Exercise #24: Urine Culture Techniques NOTES: Urine Analysis: -Urine ultra filtrate of blood.

-Toxins nutrients for bacteria -Kidneys Ureters Urinary Bladder Urethra Exterior Environment (skin urogenital surface normal flora) -Bacteruria: Bacteria in urine. - Urinary Tract Infection. -Urinary Tract Infection: -3 Routes by which UTI detected: 1. Descending or Hematogenous Route 2. Ascending Route 3. Iatrogenic Route -Organisms Causing UTIs: Normal Flora Pathogenic Escherichia, GMycobacterium, Acid Fast Bacteria Proteus, GSalmonella, GEnterobacter, GBeta-Hemolytic Streptococcus, G+ Serratia, GEnterococci, G+ RESULTS: Urine BEFORE Experiment (Physical Characteristics): A: Cloudy, light yellow, white precipitate B: Light Yellow color, clear C: Light Yellow color, cloudy, no precipitate 1. Blood Agar Plate: A: Beta Hemolysis B: None C: Gamma Hemolysis Analysis: A: Most likely infected urine sample B: Most likely normal urine sample C: Possible infected urine sample 2. MacConkey Agar Plate A: No growth B: No growth C: Growth of bacteria; Yellow color from fermentation of acid. Analysis: A: No growth, must be Gram positive bacteria since bacteria was killed B: No growth, must be Gram positive bacteria since bacteria was killed C: Growth of bacteria, so it must be Gram-negative bacteria.

3. pH Paper A: Green-weak alkaline 8.0 B: Yellow- weak acid 6.0 C: Yellow- weak acid 6.0 CONCLUSIONS: A: Infected Urine sample, Gram-positive Bacteria. This urine is infected because it showed beta hemolysis on the blood agar plate, and its physical characteristics showed it was cloudy with a white precipitate, which is abnormal. We know the bacteria is Gram-positive because it was killed on the MacConkey agar plate. B: Normal Urine sample, There was no hemolysis present, no growth on MacConkey agar plate, and the physical characteristics seemed normal since it was a light yellow and clear color. C: Infected Urine sample, Gram-negative bacteria. This urine is infected because it showed growth on the MacConkey agar plate and was fermented to a yellow color meaning it is acidic. Since there was bacterial growth on the MacConkey agar plate we know it must be Gram-negative bacteria. Questions: 1. What is a bacteriuria? When is it significant? The presence of bacteria in voided urine, when urine is voided it becomes contaminated by the normal flora of the urethra and other superficial urogenital mucous membranes is bacteriuria. The presence of bacteria in voided urine does not always indicate urinary tract infections, to confirm infection, either the numbers of organisms present or the species isolated must be shown to be significant. 2. How do microorganisms enter the urinary tract? Active infection of the urinary tract develops in one of three ways. -Microorganisms circulating in the bloodstream from another site of infection are deposited and multiply in the kidneys to produce pyelonephritis by thehematogenous route. -Bacteria colonizing the terminal third of the urethra or the external urogenital surfaces ascend the urethra to the bladder causing cystitis or pyelonephritis by the ascending route. -Microorganisms usually from the urethra are introduced into the bladder on catheters or cystoscopes. 3. List five bacteria that can cause urinary tract infection. -Escherichia coli -Klebsiella -Enterobacter -Serratia -Proteus

4. How would you instruct a female patient to collect her own urine specimen by the clean-catch technique? A male patient? -Female: All labial surfaces must be thoroughly cleansed, and the sterile container must be hold so it does not come in contact with the skin or clothing. The first stream of urine is discarded and a midstream sample is collected. When the container has been tightly closed, it is wiped clean with disinfectant. Always wash your hands and women wipe between the fold of their genitals from front to back. -Male: The procedure entails thorough washing of the plans of the penis; discard the first stream of urine, and collection of a midstream portion in a sterile container. 5. Describe a urine transport system that allows the specimen to remain at room temperature for short time periods without refrigeration. A transport system that allows the specimen to remain at room temperature would be by catheterization. To obtain a catheterized urine specimen, a sterile, polyurethane catheter tube is inserted into the urethra and passed up into the bladder. The urine drains through the catheter tube and is collected in a sterile specimen cup. If it is obtained correctly, catheterized urine is not contaminated by normal urogenital flora and represents urine obtained directly from the bladder.