Toothbrush Lab HW - lab homework PDF

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BIOL 2440-KJ Fall 2020 Toothbrush Experiment There are many microbes in our environment and as we have discussed in lecture, the majority are not directly harmful to us. Still, you might wonder just how many we have around and how often we come in contact with them. To study this question, we are going to ask the more direct question: Under which storage conditions does my toothbrush acquire the most bacteria in my bathroom? Materials 2 toothbrushes (do not brush with these) 2 Nutrient Agar Plates-Sterile and sealed with Parafilm 4 strips of new Parafilm 4 sterile disposable inoculation loops. 50ml conical tube with disinfectant for your lab space.

Procedure 1. Choose the 2 conditions you would like to test using your set of toothbrushes. You will keep your toothbrush in this condition for 1 full week. a. Toothbrush on counter, Flush the toilet with seat up b. Toothbrush on counter, Flush the toilet with seat down c. Toothbrush in cabinet, Flush the toilet with seat up d. Toothbrush in cabinet, Flush the toilet with seat down 2. If you have roommates or family that share the restroom, let them know of your plans. You may want to leave a note on the bathroom wall or mirror about the required toilet seat condition. Some suggestions for the conditions above are to tape the exposed toothbrush to the wall or put the “cabinet” toothbrush in a shoebox. 3. No need to clean the bathroom, place your toothbrushes in your chosen locations. Each day, for 5-7 days, wet the toothbrush with water from your bathroom sink and return the brush to its designated location. 4. On the last day, inoculate your plates to see what has been collected on the bristles. Each toothbrush sample goes to a different agar plate. Remember to label your plates with a sharpie marker. Indicate the condition and section off 3 areas. Innoculate one section of 3, by gently streaking the bristles across the surface of the agar.

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BIOL 2440-KJ Fall 2020 5. From the initial section, you will streak for isolation as learned in the week 1 lab. Use your disposable inoculation loops to streak for isolation by dragging from section 1 into 2. Use a fresh loop to streak from 2 into 3. Note-some kits came with swabs instead of loops. Please use the swabs in place of the loops. 6. After streaking, place the lid on the plate and seal it off with the Parafilm. Place your plates in a safe location away from food, pets, and children. Make sure the plate is agar side up and leave to incubate at room temperature for at least 3 days before observing. 7. Once growth is observed, without opening your plate, snap a clear picture for your report. Clean your incubation space with the lab disinfectant. To do this, wet a napkin or paper towel and wipe down the surface. 8. Return all supplies to your Ziploc bag and seal. Return your materials to school, in the sealed bags within the next 2 weeks. Please use schedule an appointment using the Canvas Calendar to set up your return time and/or next in-person lab.

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BIOL 2440-KJ Fall 2020

Toothbrush Experiment Homework: Lab Write-up Use the outline below to write your lab report. Your lab report should be written with complete sentences and in paragraph form. Use the bold headings to mark each section of your write-up.

Introduction (Provides background information to support your data.) 2pts 1) Make a broad statement about bacterial contamination in bathrooms. In the bathroom there are many contaminants and microorganisms that can encounter a toothbrush, many of which are pathogens. Such as the spray that comes from the toilet, the hands of the human using it, and the place in which the toothbrush is stored. 2) Explain what someone has reported about bacterial contamination in bathrooms. Search for a scientifically relevant and peer reviewed article and place the quote or summarize this paper here. Cite your article following your quote or summary with a parenthetical citation with the following format: (Author, Journal, Year of publication). In an article I read, the experiment was close to the one performed for our lab. A set of toothbrushes were put into a bathroom with a toilet for 1 and 3 months and another set of toothbrushes were placed into a bathroom with no toilet for 1 and 3 months as well. The results were the toothbrush placed into a bathroom with the no toilet for 1 and 3 months had isolated microorganisms whereas the ones placed in the bathroom with a toilet for 1 and 3 months had isolated and clumped microorganisms. They’re conclusion was, “Hard deposit on the toothbrush head between bristle tufts is a nidus for growth of micro-organisms, which not only affects the oral health but also affects the general health of an individual”(Indian J Dent, Pubmed.gov, 2011). This really made me realize how much and how many times we are exposed to bacteria. A lot of people don’t think about it because we cannot physically see the bacteria growing, however it is very important to think about as it is affecting our health. 3) Complete the hypothesis. Notice the “if,then” logic used to form this statement. a. If an aerosol of bacteria is created when the toilet is flushed, then a toothbrush left out in the open should have a lot more bacteria on it, while a toothbrush left in 3

BIOL 2440-KJ Fall 2020 a cupboard or drawer will still have bacteria on it, but should be less than a toothbrush that is left out.

Materials and Methods (Provides all information required to repeat your work) 2pts 1) What tools were used for this experiment? The materials used in this lab were two brand new toothbrushes, 2 nutrient agar plates that were sterile and sealed with parafilm, 4 strips of new parafilm, 4 sterile disposable inoculation loops, and finally 50ml conical tube with disinfectant for the lab space.

2) Outline the methods used in this experiment. Include the answers to the following questions: What experimental condition did you use to incubate you toothbrush? How long did you expose your toothbrush? How did you transfer your toothbrush back to lab? For this experiment, done in the bathroom, one condition included leaving the toilet seat up with a brand new toothbrush left out on the counter in my bathroom while the other condition included leaving the toilet seat up, but this time putting a brand new toothbrush into a cabinet in my bathroom. Each morning, I wet the toothbrush with water from the bathroom sink and then placed them back in their designated spot. I went throughout each day flushing the toilet with the seat up. I did this routine for 5 days straight. On the night of the fifth day, I labeled one agar plate counter toothbrush and the other cabinet toothbrush. I inoculated the agar plate labeled counter toothbrush with the toothbrush that was left on the counter and the other plate labeled cabinet toothbrush with the toothbrush put in my cabinet. After inoculating each plate using the streak method, I placed parafilm on each plate. Afterword, I used the disinfectant to clean the area in which I inoculated the plates. I then left the plates, agar side up, for 3 days and then observed the growth. Then again, I cleaned the space with disinfectant. I used a plastic bag to place all materials in and took them back to the lab. 3) What media and incubation time was used to culture the bacteria from your toothbrush once brought back from your home condition? The media used was nutrient agar plates and incubated for 3 days before observing.

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BIOL 2440-KJ Fall 2020

Results (Explain the tests performed and the observations from those tests) 4pts 1) Provide a rationale statement for your toothbrush experiments. Remember this is a two sentence statement where you define the experiment and give a reason for doing this experiment. The experiment is to determine where the toothbrush is most exposed to bacteria. This lab will help show how environmental factors affect the number of microorganisms on a toothbrush. 2) Write detailed observations of the bacterial growth on solid and in liquid media. Reference the Microbes in the Environment lab for descriptive terms for colony and liquid growth. Do not provide conclusions or further analysis of bacterial contamination in bathrooms in general. You will do this in your Discussion section. We did not have any liquid media for this lab. We used agar plates to grow bacteria. After inoculating each toothbrush with bacteria by putting them in different levels of exposure in the bathroom, we were able to observe bacteria growth. Most of the bacteria grown was small with entire, white or clear circular margins. I’m sure if we did use liquid media it would be cloudy in the situations where bacteria was present. 3) We have chosen 4-6 six different experimental conditions for our toothbrushes. Complete the table below with the results you observed in class. You will need to examine toothbrushes from other lab groups. Fill in the table below with your results. If your conditions are different, please change them accordingly. Also, don’t forget to give your table a descriptive title.

Table 1: Bacteria collected from the bathroom on toothbrushes from different exposure. *PLEASE NOTE: I BASED RESULTS OFF OF MY EXPERIMENT RESULTS NOT THE ONES UPLOADED ON CANVAS BESIDES POS & NEG CONTROL* * I noticed on canvas we were only supposed to see growth on the cabinet/seat up plate. However, I did see little growth on the counter/seat up plate as well.* Experimental Condition Results No incubation (neg control-see Canvas) I cannot tell by the picture for sure, looks like no bacterial growth was Direct Contamination (pos control-see Canvas) 5

shown. White/cream in color, lots of

BIOL 2440-KJ Fall 2020 bacterial colonies, looks like circular colonies, mixture of small and medium size colonies, raised, entire margins Toothbrush on counter, Flush with seat up

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produced limited bacterial colonies which were had a white color, an irregular form, raised elevation , and

BIOL 2440-KJ Fall 2020

undulate margins produced limited bacterial colonies which were had a white color, an irregular form, raised elevation , and 7

BIOL 2440-KJ Fall 2020

undulate margins limited bacterial colonies, yellowish/cream in color, irregular form, flat, entire margins, small, circular. Few in second section in Toothbrush on counter, Flush with seat down Toothbrush in cabinet, Flush with seat up

my picture Didn’t do Clear, circular colonies, raised elevation, and entire margins, punctiform, growth in second

Toothbrush in cabinet, Flush with seat down

section Didn’t do

4) Add a figure and a figure legend below the figure. See the recorded PowerPoint for instructions.

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BIOL 2440-KJ Fall 2020 Image 1

Image 2

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BIOL 2440-KJ Fall 2020

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BIOL 2440-KJ Fall 2020

Image 3

Image 4

Figure 1: Bacteria collected from the bathroom on toothbrushes from different exposures. Image 1. This was taken from experiment 3 in the table above; inoculated control. Image 2. This was taken from experiment 2 in the table above; inoculated with E. Coli as a positive control. Image 3. This was taken from experiment 1 in the table above; there was no incubation for the negative control. Image 4. This was taken from experiment 5 in the table above; inoculated control.

Discussion (Apply the observations in the results section to conclusions about microbial growth on items in your bathroom) 2pt 1) Briefly summarize your conclusions. Doing this experiment, really made me realize how important it is to be aware of how much bacteria we are exposed to. It made me understand the importance of how often you should switch out your toothbrush as well as how much bacteria are actually on the toothbrush that I use every day and night. These toothbrushes were only put in this environment for 5 days and still managed to grow bacteria on them. My toothbrush that I use has probably been in use for way longer than it should be. This experiment helped to see that many bacteria can form in a little amount of time.

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BIOL 2440-KJ Fall 2020 2) Provide an explanation for these conclusions by relating your observations to the state of bacterial contamination in bathrooms. My quote from above that I referenced from my article made me believe this experiment showed us that even though we brush our teeth daily, it doesn’t always mean that you are keeping your oral health up, it could actually be the reason you are sick.

References: Provide the full citation of the article that you referenced in the intro. -1pt if missing Karibasappa, G. N., Nagesh, L., & Sujatha, B. K. (2011). Assessment of microbial contamination of toothbrush head: an in vitro study. Indian journal of dental research: official publication of Indian Society for Dental Research, 22(1), 2–5. https://doi.org/10.4103/0970-9290.79965

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