FT EXP 1

FT 100 Food Microbiology and Toxicology: Experiment 1 - Cleaning and Sanitation Assessment: Swab Contact Method

Objectives

  • Assess the microbial load of surfaces and the human hand.

  • Determine the effectiveness of cleaning agents and the thoroughness of cleaning procedures for sanitizing surfaces and hands.

Materials

  • Glassware

  • Sterile cotton swab

  • Test tubes (10ml)

  • Sterile template

  • Pipettes (5ml)

  • Detergent

  • Petri dish

  • Hand soap

  • Erlenmeyer flask (250ml)

  • Alcohol

  • Igniter

  • Alcohol lamp

  • Peptone

  • Test tube rack

  • Plate count agar

  • Aspirator

  • School of Technology, University of the Philippines Visayas, Miagao, Iloilo, Philippines

Procedures

A. Swabbing Method for Surfaces

  1. Identify the surface you wish to assess.

  2. Position the sterile template on the unclean surface to acquire a sample.

  3. Carefully open the aluminum packet containing the sterile cotton swabs. Remove one swab from the stick end and close the packet aseptically to maintain sterility.

  4. Moisten the cotton swab end in 10ml of peptone dilution blank. This can be done by pressing and rotating the cotton bud against the wall of the test tube.

  5. Rub the moistened swab on the chosen surface through the opening in the template. Make sure to collect samples from three different points on the surface.

  6. Return the cotton swab to the test tube and break off the wooden stick, leaving about 1 inch of the stick with the swab. Ensure the cap of the test tube is replaced securely.

  7. Homogenize the sample using a vortex mixer to prepare a 10^{-1} dilution.

  8. Transfer 1 ml from the first dilution blank to a 9 ml peptone dilution blank to obtain a 10^{-2} dilution. Prepare subsequent dilutions up to 10^{-5}.

  9. Conduct pour-plating for each dilution in triplicate to ensure accurate results.

  10. Repeat steps 1-8 after cleaning and sanitizing the same surface to assess effectiveness.

  11. Incubate plates at 35°C for 24 hours to allow growth of microbial colonies.

  12. Count the colonies that develop on the plates and express the count as CFU (Colony Forming Units) per square inch to quantify the microbial load.

B. Swabbing Method for Hands

  1. Open the aluminum packet with sterile cotton swabs carefully. Remove one swab from the stick end and securely close the packet to maintain sterility.

  2. Moisten the swab end in 10ml of peptone dilution blank, removing unnecessary excess by pressing and rotating the cotton against the wall of the test tube.

  3. Rub the moistened swab on the palm of the hand, the fingertips, and the area where the fingers meet the palm to ensure comprehensive sampling.

  4. Follow steps 5 to 10 of Method A, which involves conducting pour-plating for dilution, incubating, and counting to assess microbial presence on hands.


FT 100 Food Microbiology and Toxicology: Experiment 1 - Cleaning and Sanitation Assessment: Swab Contact Method

To master and focus on both the performance and purpose of this experiment, you should concentrate on the following key concepts:

Purpose of the Experiment (Objectives)

These are the main goals and learning outcomes you should understand:

  • Assessment of Microbial Load: Understanding how to quantify the presence of microorganisms on surfaces and hands.

  • Effectiveness of Cleaning Agents: Determining how well different cleaning and sanitizing solutions reduce microbial contamination.

  • Thoroughness of Cleaning Procedures: Evaluating the efficacy of the physical cleaning methods employed in reducing microbial populations on surfaces and hands.

Performing the Experiment (Procedures)

These are the critical skills and steps you need to master during the practical execution:

  1. Aseptic Technique:

    • Maintaining sterility when handling cotton swabs, test tubes, and other materials (e.g., careful opening and closing of sterile packets, preventing contamination).

    • This is crucial to ensure that the microbial load measured truly represents the sample and not environmental contamination.

  2. Swabbing Method:

    • For Surfaces: Accurately positioning the template and rubbing the moistened swab across three different points within the designated area to collect a representative sample.

    • For Hands: Systematically rubbing the swab on the palm, fingertips, and the area where fingers meet the palm for comprehensive sampling.

  3. Sample Preparation and Dilution:

    • Properly moistening the swab in peptone dilution blank and breaking off the wooden stick while maintaining sterility.

    • Homogenization: Using a vortex mixer to ensure the sample is evenly dispersed.

    • Serial Dilution: Accurately preparing a series of dilutions (up to 10−510^{-5}) by transferring 1 ml from one dilution blank to 9 ml of peptone dilution blank. This is essential for obtaining countable plates.

  4. Pour-Plating Technique:

    • Understanding how to aseptically transfer diluted samples into Petri dishes and mix with molten plate count agar.

    • Performing pour-plating in triplicate for each dilution to ensure accuracy and statistical reliability.

  5. Incubation:

    • Ensuring plates are incubated at the correct temperature (35∘C35^{\circ}\text{C}) for the specified duration (24 hours) to allow for optimal microbial growth.

  6. Colony Counting and Quantification:

    • Accurately counting the Colony Forming Units (CFUs) that develop on the plates.

    • Expressing the microbial load correctly as CFU per square inch (CFU/in2\text{CFU/in}^2) for surfaces.