Science Buddies: "Ask an Expert"

Hello!
My daughter (7th grade) wants to sample different public places for bacterial presence. After doing quite a bit of research, we are still left with a few key questions.
1. How to collect samples: we thought to use sterile swabs to swipe the surfaces. Should they be wet or dry? If we wet them, use sterile water? Where can we find sterile swabs? Our pharmacy didn't have them.
2. Most important question of all: How to quantify the data? Her teacher said just look at the plates and express as a percent of area covered. This seems too imprecise to me. How do you do colony forming unit counts? Can we get more than one count on each plate at different times or will we have to mark the covers which will make it hard to see next time? Can she see the colonies without a microscope? (Teacher will loan a scope to take home if needed)

Thank you so much for helping us!

Hi,

Working with bacteria can be dangerous; it is highly suggested that you and your daughter conduct this experiment in a school or other kind of lab.

As for collecting samples, I believe using a dry sterile swab would be sufficient. However, you need to place your sample in a broth, and incubate this for 24 hours in 37 degrees celsius. Doing this will ensure that the bacteria sample is good. After 24 hours, take a sterile inoculating loop, and streak your plate.

When looking at the results, everything will be qualitative. Your daughter will need to look for different shapes and sizes, and how many colonies look similar. I am not sure what her teacher means by looking at the plates and express as a percent of area covered. Bacteria will grow over the entire plate, and not just in certain areas.

These websites might help for inoculation:
http://www.umsl.edu/~microbes/pdf/streakplates.pdf
http://www.madsci.org/~lynn/micro/techniques/streaking/

If your daughter is conducting this in a school lab, which is seriously preferred, she might want to stain the culture to look at the growth further.

http://polymer.bu.edu/ogaf/html/chp51exp4.htm

I don't know what you mean by "getting more than one count on each plate at different times or having to mark the covers which will make it hard to see next time." Would you be able to clarify this?

Labeling the petri dishes is always very important. Have your daughter put the data, initialize her name and put what the project is about. That way it will be easier for her. I would suggest she plate more than one petri dish, to ensure proper inoculation techniques.

Bacteria colonies can definitely be seen without a microscope. But examining growth under a microscope for further investigation is definitely the next step.

I hope this helps.

Sincerely,
Sareena

Sareena,

Thanks for answering back. I understand your answers for the most part but you are thinking from the perspective of a scientist in the lab with tissue culture facilities. Forget that! You have to think of a 12 year old girl working out of her kitchen and bedroom with no incubator or any lab facilities. There is no proper lab at her school either.

We already collected samples from several public places. We moistened a sterile swab with sterile water and swabbed the surface area to be tested. She divided the plate into 4 quadrants and streaked Quad #1, then #2, and so on.

Her positive control is already showing signs of growth (don't laugh but for a positive control we left out a chunk of ground beef on a paper plate at room temp for 6 hrs. then discarded meat, left plate out for another 2 hours before swabbing plate). The negative control is just an unopened Petri dish (we bought the pre-poured kind that is ready to go).

So I still would like to know what quantitative things she can do. You asked what I meant by getting several counts from same Petri dish. I meant that she could plot it out to get a sense of the growth curve if she counted CFU's (or whatever) on Day 1,2,3,4, etc.

As far as qualitative things, can you recommend a good yet easily attainable book/resource to help us identify what may be growing in the plates? Do you have any good guesses as to what the most likely candidates that we will see in her plates. Remember she swabbed shopping cart handles, elevator buttons, escalator handrails, computer mouse at the library, etc. etc. What are we most likely to observe?

My daughter has gotten "so into this" that now she wants to test if the commercially available antimicrobial wipes really work. I was resistant to doing this part because I felt she needed a definite source of bacteria to start with. But maybe if her positive control works out, we could repeat the strategy and use your idea of swabbing and then placing swab in test tube and then later inoculating plates so all would get same volume of the same bacteria-laden solution. Does this sound feasible? My daughter read about measuring "zone of inhibition" for this kind of experiment. Are the numbers (I suppose she would measure the diameter of the no growth zone?) likely to be very different?

Safety-wise, we are wearing gloves while doing all this and plan to use bleach on the plates at the end of experiment.

Thanks for your input.
Maureen

Hello Maureen,

I was unaware of the conditions your daughter was in while conducting this lab, and I did not assume she had all the equipment needed. Safety is of number one priority when conducting labs, and it is important that all precautions are employed. I am not a scientist, but I have conducted a lab similar to your daughter's without the use of expensive equipment.

As for her experiment, the plates seems to be inoculated well; how many hours did you leave this dishes in room temperature?

I don't exactly understand the positive and negative controls. Please correct me if I am mistaken, but wasn't your daughter's project focused on examining different kinds of bacteria that grow in public areas? If this is so, I am interested in understanding why your daughter chose these as her positive and negative controls. Based on my understanding of growing bacteria, positive and negative controls are not needed for this experiment.

As for quantitative analysis, counting the number of bacteria is possible using CFUs. The area in which she took the sample would need to be defined, and then she would be able to calculate the number of bacteria in that area using CFUs. Here is a website that will definitely help:

http://www.newton.dep.anl.gov/askasci/m ... e00323.htm

With regards to qualitative, it would simply be drawing up a chart. I was not able to find any sample charts online, but it would basically be identifying colonies, and describing the shape (circle, oval, she can even draw it), the size (large, small, medium) and whether it is indented, bumpy, round, so on.

There are many posts on Science Buddies about the efficacy of antimicrobial/bacterial products on the growth of bacteria cultures. Visit the "Mouth Microbes" thread - I believe it is on the second page - to get ideas on how to prepare the experimental set-up. I would suggest only plating one kind of bacteria (ex. E.Coli); it is important to control the experiment as best as your daughter can. However, it also depends on the purpose of her experiment. Wipes are meant to kill all bacteria on a kitchen counter, so if she wants to test kitchen counter bacteria, that is also feasible. There just might be problems that tag along with testing wipes against more than one kind of bacteria. For example, if bacteria is found on or near the substance, it would be impossible, with the facilities available to her, to detect if the wipe just doesn't work at all, or doesn't work against a specific kind/kinds of bactera.

Your daughter would need to cultivate the bacteria to ensure good inoculation results so her data is reliable. Here is some information on why broth is helpful:

http://www.newton.dep.anl.gov/askasci/m ... e00677.htm

Here are some websites to give you and your daughter some ideas:

https://www.sciencebuddies.org/mentorin ... roBio_p007
https://www.sciencebuddies.org/mentorin ... ?from=Home

As for zones, she is correct: if the wipe has antibacterial properties, no growth would be visible around the area. Quantiative data collection would be measuring the zone sizes. Zone sizes could vary from different wipes, and different results would indicate varied effectiveness in fighting microbes.

Hope this helps,
Sareena