Mycology

IntroductionIntroduction Mycology is study of fungi and fugal infection.

Fungi are define as Eukaryotic organism, consisting of microscopic branching filaments called hyphae carrying spores.

Or as unicellular yeast.

Fungi are found every where as saprophytes, may found in soil, vegetable, water and disseminated in air or as pathogenic.

Classification of fungiClassification of fungi

(1) Morphological classification 1- Filamentous form. 2- Yeast. 3- Dimorphic.

(2) Clinical classification 1- Superficial mycosis. 2- Subcutaneous mycosis. 3- Systemic mycosis. 4- Mycotoxicosis .

Yeast IdentificationYeast Identification

Candida is present as normal flora in the mouth, GIT and vagina.

Species include:

o Candida albicans (the most important)o Candida tropicaliso Candida kruseio Candida parapsilosiso Candida glabratao Candida dubliniensis

DiseasesIn normal host, candida albicans may cause:1)Oral thrush or moniliasis (patches of creamy

white exudate that cover the mucous membrane of the mouth).

2)Vulvovaginal moniliasis: itching and cheese appearing vaginal discharge. It occurs in females with:o Prolonged use of broad spectrum antibiotics

(superinfection)o Contraceptive pillso Pregnancy

3)Rash in the skin folds4)Diaper rash.5)Paronychia of the nails (thickening and loss of

the nails).

In immunocompromised individuals:

1)Esophageal thrush.2)Chronic mucocutaneous candidiasis.3)Systemic or disseminated candidiasis (even to the eye).

Specimen: It is according to the site of the lesion i.e. exudates, vaginal, oral swab, tissue, nail scrap, blood.

1.1. Microscopic ExaminationMicroscopic Examination

1- Stained smears: With Gram stain, yeast cells are seen as Gram positive, oval budding yeast cells attached to pseudohyphae.

2- 40% KOH: budding yeast, pseudohyphae.

Laboratory diagnosis of Laboratory diagnosis of Candida albicansCandida albicans

2. Culture: Grow on Sabouraud’s dextrose agar (SDA) at 37 0C.

Identification of candida on the plate is done by:

a.Morphology: Soft cream colored colonies with yeasty odor.

b.Gram film: Gram positive oval budding yeast cells.

3. Biochemical reactions: To differentiate between Candida albicans and other species.

a. Germ tube test: Candida albicans forms germ tube when incubated in serum for 1 – 2 hour at 37 0C.

b. Chlamydospore formation: Candida albicans forms chlamydospores on corn meal.

c. Sugar fermentation: Candida albicans ferment glucose and maltose with acid and gas production.

d. Inoculation of the yeast on chromogenic agar: Each candida specie produces a different color on this medium.

Germ tube Chlamydospores

CHROMagar

GTT: Germ tube testGTT: Germ tube test::

(a ) 3 2-3 Inoculate ml of horse serum & incubate for hrs.

(b ) 10 , Put drop of preparation on slide & screen with X

40 . . X for tube like structure i e germ tube.

GTT positive> production of germ tubes in plasma

after 3 hours incubation at 370C. C.

albicans.

Germ tube negative> Budding blastoconidia only

are seen.

other Candida species .

ProcedureProcedure::Using clean, dry and sterile test tubes.

Using a Pasteur pipette, dispense 3 drops of fresh

pooled human serum into the tubes.

Serum can be obtained from the Serology lab.

With a sterile wooden applicator stick, lightly touch a

yeast colony and place the stick into the serum.

Suspend the yeast in the serum. Discard the stick in a

discard container.

Incubate the test at 35oC for 2.5-3 hours.

Place a drop of the suspension on a clean microscope

slide.

Place a clean cover glass over the suspension and then

examine it with a microscope using the low power

objective.

Use the high power objective to confirm the

presence or absence of germ tubes.

Read controls and record results.

If time is not allowed to read and record the test

results add a drop of 10% formalin/formaldehyde

to each tube, seal the top with parafilm, and

store at 2-4 in the refrigerator.

A: +ve GTT B: A: +ve GTT B: -ve GTT-ve GTT