MEDIA
1)BRAIN HEART INFUSION AGAR
2)BI PHASIC MEDIUM
3)CYSTEIN HEART AND Hb AGAR
4)BLOOD AGAR
5)BIRD SEED AGAR
6)LJ MEDIUM
7)DERMATOPHYTE TEST MEDIUM
5. 8)CZAPEK`S DOX AGAR
D)MEDIA FOR STIMULATION OF
ASCOSPORE OF PERFECT FUNGI
1)ALPHACEL-YEAST EXTRACT AGAR
2)SOIL EXTRACT AGAR
E)MEDIA USED FOR BIOCHEMICAL
TESTS
1)TETRAZOLIUM REDUCTION MEDIUM
2)CARBOHYDRATE FERMENTATION MEDIA
3) CARBOHYDRATE ASSIMILATION MEDIA
4)UREASE MEDIUM
5)DIAZONIUM BLUE B REACTION
6. PEPTONE
-10 gm
AGAR
-20 gm
DEXTROSE
-40 gm
DISTILLED WATER
-1000ml
Autoclave at 121*c for 15 min.
Adjust pH to 5.5
Saprobic fungi may overgrow
It obscuring real pathogen
7.
8. INGREDIENTS
CORN MEAL
AGAR
DISTILLED WATER
TWEEN 80
-8 gm
-4 gm
-200 ml
-2 gm
9. A Heavy inoculum of yeast is streaked across a
plate containing the medium.
Cover slip is placed over it.
Streak should project beyond cover slip.
Examine under low power at edge of cover slip.
It is a sort of junction of aerobic and anerobic
condition.
Clamidiospores are best found in this area.
Shows clamidiospores seen in candida albicans
after 24-48 hrs incubation at 25*c
10.
11. Used for growing fastidious pathogenic fungi such
as
-Histoplasma capsulatum
-blastomyces dermatitis
INGREDIENTS
Brain heart infusion agar
Glucose
L cysteine hydrochloride
Agar
Distilled water
-37 gm
-20 gm
-1gm
-20gm
-900gm
12. Dissolve ingredients by boiling.
Dispense into screw capped bottles.
Autoclave at 121*c for 15 min
Cool in slanted position with one inch butt
pH adjusted to 6.7
Store in refrigerator
13. For cryptococcus neoformans
Can utilise creatinine as a source of nitrogen
Colonies are brown to black due phenoloxidase
produced by organism.
14. Niger seed ectract
Glucose
Chloromphenicol
Gentamicin
Dipheny solution
Agar
Distilled water
Autoclave at 121*c for 15 min.
Dispense into plates
-200ml
-1 gm
-400gm
-25 mg
-10ml
-20gm
-800ml
15. Used for
Histoplasma capsulatum
Blastomyces dermatitidis
Cryptococcus neoformans
Ingredients
Blood agar base
Sheep blood
Distilled water
-40gm
-50ml
-1000ml
17.
Sabrourd’s medium is used world-wide and is
generally satisfactory
Some workers prefer malt peptone agar
It is claimed that, in the latter from the syrup is
slightly more inhibitory to bacteria and produces
more rapid growth and sporulation of fungi than
sabouraud’s medium. In additional it is often
possible to distinguish mixed cultures of yeast
species by their colony morphology on malt agar
Two types of containers are used for culture
1.
Petri dish
2.
Culture tube
19. Blood culture
• Same as that of microbiology
• For manual system blood culture bottle with
agar and mycological broth or sabouraud
broth media may be employed
• Aerat the cultures periodically by shaking and
sub culture routinely rather than to wait until
the medium is cloudy
20. TISSUE
Biopsy and other tissue should be reduced in
size 1 -2mm
Put these pieces into agar medium, it should
contain antibiotics if the material is
contaminated with bacteria
21. SWABS
o
o
Heavily feed swab rotate over the surface of media several times
Secondary dilution strokes should be made with a sterile loop
CSF
o
A loop full of spun deposit should be take to inoculate the
agar media in usual manner
22. Urine
Peritoneal fluid
Spread 0.1ml un concentrated
Process as urine but take an
urine over the surface of agar
containing antibiotics
Centrifuge the specimen and
remove a loop full of
sediment to another agar
plate of the same medium
and streak out from the well
in the normal way
additional sample of 1ml of
the neat dialysate and spread
over a plate containing
mycological medium
.
23. INCUBATION
Most fungi and moulds grow at room temperature (25
30*c)
Some at body temperature (35-37*C)
Some are dimorfhic fungi
All media are incubated at 25˚ to 30˚C initially and,
when a potential dimorphic organism is isolated an
attempt is made to convert it to the tissue phase by
subeultuning it and incubating the new set of culture at
35˚C
The pathogenic fungi are aerobic organisms. A good
supply of oxygen is mandatory if they are to be isolated
in primary culture
