Shake ’n’ Incubate (VCE SSCE Biology): Revision Notes

Shake 'n' Incubate

Introduction

Hand hygiene is one of the most effective ways to prevent disease transmission. The surface of our body, particularly our hands, is covered in bacteria. While most of these bacteria are harmless, pathogenic bacteria (disease-causing microorganisms) can sometimes contaminate our hands and enter our bodies when we touch our mouth, eyes, nose, or food.

Studies have demonstrated the remarkable effectiveness of proper handwashing:

• Washing hands with soap and water reduces diarrhoeal diseases by approximately 47%
• Hand hygiene interventions could save up to one million lives that would otherwise be lost to diarrhoea
• Handwashing reduces respiratory infections by approximately 45%

Various products claim to effectively clean hands of bacteria, including water alone, regular soap, antibacterial soap, and hand sanitiser. This investigation uses yeast organisms to model pathogenic bacteria and determine which cleaning agent is most effective.

Aim

To determine the effectiveness of different skin cleaning agents in removing pathogenic microorganisms from hands.

Key concepts

Pathogenic bacteria

Pathogenic bacteria are disease-causing bacteria that can harm the body when they enter through breaks in the skin or mucous membranes. Unlike the harmless bacteria that normally live on our skin, these microorganisms can cause infections and illness.

Cellular pathogens

Cellular pathogens are disease-causing organisms that have a cellular structure, including bacteria and yeast. This distinguishes them from non-cellular pathogens like viruses.

Using yeast as a model organism

In this investigation, yeast is used to represent pathogenic bacteria. Yeast is safer to work with in a school laboratory while still demonstrating the principles of pathogen transmission and removal. When grown on nutrient agar plates, yeast forms visible colonies that can be counted and compared.

Materials

• 12 × poured sterile nutrient agar plates
• 1 × packet of yeast
• 1 × 250 mL beaker
• 1 × measuring spoon
• Sugar
• 1 × marking pen
• 1 × sterile pipette
• 12 × sterile cotton swabs
• Gloves
• Sticky tape
• 10 mL regular hand soap
• 10 mL antibacterial hand soap
• 10 mL liquid hand sanitiser
• 1 × incubator set to 37 °C
• 10% bleach solution
• Lab coat and goggles/safety glasses

Method

Part A: Preparation of agar plates

Setting up the investigation

This investigation requires groups of 5 students, with each person assigned a specific role:

• Person 1: The 'disease spreader' (applies the yeast solution)
• Person 2: Tests washing with water only
• Person 3: Tests washing with regular soap
• Person 4: Tests washing with antibacterial soap
• Person 5: Tests washing with liquid hand sanitiser

Preparing the yeast solution

1. Measure 230 mL of warm water into a 250 mL beaker
2. Add one teaspoon of sugar and the entire yeast packet
3. Gently stir the solution to dissolve all components
4. Leave for a few minutes until the solution begins to foam (this indicates the yeast is becoming active)

Labelling the agar plates

Each person washing their hands (Persons 2–5) needs three agar plates labelled as follows:

• Plate 1: 'Pre-exposure' (to test baseline bacterial levels before yeast exposure)
• Plate 2: 'Post-exposure' (to test bacterial levels after exposure to yeast)
• Plate 3: 'Post-wash' (to test bacterial levels after cleaning)

Taking the pre-exposure swab

This baseline measurement is crucial for comparison:

1. Persons 2–5 remove the lid from their 'pre-exposure' plate
2. Using a sterile cotton swab in the left hand, swab the entire surface of the right hand palm and fingers
3. Gently rub the swab across the agar surface without breaking it
4. Replace the lid and seal with sticky tape

Exposing hands to the "pathogen"

For each person (2–5), repeat the following process:

1. Person 1 puts on a glove on their left hand
2. Using a pipette, Person 1 applies 8 drops of yeast solution to the gloved palm
3. Person 1 rubs their hands together to spread the yeast over their right hand
4. Person 1 removes and disposes of the glove
5. Person 1 shakes hands with the participant's right hand for 10 seconds straight (simulating pathogen transmission)

Taking the post-exposure swab

1. Using a fresh sterile cotton swab, swab the participant's right hand
2. Gently rub the swab across the 'post-exposure' agar plate
3. Replace the lid and seal with sticky tape

Washing hands with the assigned method

• Person 2: Wash hands with water only from the tap, scrubbing for 20 seconds, then air dry
• Person 3: Wash hands with normal hand soap and water for 20 seconds, then air dry
• Person 4: Wash hands with antibacterial soap and water for 20 seconds, then air dry
• Person 5: Apply hand sanitiser to hands and rub until dry

Taking the post-wash swab

1. Using a fresh sterile cotton swab, swab the participant's right hand again
2. Gently rub the swab across the 'post-wash' agar plate
3. Replace the lid and seal with sticky tape

Incubation

Place all sealed dishes in an incubator set at 37 °C for 24–36 hours. This temperature mimics human body temperature and allows optimal yeast growth.

Part B: Examination of plates

Recording results

After the incubation period:

1. Carefully examine each plate and observe the level of growth
2. Count the number of colonies on each plate
3. Record results using the standardised notation system:
   • 0 = no growth
   • + = small amount of growth (<5 colonies)
   • ++ = moderate growth (6–20 colonies)
   • +++ = heavy growth (20–50 colonies)
   • ++++ = extremely heavy growth forming a 'lawn' (uncountable individual colonies)

Safe disposal procedure

Results

Record your observations in a table with the following structure:

Treatment	Swab	Amount of growth and number of colonies
Water only	Pre-exposure
	Post-exposure
	Post-wash
Normal soap	Pre-exposure
	Post-exposure
	Post-wash
Antibacterial soap	Pre-exposure
	Post-exposure
	Post-wash
Hand sanitiser	Pre-exposure
	Post-exposure
	Post-wash

Understanding the experimental design

Independent and dependent variables

The independent variable is the type of handwashing agent used (water, normal soap, antibacterial soap, or hand sanitiser). This is what the experimenter deliberately changes.

The dependent variable is the amount of bacterial/yeast growth on the agar plates, measured by colony counts. This is what is measured in response to the independent variable.

Control elements

The 'pre-exposure' swabs serve as a control, showing the baseline level of bacteria on hands before exposure to the yeast "pathogen". This allows for comparison and helps determine if changes are due to the treatment rather than individual variation.

Replication

This experiment includes replication through:

• Multiple participants testing the same condition
• Three separate measurements for each participant (pre-exposure, post-exposure, post-wash)

Discussion questions

Understanding cellular pathogens

What are cellular pathogens?

Cellular pathogens are disease-causing organisms that have a cellular structure, such as bacteria, fungi (including yeast), and protozoa. They differ from non-cellular pathogens like viruses, which lack cellular structure.

What is the faecal-oral route?

The faecal-oral route is a mode of disease transmission where pathogens from faeces are transferred to the mouth, typically through contaminated hands, food, or water. This commonly occurs when people don't wash their hands after using the toilet, then touch food or their mouth.

Disinfectants versus antiseptics

• Disinfectants are chemical agents used to kill microorganisms on non-living surfaces (e.g., bench tops, floors)
• Antiseptics are chemical agents used to kill microorganisms on living tissue (e.g., skin)

Both antibacterial soap and hand sanitiser are antiseptics, as they are designed for use on skin.

Analyzing experimental results

When examining your results, consider:

• Did the post-exposure swabs show more growth than pre-exposure swabs? This would demonstrate pathogen transmission through hand-to-hand contact
• Which treatment showed the greatest reduction in growth from post-exposure to post-wash?
• Were there any treatments that showed little to no difference?

Identifying experimental errors

Types of errors:

Personal errors occur due to human mistakes, such as:

• Breaking the agar surface while swabbing
• Not scrubbing hands for the full 20 seconds
• Touching surfaces after washing hands

Systematic errors consistently affect results in one direction, such as:

• Incubator temperature being incorrectly set
• Using expired yeast that doesn't grow properly

Random errors occur unpredictably and can affect results either way, such as:

• Variations in swabbing technique
• Different amounts of yeast solution transferred during handshaking

Writing your conclusion

Your conclusion should address four key areas:

1. Achievement of aim

State whether the investigation successfully determined the effectiveness of different skin cleaning agents. Reference your specific results (e.g., "Hand sanitiser showed the greatest reduction in colony growth from post-exposure to post-wash")

2. Limitations

Identify weaknesses in the experimental design, such as:

• Small sample size (only one person per treatment)
• Use of yeast instead of actual pathogenic bacteria
• Difficulty ensuring consistent handshaking pressure and duration
• Variability in swabbing technique

3. Improvements

Suggest specific ways to enhance the experiment:

• Increase sample size by having multiple people test each treatment
• Standardise the handshaking procedure
• Use a mechanical device to ensure consistent swabbing
• Test additional variables like water temperature or scrubbing duration

4. Broader implications

Consider the wider significance:

• How these findings relate to public health recommendations
• Applications in healthcare settings
• Further research questions (e.g., testing water temperature, testing different types of sanitiser)

Remember!