Urine Culture Using Calibrated Loop

Principle:

The number of microorganisms per milliliter recovered on urine culture can aid in the differential diagnosis of UTI. Plastic or wire loops, available commercially, have been calibrated to deliver a known volume of liquid when handled correctly, thus enabling the microbiologist to estimate numbers of organisms in the original specimen based of CFU of growth on cultures.

Method:

• Flame the calibrated wire loop and allow it to cool without touching any surface. Alternatively, aseptically remove a plastic calibrated loop from its package.
• Mix the urine thoroughly and remove the top of the container. If the urine is in small- diameter tube, the surface tension will alter the amount of specimen picked up by the loop. A quantitative pipette should be considered it he urine cannot be transferred to a larger container.
• Insert the loop vertically into the urine to allow urine to adhere to the loop.
• Spread the loopful of urine over the surface of the agar plate (see figure below) .A standard quadrant streaking technique is also acceptable.

• Without reflaming, insert the loop vertically into the urine again for transfer of a loopful to a second plate. Repeat for each plate.
• Incubate plates for at least 24 hours at 35⁰C to 37⁰C in air. Colonies are counted on each plate. The number of CFUs is multiplied by 1000 (if a 0.001 mL loop was used) to determine the number of microorganisms per milliliter in the original specimen.
• Because antimicrobial treatment or other factors may inhibit initial growth, reincubate plates with no growth or tiny colonies for an additional 24 hours before discarding plates.
• To store the inoculating loop, place handle down in a test tube taped to the wall, rather than flat on the bench, to prevent bending, which would destroy the calibration.