Stool examination

Beneficial for students

1. STOOL EXAMINATION 10/29/2021 1 Analysis of Stool Examination

2. INTRODUCTION Examination of sample of stool is easily done Useful in the evaluation of diarrheal diseases, parasitic infestations, colorectal carcinoma and malabsorption . 10/29/2021 2 Analysis of Stool Examination

3. COLLECTION CONTAINER: Stool sample is collected either in a wide mouthed glass or plastic jars with a screw cap Container should be clean and dry METHOD OF COLLECTION: Sample is transferred from a clean bed pan or toilet into the container AMOUNT OF SAMPLE : Stool required is small and about 2 -5g of the stool sample is adequate 10/29/2021 3 Analysis of Stool Examination

4. PRCECAUTIONS IN COLLECTION Morning sample is preferred Sample should be labeled and the time of collection to be mentioned Contamination with either urine or other substances in the bed pan or toilet should be avoided Stool must be fresh Examine within 1 hour of collection If blood/mucus/any other abnormal gross features are present in the stool, it should be included in the sample collected 10/29/2021 4 Analysis of Stool Examination

5. PRESERVATION Formal –saline can be used as preservative which preserves morphology of protozoa and helminthic eggs 10/29/2021 5 Analysis of Stool Examination

6. STOOL EXAMINATION PHYSICAL EXAMINATION QUANTITY : The quantity of stool varies from 100-250g/day depending on the type of diet consumed CONSISTENCY AND FORM: Normal feces is well –formed Extensively hard stool is observed during constipation Large bulky, frothy, pale, foul smelling stool which floats on water is characteristic of steatorrhea (Poor fat digestion ) Rice water stool- cholera Watery/semisolid –diarrhea, dysentry 10/29/2021 6 Analysis of Stool Examination

7. COLOR : Normal – Golden brown – stercobilin (Pigment derived from bilirubin metabolism) Black tarry stool – Usually due to altered blood in stool –melena Source of blood – Bleeding from the upper gastrointestinal tract Black tarry stool – observed in iron administration Bright red color: Due to bleeding from the lower GIT like bleeding piles Clay colored stools: observed in obstructive jaundice 10/29/2021 7 Analysis of Stool Examination

8. ODOR : Normal odor of stool is due to indole and skatole formed by intestinal fermentation and putrefaction Odor varies according to the pH of the stool BLOOD AND MUCUS IN STOOL: Observed in either amebic dysentery or bacillary dysentery PARASITES: Stool sample may show adult worms/segments of worms( eg . roundworm, pinworm, whipworm, hookworm or tapeworm) 10/29/2021 8 Analysis of Stool Examination

9. CHEMICAL EXAMINATION The chemical examination of stool includes: REACTION AND pH Normal stool pH : Ranges from 5.8 to 7.5 Strongly acidic stool : Observed with excess carbohydrate diet or fermentation due to lactose intolerance Strongly alkaline stool : observed with excess proteins in diet 10/29/2021 9 Analysis of Stool Examination

10. OCCULT BLOOD : Small amount of blood in stool cannot be seen as gross examination Chemical test is necessary to detect occult blood in stool Presence of blood/hemoglobin in the stool which is detected by a ‘chemical test’ and not by the naked eye is known as ‘occult(hidden) blood’ TEST FOR OCCULT BLOOD Benzidine test: sensitive test Principle : Heme compounds derived from hemoglobin molecule in red cells(from blood in the stool) have peroxidase activity. This peroxidase converts hydrogen peroxide(present in the reagent) to nascent oxygen 10/29/2021 10 Analysis of Stool Examination

11. The released oxygen oxidizes the benzidine (in the reagent) in an acidic pH to colored oxidation products which are blue or green in color PROCEDURE The benzidine reagent consists of 4g benzidine base in 10ml glacial acetic acid Stable for 2-4 months Emulsify a bit of stool in 5ml water Mix 1ml of emulsion with 1ml of benzidine reagent in a test tube Add several drops of 3% hydrogen peroxide Interpretation - Appearance of blue color indicates the presence of blood Precaution - Benzidine is carcinogenic Other test - Guaiacum test using gumguaiacum 10/29/2021 11 Analysis of Stool Examination

12. Causes of false –positive reactions: Presence of substances like myoglobin and hemoglobin (Presence in red meat) in diet Presence of vegetable peroxidases ( eg.Horse radish, bananas, black grapes,pears , plums, melons) Leukocytes and bacteria Drugs like boric acid, bromides, iodine and oxidizing agents. Cause of false –negative reactions: Use of vitamin C and other oxidants SIGNIFICANCE Determining the cause of microcytic hypochromic anemia due to chronic blood loss Cause of chronic blood loss may be: GIT neoplasms ( eg . colon, stomach cancer) 10/29/2021 12 Analysis of Stool Examination

13. Ulcerative diseases of the GIT Hookworm infection Drugs like aspirin, steroids and indomethacin can be associated with increased gastrointestinal bleeding OTHER CHEMICAL TESTS Quantitative fecal fat estimation: Increase in fecal fat (more than 6g per day) is found in malabsorption syndromes or diseases of pancreas. Presence of reducing substances like lactose in stool may be found in infants with diarrhea 10/29/2021 13 Analysis of Stool Examination

14. 10/29/2021 Analysis of Stool Examination 14 Characteristics Amoebic dysentery Bacillary dysentery Nature of stool Blood and mucous mixed with fecal matter Blood and mucous without fecal matter Consistency Does not adhere to the container Adheres to the container Odor Offensive Odorless Reaction Acidic Alkaline Microscopy RBC’s Pus cells Macrophages Parasites Charcot –Leyden crystals In clumps Scanty Scanty Trophozoites Present Discrete Plenty Plenty Nil Absent

15. MICROSCOPIC EXAMINATION A fresh sample of stool without any contamination with disinfectants is used for microscopic examination On microscopy, examine the stool for: Leukocytes(pus cells), red blood cells, muscle fibres , fat globules, crystals, cysts and yeast cells and are expressed as number seen per high power field Protozoa, eggs, larvae and cysts of parasites, flagellates and ciliates They are reported as scanty, few, moderate or many 10/29/2021 15 Analysis of Stool Examination

16. MICROSCOPIC EXAMINATION Methods for the preparation of stool for microscopy Saline preparation A small amount of stool sample is picked up with the help of tooth pick and taken on a glass slide. Mix with normal saline to make a thin emulsion so that the fine print can be seen through it Cover with a cover slip This is useful for the demonstarion of motility especially of Entamoeba histolytica 10/29/2021 16 Analysis of Stool Examination

17. IODINE PREPARATION Gram’s Iodine is used instead of saline Iodine imparts brown color to chromatin granules (nuclei ) of amoebic cysts and also glycogen vacuole But iodine kills living parasites thereby preventing identification of motility 10/29/2021 17 Analysis of Stool Examination

18. STOOL CONCENTRATION When ova and cysts are few in number and are not detected by routine methods, concentration method will be of help These methods are as follows: Floatation method: Stool sample is mixed with either zinc sulphate or magnesium sulfate which has a high specific gravity and causes the parasite to float in the solution Used for concentration of cysts, larvae and most of the helminthes eggs 10/29/2021 18 Analysis of Stool Examination

19. MICROSCOPIC EXAMINATION Sedimentation methods: The parasites sediment and get deposited at the bottom by centrifugation It can be done by either simple sedimentation method or by formal – saline ether sedimentation method Simple sedimentation method: A small amount of stool sample is mixed with saline in a tube or bottle and sleved through a strainer The sieved contents are centrifuged and the supernatant fluid discarded 10/29/2021 19 Analysis of Stool Examination

20. The deposit is resuspended in more saline, mixed and centrifuged This is repeated till the supernatant fluid appears clear The deposit is examined directly on a glass slide The sieved contents are centrifuged and the supernatant fluid discarded The deposit is resuspended in more saline, mixed and centrifuged This is repeated till the supernatant fluid appears clear The deposit is examined directly on a glass slide 10/29/2021 20 Analysis of Stool Examination

21. Formal- saline ether sedimentaion method : This yields a good concentration of parasites and is recommended for routine work But this method cannot be used to concentrate living parasites because the formalin used kills the parasites 10/29/2021 21 Analysis of Stool Examination

22. STOOL CULTURE AND SENSITIVITY Collection - for culture should be collected in a sterile wide mouthed container Culture Media used - Mac conkey’s agar ,nutrient agar or selective media depending on the suspected organism Procedure: Take stool in culture media and incubate at 37C for 18-20 hours Suspected colonies are tested by using oxidase test Causative organisms are identified by using biochemical tests Organism may be confirmed by agglutination using specific antisera Antibiotic sensitivity testing is done for the identified pathogenic organism 10/29/2021 22 Analysis of Stool Examination