FECAL ANALYSIS ROUTINE FECAL EXAMINATION macroscopic, microscopic and chemical analysis of feces for the early detection

n of: o gastrointestinal (GI) bleeding o causes of diarrhea and steatorrhea o liver and biliary duct disorders o detection of pathogenic bacteria and o maldigestion/malabsorption syndromes parasites o inflammation

PHYSIOLOGY Normal fecal specimen contains: o bacteria (many of which make up the normal flora of the intestines o cellulose o other undigested foodstuffs o GI secretions BACTERIAL METABOLISM produces the strong odor associated with feces and intestinal gas (flatus)

o o o o

bile pigments cells from the intestinal walls electrolytes water

CARBOHYDRATES especially oligosaccharides resistant to digestion pass through the upper intestine unchanged but are metabolized by bacteria in the lower intestine, producing large amounts of flatus excessive gas production: LACTOSE-INTOLERANT INDIVIDUALS when the intestinal bacteria metabolize the lactose from consumed milk or lactose-containing substances

ALIMENTARY TRACT where digestion of ingested proteins, carbohydrates and fats take place SMALL INTESTINE primary site for the final breakdown and reabsorption of these compounds

DIGESTIVE ENZYMES SECRETED INTO THE SMALL INTESTINE BY THE PANCREAS trypsin chymotrypsin amino peptidase lipase

BILE SALTS provided by the liver aid in the digestion of fats

* A deficiency in any of these substances causes the inability to digest, and, therefore, to reabsorb certain foods

EXCESS UNDIGESTED OR REABSORBED MATERIAL appear in feces patient exhibits symptoms of maldigestion and malabsorption INGESTED FLUID, SALIVA, GASTRIC, LIVER, PANCREATIC AND INTESTINAL SECRETIONS 9000 mL/day DIGESTIVE TRACT 500 to 1500 mL LARGE INTESTINE 150 mL FECES WATER AND ELECTROLYTES readily absorbed in both intestines fecal electrolyte content similar to plasma

LARGE INTESTINE can absorb approximately 3000 mL of water

DIARRHEA occurs when amount of water reaching the large intestine exceeds 3000 mL water is excreted with the solid fecal material

CONSTIPATION provides time for additional water to be reabsorbed from the fecal material small, hard stools DIARRHEA DIARRHEA increase in daily stool weight above 200 g increased liquidity frequency: more than 3X/day can be classified based on: a. Duration of the illness o ACUTE: less than 4 weeks o b. CHRONIC: more than 4 weeks Mechanism Osmotic gap = 290 [2 (fecal Na+ + fecal K+)

Osmotic diarrhea = >50 mOsm/kg, negligible electrolytes Secretory diarrhea = <50 mOsm/kg, increased electrolytes

MECHANISM

ACTION

CAUSATIVE AGENTS

OTHER CAUSES Drugs Stimulant laxatives Hormones Inflammatory bowel disease o Crohn disease o Ulcerative colitis o Lymphocytic colitis o Diventiculitis Endocrine disorders o Hyperthyroidism o Zollinger-Ellison syndrome o Vipoma Neoplasms Collagen vascular disease

Secretory

Increase in secretion of water and electrolytes which override the reabsorptive ability of the large intestines

E. coli Clostridium V. cholerae Salmonella Shigella Staphylococcus Campylobacter Protozoa Cryptosporidium

Incomplete breakdown or reabsorption of food presents increased fecal material to the large intestine, resulting in the retention of water and electrolytes in the large intestine Conditions of enhanced motility (hypermotility) or slow motility (constipation)

Osmotic

Entamoeba histolytica

Disaccharidase deficiency (lactose intolerance) Malabsorption (Celiac sprue) Poorly absorbed sugars (lactose, sorbitol, mannitol) Laxatives Magnesium-containing antacids Antibiotic administration

Altered Motility

Vagotomy Diabetic neuropathy Complication of menstruation Hyperthyroidism

IRRITABLE BOWEL SYNDROME (IBS) both hypermotility and constipation are seen a functional disorder in which the nerves and muscles of the bowel are extra-sensitive, causing: o cramping o diarrhea o bloating o constipation o flatus triggered by: o food o chemicals o emotional stress o exercise RAPID (ACCELERATED) GASTRIC EMPTYING (RGE) DUMPING SYNDROME hypermotility of the stomach shortened gastric emptying half-time, causing the small intestines to fill too quickly with undigested food from the stomach hallmark of Early Dumping Syndrome (EDS) healthy individuals: gastric emptying half-time range of 35-100 minutes (varies with age and gender) RGE: less than 35 minutes caused by disturbances in the gastric reservoir or in the transporting function normal gastric emptying is controlled by the FUNDIC TONE, DUODENAL FEEDBACK and GI HORMONES

START

Early Dumping

10-30 minutes following meal ingestion

Late Dumping

2-3 hours after a meal

SYMPTOMS Nausea Vomiting Bloating Cramping Diarrhea Diziness Fatigue Weakness Sweating Dizziness

COMPLICATION Hypoglycemia

CAUSES Gastrectomy Gastric bypass surgery Postvagotomy status Zollinger-Ellison syndrome Duodenal ulcer disease Diabetes mellitus

STEATORRHEA increase in stool fat that exceeds 6 g/day due to absence of bile salts that assist pancreatic lipase in the breakdown and subsequent reabsorption of triglycerides detection is useful for the diagnosis of pancreatic insufficiency and small boweled disorders that cause malabsorption disease association: o Cystic fibrosis o Chronic pancreatitis o Carcinoma Steatorrhea may be present Maldigestion and Malabsorption 9000 mL/day D-Xylose Test urine D-Xylose is low Malabsorption D-XYLOSE a sugar that does not need to be digested but does need to be absorbed to be present in urine if low: Malabsorption if normal: Pancreatitis MALABSORPTION CAUSES o o o o o c. d. Bacterial overgrowth Intestinal resection Celiac disease Tropical sprue Lymphoma o o o o Whipple disease Giardia lamblia infestation Crohn disease Intestinal ischemia

Severity Stool characteristics

SPECIMEN COLLECTION 1. 2. Collect specimen in a clean container, such as a bedpan or disposable container. Transfer specimen to laboratory container.

Precaution: SPECIMEN MUST NOT BE CONTAMINATED WITH URINE OR TOILET WATER, which may contain disinfectants. KITS FOR OCCULT BLOOD contain paper that can be floated in toilet bowl to collect the specimen

RANDOM SPECIMENS for qualitative testing for blood microscopic examination for leukocytes, muscle fibers and fecal fats container: PLASTIC OR GLASS CONTAINERS WITH SCREW-CAPPED TOPS

TIMED SPECIMENS for quantitative testing for fecal fats most representative sample: 3-DAY COLLECTION o due to variability of bowel habits and transit time required for food to pass through the digestive tract container: PAINT CANS to accommodate specimen quantity and facilitate emulsification prior to testing

MACROSCOPIC SCREENING first indication of GI disturbances: o changes in brown color o formed consistency

COLOR BROWN PALE YELLOW, WHITE, GRAY BLACK, TARRY Esopaghus bleeding Stomach bleeding Duodenal bleeding o all of these take 3 days to appear in stool Iron ingestion Charcoal ingestion Bismuth ingestion (antacids) RED GREEN Oral antibiotics (bilirubin to biliverdin) Ingestion of increased amounts of green vegetables Food coloring

Normal: Stercobilinogen to Urobilin

Blockage of the bile duct Diagnostic procedures that use barium sulfate

Lower GI bleeding Medications Food, especially Beets

APPEARANCE SLENDER, RIBBONLIKE STOOLS

WATERY CONSISTENCY

SMALL, HARD STOOLS

BULKY, FROTHY, FOUL ODOR, GREASY, MAY FLOAT

MUCUS-COATED STOOLS

BLOOD-STREAKED MUCUS-streaked

Diarrhea

Constipation

Intestinal constriction

Biliary obstruction Steatorrhea Pancreatic disorders

Intestinal inflammation or irritation

Bacterial or amebic dysentery

MICROSCOPIC EXAMINATION OF FECES FECAL LEUKOCYTES primarily neutrophils seen in infections that affect intestinal mucosa such as ULCERATIVE COLITIS and BACTERIAL DYSENTERY PRELIMINARY TEST to determine causative agent INVASIVE BACTERIAL PATHOGENS CAUSE THE APPEARANCE OF FECAL LEUKOCYTES Salmonella Shigella Campylobacter Yersinia Enteroinvasive E. coli STAIN Methylene blue Dried smear Wrights Grams TOXIN-PRODUCING BACTERIA USUALLY DO NOT CAUSE THE APPEARANCE OF FECAL LEUKOCYTES Staphylococcus aureus Vibrio spp.

Wet preparation

ADVANTAGES Faster Provide permanent slides Observation of Gram (+) and Gram (-) bacteria

DISADVANTAGES More difficult to interpret

All slide preparations: FRESH SPECIMEN 3 neutrophils/hpf: INVASIVE CONDITION OIO: finding of neutrophils has approximately 70% sensitivity for the presence of invasive bacteria Lactoferrin Latex Agglutination Test o detects fecal leukocytes and remains sensitive in refrigerated and frozen specimens o LACTOFERRIN: granulocyte secondary granule

MUSCLE FIBERS Indicative of pancreatic insufficiency, such as CYSTIC FIBROSIS, BILIARY OBSTRUCTION and GASTROCOLIC FISTULAS emulsify a small amount of stool in 10% alcoholic eosin, which enhances the muscle fiber striations slide is examined for 5 minutes number of RED-STAINED FIBERS WITH WELL-PRESERVED STRIATIONS are counted UNDIGESTED FIBERS: visible striations running both vertically and horizontally; ONLY ONES COUNTED PARTIALLY DIGESTED FIBERS: striations in only one direction presence of MORE THAN 10 UNDIGESTED FIBERS are considered increased representative sample: RED MEAT IN DIET PRIOR TO COLLECTION; examined within 24 hours

QUALITATIVE FECAL FATS specimens suspected of steatorrhea: MICROSCOPIC EXAMINATION FOR EXCESS FECAL FAT o monitoring patients undergoing treatment for malabsorption disorders stains: Sudan III (routine), Sudan IV, Oil Red O NEUTRAL FATS (TRIGLYCERIDES) o stained by Sudan III as large orange-red droplets often located near the edge of cover slip o >60 droplets/hpf: steatorrhea o SPLIT FAT STAINING total fat content; breakdown of neutral fats by bacterial lipase and hydrolysis of neutral fats may lower neutral fat count determine whether maldigestion or malabsorption causes steatorrhea SOAPS and FATTY ACIDS o do not stain with Sudan III o second slide mixed with acetic acid and heated stained droplets: free fatty acids, fatty acids from soap and neutral fat hydrolysis o Normal: 100 small droplets, <4m/hpf

o Slightly elevated: 100 small droplets, 1-8m o Increased: 100 droplets measuring 6-75 m CHOLESTEROL o stained by Sudan III after heating o forms crystals upon cooling

CHEMICAL TESTING OF FECES OCCULT BLOOD Fecal Occult Blood Test (FOBT) detection of hidden blood most frequently performed fecal analysis bleeding >2.5 mL/150 g of stool: pathologically significant but may not produce signs of bleeding early detection of colorectal cancer principle: PSEUDOPEROXIDASE ACTIVITY OF HEMOGLOBIN chromogens in order of decreasing sensitivity: o benzidine o ortho-tolidine o gum guaiac: routine commercial testing kits o Guaiac-impregnated filter paper: feces + H2O2 Contraindications: 3 days: o red meat o horseradish o melon o raw broccoli o cauliflower o radish o turnip o Vitamin C o Iron supplements 7 days o Aspirin o NSAIDs other than Acetaminophen Hemoquant o fluorometric test for hemoglobin and porphyrin Immunochemical Fecal Occult Blood Test (iFOBT) o specific for the globin portion of human hemoglobin and uses anti-human hemoglobin Abs

QUANTITATIVE FECAL FAT TESTING confirmatory test for steatorrhea Van de Kamer titration (gold standard) 3-day specimen with a regulated intake of fat (100 g/d) fecal lipids are converted to fatty acids and titrated to a neutral endpoint with NaOH Coefficient of fat retention = (dietary fat fecal fat) X 100 dietary fat ACID STEATOCRIT o rapid test to estimate amount of fat excretion o monitor therapy and screen for steatorrhea in pediatric populations NEAR-INFRARED REFLECTANCE SPECTROSCOPY (NIRA) o 48-72hr stool that does not require reagents after homogenization o reflectance of fecal surface + IR light between 1400 nM and 2600 nM o quantitates water, fat and nitrogen in g/24h

APT TEST (FETAL HEMOGLOBIN) grossly bloody stools and vomitus: SWALLOWING MATERNAL BLOOD DURING DELIVERY material is emulsified in water to release Hb, centrifuged, + 1% NaOH to pink Hb-containing supernatant in the presence of alkali-resistant fetal Hb, the solution remains pink (Hb F), whereas denaturation of the maternal Hb (Hb A) produces a yellow-brown supernatant after standing for 2 minutes the test can also distinguish fetal Hb from Hb A, Hb AS, CS and SS stool specimens should be tested when fresh

FECAL ENZYMES supplied by the pancreas for digestion of dietary proteins, carbohydrates and fats decrease (pancreatic insufficiency) is associated with chronic pancreatitis and cystic fibrosis steatorrhea occurs, presence of undigested food in feces analysis focuses on TRYPSIN, CHYMOTRYPSIN and ELASTASE I Trypsin: (historically) absence has been screened for by exposing x-ray paper to stool emulsified in water o If present: it digests gelatin on the paper o Detects only severe cases of pancreatic insufficiency o False-negative results: intestinal degradation of trypsin and possibly trypsin inhibitors in feces, bacterial enzymes Chymotrypsin: more resistant to intestinal degradation, more sensitive indicator of less severe cases of pancreatic insufficiency o remains stable in feces for 10 days at RT o also capable of gelatin hydrolysis but most frequently measured spectrophotometrically Elastase I: isoenzyme of elastase; enzyme form produced by pancreas o strongly resistant to degradation o 6% of secreted pancreated enzymes o pancreas-specific with concentrations 5X higher than in pancreatic juice o not affected by motility disorders and mucosal defects o measured using ELISA that provides a very sensitive indicator of exocrine pancreatic insufficiency

CARBOHYDRATES an increase indicates OSMOTIC DIARRHEA by the osmotic pressure of unabsorbed sugar in intestine drawing in fluid and electrolytes may be present due to: o intestinal inability to absorb carbohydrates: CELIAC DISEASE o lack of digestive enzymes such as lactase: LACTOSE INTOLERANCE Carbohydrate malabsorption or intolerance (maldigestion): primarily analyzed by serum and urine tests o COPPER REDUCTION TEST (Clinitest tablet) in fecal specimen Detects congenital disaccharidase deficiencies as well as enzyme deficiencies due to nonspecific mucosal injury Infant diarrhea: fecal carbohydrate testing + pH determination o normal pH of feces: 7.0 8.0 o carbohydrate disorders: pH 5.5