Experiment No. 4 - ROUTINE STOOL EXAMINATION: MICROSCOPIC

Question 1

The microscopist should be familiar with the different structures found in the feces such as

Answer 1

trophozoites and cysts of Amoeba , helminth eggs and larvae macrophages, WBC, fungi, plant cells (pollen grains and spo , RBC, res), epithelial cells, crystals like Calcium oxalate and triple phosphate, bacteria, plant fibers, root hairs and animal cells are similar to helminth ova.

Question 2

Most common errors in the identification of parasites may be due to the following:

Answer 2

(1) Lack of familiarity with the parasites
(2) Cursory examination of the slide
(3) Confusion of parasite with artifacts.

Question 3

This is the simplest and moderately efficient procedure for examinations of feces.

Answer 3

Direct Fecal Smear (DFS) or Direct Wet Mount Technique

Question 4

The (?) of the intestinal tract usually results in an even distribution of parasites in stool.

Answer 4

mixing action

Question 5

DFS preparations from (?) specimens should be mandatory.

Answer 5

soft, loose and watery

Question 6

It is highly recommended for the detection of (?) in liquid, diarrheic and bloody mucoid specimens, however, if the number of organisms are few, this method may be of low sensitivity and insufficient to reveal their presence.

Answer 6

motile trophozoites

Question 7

Direct Fecal Smear (DFS) or Direct Wet Mount Technique

1. Comminute approximately (?) of feces with a drop of NSS on a clean slide using an applicator stick until a uniform suspension without fibers or gritty materials obtained.
2. Place the cover slip above the preparation and immediately examine under the LPO of the microscope in a (?) (figure A). Confirm identification under HPO.
3. Wet mounts may be preserved for several hours by ringing the cover glass with (?).

Answer 7

2 mgsy
stematic method
paraffin or nail polish

Question 8

The amount of feces used for the direct mount is important. The (?) recommended approximates what would form a low cone at the end of a wooded applicator stick.

Answer 8

2 mg

Question 9

When less than 2 mg of feces is used, the suspension will be (?) and may have (?), whereas the use of more than 2 mg results in a suspension that is and parasites may be (?).

Answer 9

too thin
blank spaces

too thick
hidden under fecal debris

Question 10

(?) may be used with wet mount preparations to aid in location and identification of protozoa but are not necessary for eggs and larvae.

Answer 10

Temporary stains

Question 11

(?) are the most commonly used temporary stains and are most useful for recognition of cyst stage; however, they kill and distort the trophozoites.

Answer 11

Dilute iodine solutions

Question 12

In cysts, the visibility of (?) is enhanced so that their number and morphological features are more clearly seen.

Answer 12

nuclei

Question 13

The use of (?) is not recommended since they do not stain organisms well.

Answer 13

weak iodine solutions

Question 14

Likewise, the use of an (?) will stain the organisms so darkly that morphological features cannot be seen.

Answer 14

iodine solution that is too strong

Question 15

Dilute iodine solutions that are recommended are (?).

Answer 15

D’ Antoni’s, Dobell and O’Connor’s and Lugol’s

Question 16

This technique is based on the fact that when a layer of stool is in contact with glycerin-soaked cellophane for a period of time, it becomes clarified and helminth eggs become visible.

Answer 16

Kato - Thick Smear Preparation

Question 17

Kato - Thick Smear Preparation

1. Place a pea-size stool sample at the center of glass slide and cover with a square piece of (?).
2. With the aid of a (?), press the cellophane gently to spread the stool, taking care that the specimen does not spread beyond the cellophane cover, the cellophane also serves as a cover slip.
3. Leave the prepared slide at room temperature for (?). During this time the microscopic field becomes clear due to the action of glycerin on the stool constituents.
4. The slide should be examined (?). Allowing the slide to stand for long period of time will cause drying and shells of hookworm ova will become be difficult to see.

Answer 17

pre-treated cellophane
rubber stopper
10-20 minutes
after 10-20 minutes or within one hour after preparation

Question 18

virtually ensures the detection of even small numbers of organisms that would go undetected if only direct smears of permanent-stained slides were examined.

Answer 18

Formalin - Ether Concentration Technique (FECT)

Question 19

Concentration procedures generally fall into two categories - (?).

Answer 19

flotation and sedimentation

Question 20

(?) can be achieved by simple gravity or centrifugation.

Answer 20

Sedimentation

Question 21

Since the sediment will generally contain all the parasites occurring in the stool sample, this procedure has greater diagnostic sensitivity.

Answer 21

Sedimentation

Question 22

An added advantage is that the technique can be readily applied to both fresh and preserved feces.

Answer 22

Sedimentation

Question 23

Formalin - Ether Concentration Technique (FECT)

1. Thoroughly comminute approximately [?] of fresh feces in [?] in a suitable container. Let it stand for [?] or longer to achieve adequate fixation. For very loose or watery fecal sample, use [?] of material.
2. Strain the suspension through two layers of wet gauze and pour into a [?] conical centrifuge tube.
3. Fill tube with NSS; centrifuge at [?].
4. Discard the supernatant if [?]; the sediment should be resuspended then centrifuged again using NSS. Proceed to the next step only after the supernatant is clear.
5. Resuspend the sediment in 10% formalin to a volume of [?]; add [?] of ether (or ethyl acetate) then shake vigorously for [?]. If ether is used, hold tube with stopper directed away from face.
6. Centrifuge at [?]. .When tube is removed, it will be seen to consist of four layers
7. Decant tube, discard [?], mix small amount of fluid left with the remaining sediment and pipette out.
8. Prepare [?] for examination

Answer 23

1.0 to 1.5 g

10 ml 10% formalin

30 minutes

5-6 ml

15 ml

400 - 500 g for 1 – 2 minutes

cloudy

10 ml

3 ml

30 seconds

400 - 500 g for 2 -3 minutes

iodine and unstained wet mounts

iodine and unstained wet mounts

iodine and unstained wet mounts

top 3 layers

Question 24

When tube is removed, it will be seen to consist of four layers:

Answer 24

a. a top layer of ether
b. a plug of debris adhering to the wall
c. a layer of formalin
d. sediment for examination

Question 25

Recently, concern over storage and use of ether, a potentially flammable and explosive material, has led to the use of (?) as a substitute.

Answer 25

ethyl acetate

Question 26

(?) appears to be somewhat more efficient than ether when specimens contain Taenia or Hymenolepis nana eggs or Giardia cysts since there are fewer tendencies for these eggs or cysts to become trapped in the plug of debris.

Answer 26

Ethyl acetate

Question 27

However, one disadvantage is that ethyl acetate, when used as a solvent for fresh feces, is not as efficient as ether in (?) which may interfere with the microscopic examination of the sample

Answer 27

extraction of fatty or mucoid materials

Question 28

A. Direct Fecal Smear (DFS) or Direct Wet Mount Technique

Simplified:
● [?] feces to a glass slide
● Add [?] then mix w/ applicator slip then cover w/ a coverslip
● View under [?]; [?] for parasite
● Count by [?]. Take a picture of each field

Answer 28

2 mg

NSS

LPO; HPO

battlement method

Question 29

B. Kato - Thick Smear Preparation

Simplified:
●[?] feces to a glass slide
● Use.a [?]
● examine after [?]

Answer 29

2 mg

single glycerin strip

10-20 mins

Question 30

C. Formalin - Ether Concentration Technique (FECT)

Simplified:
● [?] feces to a test tube w/ cap; [?] for watery stool
● add [?] to preserve
● stand for [?]
● strain suspension in [?] of wet gauze
● put in a [?]
● fill tube w/ NSS until capacity of the tube (?)
● centrifuge for [?]
● discard supernatant (liquid NSS) if [?] by decanting unit NSS is clear
● resuspend sediments in [?]
● add [?] of ether
● shake for [?] vigorously
● centrifuge for [?]
● examine [?] (ask to sign)
● decant the tube, discard [?], leave fecal material
● prepare [?] for examination (1 drop of iodine and NSS)

Answer 30

1-1.5 grams; 5-6 teaspoon

10 mL % formalin

30 mins

2 layers

centrifuge tube w/ cap

15 mL

1-2 mins

cloudy

10% formalin

3 ml

30 secs

4 layers

3 layers

iodine and unstained wet mount