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LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
Practi cal No. 3
(L a b o r a to r y e xa m i n a ti o n o f fu n g i )
B. Yeas t s
Ob j e c t i v e s :
1- Describe the general structures of the yeasts.
2- To be familiar with the most common methods to identify the yeasts.
Gen er al s t r u c t u r es :
1- Yeast cells usually grow as la r g e , s in g le c e lls . Most reproduce by an asexual
process called ''b u d d i n g '' .
2- Many types of yeast also produce structures called ''p s e u d o h y p h a e '' .
3- Another structure produced by a few species of yeasts is the ''g e r m tu b e '' . It
is a tubal-like structure that grows from the surface of the yeast cell. It has
parallel, non-constricted walls. Germ tubes are seen in direct examination of
specimens or following growth of yeast cells in media that stimulates their
production.
Fi gu r e 3 . 1 : Yeast cells
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Fi gu r e 3 . 2 : Germ tube
Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
Fi gu r e 3 . 3 : Pseudohyphae
Ex a m i nat i o n o f ye as t s :
I.
Ma c r o s c o p i c e x a mi n a t i o n o f y e a s t s :
Yeast colonies generally look similar to bacterial colonies. There are a few basic
elements that you can identify for all colonies:
1. Form
2. Size
3. Color
4. Elevation
5. Margin
6. Surface (texture)
7. Chromogenesis (pigmentation)
II.
Mi c r o s c o p i c e x a mi n a t i o n o f y e a s t s :
Yeasts are examined by microscope by one of these methods:
1- Gram stain.
2- Wet mount.
3- Germ tube test.
4- India ink test.
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
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LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
Yeast can be identified microscopically by the presence of:
1- Yeast cells, which may be divided by budding.
2- Pseudohyphae.
3- Germ tube.
4- Capsule arround the yeast cell.
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
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LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
1. Can di da al bi c an s
Ma c r o s c o p i c e x a mi n a t i o n :
1. For m : circular
2. Si ze : small
3. Col or : off-white
4. El e va t i o n: convex
5. Ma r g i n : smooth, entire
6. Surfa ce : buttery
Mi c r o s c o p i c e x a mi n a t i o n :
1- Gram positive, small, budding and oval in shape (Figure 3.4).
2- In stained smear, the yeast cells can often be seen attached to pseudohyphae.
3- Germ tube positive (Figure 3.5).
Fi gu r e 3 . 4 : Gram positive yeast
Fi gu r e 3 . 5 : Germ tube (C. albicans)
Ot h e r e x a mi n a t i o n :
• CHR O M agar Can di da: this medium has proven to be useful for detection of
mixed cultures of Candida species within a single specimen.
• Cor n m eal agar : presence of blasoconidia, chlamydospores or hyphae.
• API 2 0 C : depends on carbohydrate assimilation.
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
2. Cr y pt oc oc c u s n eof or m an s
Ma c r o s c o p i c e x a mi n a t i o n :
1- For m : circular
2- Si ze : small to medium
3- Col or : beige
4- El e va t i o n: convex
5- Ma r g i n : smooth, entire
6- Surfa ce : creamy
Mi c r o s c o p i c e x a mi n a t i o n :
1- Gram positive, single and budding oval or spherical yeast cells.
2- India ink: positive; the cells are surrounded by a thick capsule (Figure 3.6).
Fi gu r e 3 . 6 : Capsule (C. neoformans)
Ot h e r ex am i n at i on :
• Ur eas e t es t : positive for C. neoformans.
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
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LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
How t o i den t i f y C. al bi c an s , C. sp p a nd C. n eof or m an s ??
Gr am s t ai n
All +ve
Ger m t ube
– ve
+v e
C. albicans
C. neoformans
C. spp
API 20C for non--‐C. albicans
In d i a i n k
+v e
- ve
C. spp
C. neoformans
(capsulated)
API 20C for non--‐C. albicans
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
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LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
Worksheet
Ma t e r i a l s :
1- Yeast culture on SDA.
2- Micsoscope.
3- Microscopic slides.
4- Coverslips.
5- Pasteur pipette.
6- Tube.
7- Wooden applicator stick.
8- India ink.
9- Human serum.
Exe rci se 1 :
Perform gram stain from the given plate.
Result:
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
Exe rci se 2 :
Perform wet preparation from the given plate.
Result:
Exe rci se 3 :
Perform the india ink test. Record your result with drawing.
Result:
3rd
Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
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LABORATORY DIAGNOSIS OF FUNGI 2
Practical No. 3
Exe rci se 4 :
Perform germ tube test from the given plate. Record your result with drawing.
Method:
Step (1):
a. Using a Pasteur pipette, dispense 3 drops of fresh-pooled human serum
into the tubes.
b. With a sterile wooden applicator stick, lightly touch a yeast colony and
place the stick into the serum.
c. Incubate the tube at 35oC for 1-2 hours.
Step (2):
a. Using a pasteur pipette, place a drop of the suspension on a clean microscopic
slide.
b. Place a clean coverslip over the suspension and then examine it under the
microscope.
Result:
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Dr. Nessrin AL-abdallat
year Laboratory Medicine, 1434-1435H
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