Interrupting the transmission of microorganisms  

STUDENT HANDOUT:

• Interrupting the transmission of microorganisms Student Handout

LABORATORY REPORT FORM:

• Interrupting the transmission of microorganisms Laboratory Report Form

INTRODUCTION:

Individuals play a crucial role in limiting the spread of infectious diseases. In fact, a single person can, through unknowing, careless, or willful intent, cause devastating illness and even death in others.  

A classic example of the transmission of a bacterial infection is the story of Typhoid Mary.  Mary Mallon was a cook who lived in NYC in the late 1800s and early 1900s.  In 1906, Mary worked for a wealthy banker who rented a home in Long Island for the summer.  In late August to early September, 6 of 11 people in the home became infected with typhoid fever, a potentially deadly gastrointestinal disease caused by the bacterium, Salmonella typhi. Initially, no one knew the source of the S. typhi bacteria that was causing the infection.  

The person who investigated the outbreak was George Sober, a “sanitary engineer”.  Initially he thought the source of the bacteria might be the clams served at the home, but since not everyone who was sick had eaten the clams, he began to suspect the cook, Mary Mallon, who had a mild form of the disease, but was continuing to cook and serve food!  Mary was described as a “healthy carrier”, someone who harbors disease-causing bacteria in their body but is not sick.  George Sober was able to piece together the fact that Mary had worked for 8 families in the past.  There were 22 Salmonella infections in 7 of these 8 families and some of the infected people died.  Amazingly, that same year, there were around 3,000 Salmonella infections in NYC and Mary is thought to be the main cause of the outbreak!  

Since there were no effective vaccines or treatments for S. typhi at the time; the solution was to limit Mary’s contact with other people.  In late 1907, the police and the health department pursued Mary, who eluded them for 5 hours, and finally brought her to a hospital.  She was confined to a cottage on the hospital grounds, forced to stay there for 2 years.  It is unclear if anyone adequately explained the concept of a carrier to Mary. 

In 1910, a new health commissioner allowed Mary to be released to work in a non-food related position.  However, Mary obtained a position as a cook at Sloane Maternity in Manhattan, where she infected at least 25 people, 2 of which died.  

After that, she was given the name “Typhoid Mary” in cartoons and newspapers (Figure 1), and even in medical journals.  After the hospital outbreak, Mary was placed back in Riverside Hospital where she was forced to live for the rest of her life.  She died in 1932.

Figure 1. Newspaper article about Typhoid Mary. Image from Wikimedia Commons. 

Microorganisms are present on the entire surface of the human body and its mucosal surfaces. In addition, microbes are present in the environment-in our food and water, in the soil, and in and on the surfaces of animals and plants. Spread of microorganisms between people may occur through close contact (e.g., coughing, sneezing, and handshaking). Spread from the environment to humans may occur through touching of contaminated materials, ingestion of contaminated food or drink, or inhalation of contaminated air.

To stop or minimize the transmission of communicable diseases, it is necessary to interrupt the chain of their spread by eliminating the source of microorganisms or at least greatly reducing their numbers. This can be done through disinfection of environmental surfaces using disinfectants and handwashing with specialized chemicals called antiseptics.

PRECAUTIONS:

1. Wear a disposable apron.

2. Wash your hands with an antimicrobial soap such as liquid Dial® every time you enter or leave the lab.

3. Wipe tabletops thoroughly with Lysol® or Clorox Wipes.

4. Be extremely careful when using the Bunsen burner.

5. Once the inoculating loop is sterilized, do not touch it to the table, hands, etc.

6. Avoid contact of bleach with your skin, eyes, and clothing.

7. Discard all materials in biohazard containers for proper disposal.

8. Autoclave ALL materials as soon as you no longer need them.

9. NEVER discard cultures in the trash can.

BEFORE BEGINNING, YOUR TEACHER SHOULD DIVIDE YOUR CLASS INTO 4 GROUPS (A, B, C,  & D).

You will then pair up (A & B are one group, C & D are another group).

EXPERIMENT #1:  How microorganisms are transmitted (A & B)

OBJECTIVES:

1. Demonstrate how bacteria on environmental objects can be transferred to a person and possibly to a group of persons.

2. Demonstrate how bacteria can be transmitted or transferred directly from one person to another, and possibly to a group of persons.

3. Quantify the number of bacterial cells on surfaces.

MATERIALS:

1. Ground meat, or a house plant with very moist soil

2. Wooden cutting board

3. Antimicrobial soap

4. Permanent marker

5. Dishwashing soap

6. 4 Trypticase Soy agar (TSA) plates for fingertip colony counts

7. 37°C incubator

8. Biohazard bag

9. Masking tape

PROCEDURE:

Student A

1. Label two TSA plate with your initials and the date.  

• Label one "hand before use”

• Label one "hand after use"

1. Take a culture sample of the fingertips of your right hand as follows:

(a)  Remove the top lid of the petri dish labeled "hand before use."

(b)  Gently press the tips of your five fingers on the surface of the agar for about 15 seconds so that you leave an impression of each tip. Be sure that your fingertips completely touch the agar surface.

(c)  Close the lid and tape it to the base using two small pieces of masking tape.

3. Using both hands, roll food (or soil) on the cutting board. Be sure your fingertips make contact with the food! Don't allow your hands to touch any other object!

4. Take a culture sample of the fingertips of your right hand using the procedure followed in step 2 using the "hand after use" plate.

5. Again, roll meat (or plant soil) on the cutting board.

6. Using right hands, shake hands with student B after student B completes step 2 in the "Student B" section. Be sure to grip tightly! Don't allow your hands to touch any other object!

7. Dispose of meat or soil.

8. Clean the cutting board with antibacterial soap and warm water.

9. Wash your hands well with the antibacterial soap.

Student B

1. Label two TSA plate with your initials and the date.  

• Label one "hand before use”

• Label one "hand after use"

2. Take a culture sample of the fingertips of your right hand as follows:

(a)  Remove the top lid of the petri dish.

(b)  Gently press the tips of your five fingers on the surface of the agar for about 15 seconds so that you leave an impression of each tip. Be sure that your fingertips completely touch the agar surface.

(c)  Close the lid and tape it to the base using two small pieces of masking tape.

3. Shake right hands with student A, after student A completes step 5.

4. Following the procedure described in step 2, take a culture sample of the fingertips of your right hand used in the handshake with student A using the "hand after use" plate.

5. Wash your hands well with the antimicrobial soap.

Students A & B

1. Tape plates together with two pieces of masking tape. Incubate all plates in an upside-down position in a 37°C incubator for one or two days OR incubate on a window sill at room temperature for at least two days or until bacterial colonies are visible. At the end of the lab, dispose of plates in biohazard bag.

OBSERVATIONS/CONCLUSIONS:

1. Examine the culture plates.

2. Count the number of bacterial colonies on each plate and record your results on Table 1 of the Laboratory Report Form.

3. Compare the “before” and “after” plates.

4. Is there a difference in the number of colonies on the “before” and the “after” plates?  Summarize your conclusions.

QUESTIONS:

1. How are bacterial colonies detected and counted?

2. Did all the bacterial colonies look alike? If not, what could account for differences in appearance?

3. Which cultures, "before" or "after," gave rise to the largest bacterial counts?

4. Did the handshake with student A, who contacted food or soil, spread microbes to student B?

5. If student A, who contacted food or soil, had shaken the hands of students X, Y, and Z (assuming that they have clean, washed hands), and you had taken culture samples of students X, Y and Z's hands after the handshakes, would the bacterial counts of their cultures increase or decrease after the handshakes?

EXPERIMENT #2:  Transmission of microorganisms between the environment and humans (C& D)

OBJECTIVES:

1. Demonstrate how to interrupt the transmission of microorganisms between the environment and people.

2. Demonstrate how to interrupt the transmission of microorganisms between persons.

3. Quantify the numbers of bacterial cells on inanimate surfaces.

MATERIALS:

1. Ground meat or a house plant with very moist soil

2. Wooden cutting board

3. Antimicrobial soap

4. Household bleach

5. Permanent markers

6. Paper towels

7. 3 Trypticase Soy agar (TSA) plates for fingertip colony counts

8. 3 RODAC® or other TSA plates with colony counting grids for sampling of cutting board

• There are special agar plates used for culturing surfaces.  These plates are made so that the surface of the growth medium (nutrient agar) is raised over the edge of the dish.  These plates are known as RODAC® Plates.  Information about the RODAC® plate can be found in the Laboratory Media Appendix.

9. Masking tape

10. Dishwashing soap

11. 37°C incubator

12. Biohazard bag

PROCEDURE:

Student C

1. Label three RODAC® plates with your initials and the date.

• Label one "board before use,”

• Label one "board after use"

• Label one "board after wash"

2. Take a culture sample of a clean cutting board as follows:

(a)  Remove the lid of the RODAC® petri dish.

(b)  Take the bottom lid, containing the mound of culture medium, and roll it on the surface of the board (Figure 2).

(c)  Make sure that the entire surface of the agar firmly contacts the sample surface by gently pressing in the bottom of the dish with the knuckles of your second or third finger.

(d)  Close the lid and tape it to the base using two small pieces of masking tape.

Figure 2.  Working with RODAC plates.

3. Using both hands, roll meat (or soil) on the cutting board. Be sure to make complete contact between the food and the cutting board!

4. Gently remove the largest amounts of food/soil from the board and dispose of them in biohazard bags.

5. Thoroughly wash your hands with antimicrobial soap and warm water.

6. Following the procedure described in step 2, take a sample of the cutting board using the plate labeled "board after use."

7. Thoroughly wash the cutting board with dishwashing soap and warm water.

8. Rinse well. Carefully pour bleach on the board and let it stand for 15 minutes. Take care not to splash bleach on clothes or in eyes!

9. Wipe the board dry with paper towels.

10. Following the procedure described in step 2, take a sample of the cutting board using the plate labeled "board after wash."

Student D

1. Label three TSA plate with your initials and the date.  

• Label one "hand before use,”

• Label one "hand after use"

• Label one "hand after wash"

2. Take a culture sample of the fingertips of your right hand as follows:

(a)  Remove the top lid of the petri dish labeled "hand before use".

(b)  Gently press the tips of your five fingers on the surface of the agar for about 15 seconds so that you leave an impression of each tip. Be sure that your fingertips completely touch the agar surface.

(c)  Close the lid and tape it to the base using two small pieces of masking tape.

3.  Using both hands, roll the food (or soil) on the cutting board. Be sure to contact your fingertips with the food! Don't allow your hands to touch any other object!

4.  Take a culture sample of the fingertips of your right hand using the procedure followed in step 2 using the "hand after use" plate.

5.  Thoroughly wash your hands with antimicrobial soap and warm water.

6.  Completely dry your hands with paper towels.

7.  Take a culture sample of the fingertips of one of your washed hands using the procedure followed in step 2 using the "hand after wash" plate.

Students C & D

1. Tape plates together with two pieces of masking tape. Incubate all plates in an upside-down position in a 37°C incubator for one or two days OR incubate on a window sill at room temperature for at least two days or until bacterial colonies are visible. At the end of the lab, dispose of plates in biohazard bag.

OBSERVATIONS/CONCLUSIONS:

1. Examine the culture plates.

2. Count the number of bacterial colonies on each plate and record your results on Table 2 of the Laboratory Report Form.

3. Compare the “before use,” “after use,” and “after wash” plates.

4. Is there a difference in the number of colonies on the plates? Summarize your conclusions.

KEY TERMS:

• antiseptic 

• contaminated material

• disinfectant 

• disinfection

• mucosal surface

SUGGESTED READINGS:

Marineli, F., G. Tsoucalas, M. Karamanou and G. Androutsos. Mary Mallon (1869-1938) and the history of typhoid fever. Annals of Gastroenterology (2013) 26, 132-134