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[MT 6322] Mycology & Virology Laboratory > [2S] Exercise 6: Laboratory Diagnosis of Common Fungal Diseases > Flashcards

[2S] Exercise 6: Laboratory Diagnosis of Common Fungal Diseases Flashcards

1
Q

constitute a major part of the ecosystem and are also known potential pathogen in human and animals

A

Fungi

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2
Q

are specialized fungi that produce the enzyme keratinases that degrade keratin for their nutrition

A

Keratophilic fungi

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2
Q

Soil rich in _______ materials favours the growth of these fungi

A

keratinous

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3
Q

specific method for isolation of keratophilic fungi in soil

A

Hair Baiting Technique

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3
Q

PRINCIPLE: Temperature, pH, soil humidity, texture, as well as depth of soil are the climactic factors to be considered for cultivation of keratophilic fungi. Human hair is a keratin-rich substrate often used as bait to trap and isolate keratophilic fungi

A

Hair Baiting Technique

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4
Q

source of nutrition

A

Keratin

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5
Q
  • Substrate
  • Rich in keratine
A

human hair

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5
Q

Mainly composed of Hyphomycetes

A

Keratinophilic fungi

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6
Q

is a keratin-rich substrate often used as bait to trap and isolate keratophilic fungi

A

human hair

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7
Q

Media used for Hair Baiting

A

Saboraud Dextrose agar (with cycloheximide, penicillin and streptomycin)

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8
Q

HAIR BAIT PROCEDURE

T/F: The collected hairs should be washed thoroughly with water then air dry at room temp

A

T

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9
Q

HAIR BAIT PROCEDURE

T/F: Cleanse the collected hair baits with 70% alcohol. Air dry

A

T

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10
Q

HAIR BAIT PROCEDURE

Sterilized the cleaned hair by autoclaving at ____ for __ min

A

15 lbs./sq inch; 20 min

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11
Q

HAIR BAIT PROCEDURE

Sterile petri dishes are then half-filled with soil and moistened with _____ of sterile distilled water

A

15 to 30 mL

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12
Q

HAIR BAIT PROCEDURE

Cover the plates and incubate the preparation at _______ in a ______ or cupboard for about 1-2 months. Soil may be moistened when necessary

A

room temperature (20—25°C); dark place

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13
Q

HAIR BAIT

Examine the plates periodically for the development of ______ on the hair filaments. This can be done by taking a small amount of colony on hair bait, mount on a clean slide with saline or LPCB and examine the preparation under the microscope

A

mycelium

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14
Q

HAIR BAIT

After fungal colonies have grown around the hair, subculture the colonies in a ____________ medium containing cycloheximide.

A

Sabouraud Dextrose agar

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15
Q

HAIR BAIT

The keratinophilic fungi isolated and identified in this technique, particularly if it is a _____________ or its variant, may be used as test fungi for the Hair Perforation test

A

Trichophyton spp.

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16
Q

The ability to perforate hair and utilize its keratin content through digestion by its keratinase enzymes is evident in the majority of the dermatophytes

A

Hair Perforation Technique

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17
Q

An in vitro hair perforation test used to differentiate
keratophilic dermatophytes, particularly the Trichophyton spp. and its variant

A

Hair Perforation Technique

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17
Q

HAIR PERFORATION

Positive control strains

A

Trichophyton mentagrophytes

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18
Q

The ability to perforate hair and utilize its keratin content through digestion by its ______ enzymes is evident in the majority of the dermatophytes

A

keratinase

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19
Q

PRINCIPLE: Petri dish containing sterile water and yeast extract is inoculated with test fungi and human hair samples

A

Hair Perforation Technique

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20
Q

PRINCIPLE: After 7 days of incubation at room temperature, hair is examined microscopically and observed for deep, narrow wedge-shaped tears or slashes indicative of presence of hair perforations

A

Hair Perforation Technique

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21
Q

HAIR PERFORATION

Negative control strains

A

Trichophyton rubrum

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22
Q

HAIR PERFORATION PROCEDURE

Collect about 10-20 pieces of hair samples and cut it into short pieces about ___ in length

A

1 cm

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23
Q

HAIR PERFORATION PROCEDURE

Transfer the sterile short pieces of hair into a screw
capped tube containing ___ of sterile water

A

5 mL

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23
Q

HAIR PERFORATION PROCEDURE

Incubate the tubes at _______

A

room temperature

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24
Q

HAIR PERFORATION PROCEDURE

Sterilize the cut hair samples by autoclaving at ___C for ___ minutes and store in a sterile container

A

121 °C for 10

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25
Q

HAIR PERFORATION PROCEDURE

After _ days of incubation, take an aliquot amount of
the hair sample and place in a clean glass slide with
lactophenol cotton blue (LPCB) stain.

A

7

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25
Q

HAIR PERFORATION

T/F: Steps 6 and 7 may be repeated at intervals of 7
days until the 4th week.

A

T

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26
Q

HAIR PERFORATION PROCEDURE

After 7 days of incubation, take an aliquot amount of
the hair sample and place in a clean glass slide with
_______ stain.

A

lactophenol cotton blue (LPCB)

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26
Q

HAIR PERFORATION

can be used as positive control and should be processed and tested in the same manner as that of the specimen or test sample

A

Prepubertal hair or child’s scalp hair

27
Q

INTERPRETATION OF RESULTS

Hair inoculated with isolates of _______ _______ produce marked localized areas of pitting and marked erosion or deep, narrow wedge-shaped scars or slashes indicative of presence of hair perforations

A

POSITIVE: Trichophyton
mentagrophytes

28
Q

INTERPRETATION OF RESULTS

Absence of deep, narrow wedge-shaped perforations on surface of hair sample inoculated with _______ _____

A

NEGATIVE: Trichophyton rubrum

29
Q

RAPID TEST FOR DIAGNOSIS OF FUNGAL DISEASES

  • non-culture-based diagnostic tool
  • Presumptive
A

Detection of patient antibodies (serologic tests)

30
Q

RAPID TEST FOR DIAGNOSIS OF FUNGAL DISEASES

Standard method for diagnosing cryptococcosis, and to some extent, aspergillosis, and candidiasis

A

Antigen detection test

30
Q

T/F: Establishing the presence of antibodies to fungi in serum/plasma and spinal fluid or the direct detection of the fungal antigens themselves, can be vital in the diagnosis of bacterial disease

A

F; mycotic disease

31
Q

Caused by opportunistic fungi C. neoformans and C. gattii

A

Cryptococcosis

31
Q

RAPID TEST FOR DIAGNOSIS OF FUNGAL DISEASES

Routine serologic assay identifying many fungal diseases including coccidioidomycosis, histoplasmosis, aspergillosis

A

Antibody detection

32
Q

RAPID TEST FOR DIAGNOSIS OF FUNGAL DISEASES

3 Important serologic techniques for the rapid diagnosis of fungi

A
  • Immunodiffusion (ID)
  • Complement Fixation (CF)
  • Enzyme immunoassay (EIA)
33
Q

Cryptococcosis MOT

A

inhalation of dust from pigeon droppings or bird feces contaminated with fungi

33
Q

A latex agglutination test wherein a carrier particle such as latex coated with antibodies specific to Cryptococcus capsular antigen

A

Cryptococcal antigen test

34
Q

T/F: At risk of Cryptococcosis are individuals with lowered or impared cell-mediated immune function causing meningoencephalitis, pneumonia, septicemia

A

T

35
Q

Cryptococcosis cause (3)?

A

Meningoencephalitis, pneumonia, septicemia

36
Q

Cryptococcal antigen test is mixed with what specimen?

A

CSF / Serum / Plasma

37
Q

Reagent used for Cryptococcal antigen test

A

Pronase reagent

37
Q

Positive result for Cryptococcal antigen test

A

formation of visible clumps or aggregates

38
Q

To detect Cryptococcal capsular polysaccharide antigen could be taken as the early diagnostic method of cryptococcal meningitis

A

Latex agglutination test

39
Q

CAT PREPARATIONS: CSF

Centrifuge specimen at _____ g for __ mins

A

1000 x; 15 mins

40
Q

CAT PREPARATIONS: CSF

Using a serologic pipette, aspirate ____ of CSF and place it in a sterile glass tube

A

0.5 or 1 mL

41
Q

CAT PREPARATIONS: CSF

Inactivate the CSF sample in a water bath set at __C for _ minutes. Allow to cool for ___ mins before testing

A

56C; 5 mins; 3-4 mins

42
Q

CAT PREPARATIONS: CSF

Prepare 1:100 dilution of CSF sample by mixing ___ of the heat inactivated specimen with ____ of sterile normal saline

A

25 uL; 2475 uL

43
Q

CAT PREPARATIONS: CSF

Prepare _____ dilution of CSF sample by mixing 25 uL of the heat inactivated specimen with 2475 uL of sterile normal saline

A

1:100

44
Q

CAT PREPARATIONS: SERUM

In a clean glass tube, place ____ of serum and add ___ of pronase reagent.

A

300 uL; 50 uL

45
Q

CAT PREPARATIONS: SERUM

Centrifuge ______ at 1000x g for 15 mins.

A

clotted blood

46
Q

CAT PREPARATIONS: SERUM

Mix the specimen and reactant by using a ______.

A

vortex mixer

47
Q

CAT PREPARATIONS: SERUM

Inactivate the mixture by heating it in a water bath set at __C for ___ minutes.

A

56C, 30 mins

48
Q

CAT PREPARATIONS: SERUM

Prepare 1:100 dilution of CSF sample by mixing ___ of the heat inactivated specimen with ____ of sterile normal saline

A

25 uL; 2475 uL

49
Q

CAT TEST PROCEDURE

T/F: 1. Label the test/reaction areas of the disposable card as follows: one for positive control, one for negative control, one for the undiluted heat inactivated specimen, and one for 1:100 dilution of clinical specimen (e.g., CSF, serum/Plasma)

A

T

50
Q

CAT TEST PROCEDURE

  1. On separate areas on the disposable card slide, place __ uL of heat inactivated specimen, __ uL of the 1:100 diluted specimen, and __ uL of positive and negative controls respectively. Spread each sample on the labeled reaction area.
A

25

51
Q

CAT TEST PROCEDURE

Add __ uL of latex reagent in each of the four labeled wells.

A

25

52
Q

CAT TEST PROCEDURE

Mix well the specimen-latex reagent mixture using a disposable _____, making sure that the mixture covers the entire reaction area.

A

stirrer

53
Q

CAT TEST PROCEDURE

Continue mixing by rotating the reaction card at ___ rpm for __ minutes using a mechanical rotator

A

100 rpm, 15 mins

54
Q

CAT TEST PROCEDURE

T/F: All positive test results should be doubly titrated and maximum titer showing agglutination reported

A

T

55
Q

CAT INTERPRETATION OF RESULT

Denotes presence of clumping or granular aggregates, indicative of the presence of cryptococcal antigen

A

Positive

56
Q

CAT INTERPRETATION OF RESULT

Presence of homogenous suspension

A

Negative

57
Q

Principle of Rapid Cryptococcal Antigen Test

A

Immunochromatographic assay

58
Q

will bind to the gold-conjugated antibodies

A

CrAg

59
Q

gold-conjugated antibody-CrAg complex moves up the test strip through ______ ______

A

capillary action

59
Q

gold-conjugated antibody-CrAg complex is captured/immobilized with ________ ________ against CrAg, forming a red band

A

monoclonal antibodies

60
Q

Specimen used for RCAT

A
  • Cerebrospinal fluid (CSF)
  • Serum/Plasma
  • Urine
60
Q

RCAT PROCEDURE

  1. On a clean kahn test tube or ____ tube, add 1 drop of specimen diluent
A

Eppendorf

61
Q

RCAT PROCEDURE

  1. Using a disposable dropper pipette, place ___ (1 drop) of the patient specimen to the tube.
A

40 uL

62
Q

RCAT PROCEDURE

  1. Insert the ______ strip into the tube
A

lateral flow assay (LFA)

63
Q

RCAT PROCEDURE

  1. Immerse the strip into the specimen for ____
A

10 minutes

64
Q

RCAT INTERPRETATION OF RESULT

Absence of red line or a line only at the test region

A

Invalid

65
Q

RCAT INTERPRETATION OF RESULT

Presence of two red lines-one at the control region and the other at the test region

A

Positive

66
Q
A

Invalid

66
Q

RCAT INTERPRETATION OF RESULT

Only one red line at the control region

A

Negative

[MT 6322] Mycology & Virology Laboratory (10 decks)

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