Gunjan in the mouthwash trial will show resistance to

 Gunjan PatelIB Biology HL Yr:2 TheEffects of mouthwash on the human mouth and how it affects the production ofbacteria Research Question:How doesmouthwash affect the growth of bacteria in the human mouth? Aim:Thisinternal assessment aims to determine how variations of mouthwash affect theproduction of bacteria in the human mouth. This is going to be achieved byusing sterile cotton swabs to swab the mouths (teeth and gum lining) of fiveindividuals and then placing it in a zig-zag pattern on a petri dish lined withagar. It will be measured by using different types of mouthwashes, such asorganic, different brands and different contents in the mouthwash.  It will attempt to determine if bacteria inthe mouthwash trial will show resistance to the mouthwash during the week byrecording the growth daily in a qualitative measure.

 Background:The two types of mouthwash that people can buyare, cosmetic and therapeutic. People can buy therapeutic mouthwashes in thestore or also by prescription from a doctor. Meanwhile, cosmetic mouthwash relievesbad breath for a short period of time and in place of it, will be a refreshingtaste but it does not have any active ingredients while therapeutic does.Ingredients in a therapeutic mouthwash may include: cetylpyridinium chloride,fluoride and peroxide. Mouthwash is also helpful because it can accessplaces in the mouth where a toothbrush cannot.

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One of the most common uses oftherapeutic mouthwash is to treat halitosis. That is also caused from thebuildup by food fragments within the mouth that was not removed due to lack ofdental hygiene such as brushing for a short time and/or not using floss. Peoplewith halitosis have cavities or gum disease.

However, people that do not have halitosis,also have buildups of bacteria in the mouth, such as Staphylococcus aureus,Streptococcus pyogenes, Helicobacter pylori, Streptococcus mutans, andPorphyromonas gingivalis. People have studied the efficacy of different typesof mouthwash and have shown that it provides an effective control of bacterialgrowth as well as decreased pain following dental procedures. Anotherpopular type of mouthwash is organic mouthwash, as it only contains naturalingredients which only removes the bad bacteria.

Xylitol, oils, and plant-basedextracts are all ingredients in a typical organic mouthwash. While Listerine isthe most popular/used ‘name brand’ mouthwash.  Personal engagement:I choseto conduct this experiment because I have always been curious why eachmouthwash says that they are number one. I wanted to find a way to prove whichmouthwash is the best at reducing the count of colonies of bacteria and to dothis I had four members of my family swab their mouths at the beginning of theweek and put it in four agar plates to let it grow throughout the week. Eachday after that, I assigned each member of my family a specific mouthwash and atthe same time making sure each person ate the same type of food to keep my dataaccurate. They would use the mouthwash once at the end of each day.

And at theend of the week, I would swab their mouths and put it on an agar plate to letthe bacteria grow.  Mouthwash:The mouthwashes that were used and the person who used the mouthwashare listed below:-       ListerineAntiseptic Mouthwash containing alcohol (Person #1)-       CrestScope Mouthwash (Person #2)-       CrestPro-health CPC Antiplaque Mouthwash (alcohol-free) (Person #3)-       TheraBreath Oral Rinse (Person #4)o  This mouthwash is clinically tested and usesOXYD-8 to attack only harmful bacteria and eliminate the bad taste in yourmouth.  Hypothesis: This experiment is to determine how variationsof mouthwash affect the growth of bacteria, and it is expected that, out of thelist of the used mouthwashes in my experiment, Listerine and Scope brands will havethe least number of bacteria grown. This is due too that they are the only onesthat contain alcohol as an active ingredient, which is a known antiseptic thatis used worldwide as a disinfectant in multiple places such as a hospital.Furthermore, the “organic” mouthwashes that are recommended by dentists,contain natural/herbal products. The data on their efficacy are lackingcompared to the “name brand” products. Bacteria that are exposed to mouthwashdaily, will develop resistance and will determine if the bacteria show anysigns of resistance for one week.

I hypothesize, that the Listerine antiseptic mouthwashes will be themost effective in inhibiting the growth of bacteria with the organic”herbal” brand being the least effective.  Variables: Dependent Variable: –       The amount of bacteria present –       Changes due to mouthwash use –       Number of colonies Independent Variable: –       Type of mouthwash –       Time Controlled Variable: –       Temperature where petri dishes are stored  –       Light conditions where petri dishes are stored –       Size of Petri Dish –       Type of Agar –       Time between both trials –       Trial without mouthwash  Materials:-       (10)Pre-poured sterile LB-agar plates-      (10) 6-inch sterile cottonswabs –      fourdifferent brands of mouthwash (different ingredients) –       permanentmarkers –       Rulers-       Scotchtape-       Agarplates –       Incubator-       Heatlamps Method:1.     Obtaineight agar plates and sterile swabs from Amazon and get a heat lamp in theready position.

2.     Usinga sterile cotton swab, swab the inside of four individual’s mouths and makesure to swab the front teeth along with the gums while twirling the swab in acircular motiona.    Ensure that the individuals are eating the samecuisine until the end of the trial3.     Afterswabbing for individuals, remove the lid of the agar plates and carefully twirlthe cotton swab across the plate in a semi zig-zag pattern, and then label eachagar plate with the name of the individual (also mouthwash when repeatingmouthwash)4.     Closethe lid of the agar plates and slowly place them upside down and move theplates to under the heat lamp.5.     Leavethe agar plates under the heat lamp for 48 hours and after time has passed,remove the plates from the lamp and take a picture of the plates to documentthe growth6.

     Putthe agar plates on a table and near the lamp and put it back under the heatlamp for 4 days. Continue to take pictures every 24 hours 7.     Obtainfour different types of mouthwashes, such as Listerine antiseptic, Crestnon-alcoholic, Crest antiseptic, and organic mouthwash, to determine whichmouthwash is most effective in killing bacteria.8.     Afterthe mouths have been swabbed (step 2), assign a mouthwash to each individual(different ingredients, such as organic/non-alcoholic).

And after dinner everyday, each individual will need to use the mouthwash they have been assigned9.     Afterfour days have passed, repeat steps 2-6 but with using mouthwash and also usingan incubator to let the plates with agar grow on it.Data and Analysis:Table 1: The data recorded, describing the growth of bacteria in the trial withoutmouthwash, in relation to it filling the agar plate Days Agar plate, person #1 Agar plate, person #2 Agar plate, person #3 Agar plate, person #4  Day 1 (Monday) None (0%) None (0%) None (0%) None (0%) Day 3 (Wednesday) Light growth (<25%) Light growth (<25%) Mild growth (25-50%) None (0%) Day 4 (Thursday) Mild Growth (25-49%) Mild Growth (25-49%) Intermediate growth         (50-75%) Light growth (<25%) Day 5 (Friday) Mild Growth (25-49%) Intermediate growth       (50-75%) Intermediate growth         (50-75%) Mild Growth (25-49%)                  Person #1, Day 3:                                                Person#1, Day 5:ThePictures above demonstrate the growth of bacteria at the end of the firsttrial, which was day #5. With person #1 being on the left and person #4 beingon the right. On days 1,3,4,5 I recorded the growth of bacteria on the agarplate and took pictures to show the growth of it on the agar plate. Calculatinghow much of the colonies of bacteria, filled up on the plate in table #1. Wecan see that the number of colonies differs in each person due to the meal theyate and the bacteria present in the mouth.

 Table 2: The data recorded with the use ofmouthwash in the trial, describing the growth of bacteria in the trial withoutmouthwash, in relation to it filling the agar plate Days Agar plate, person #1 Agar plate, person #2 Agar plate, person #3 Agar plate, person #4  Day 1 (Monday) None (0%) None (0%) None (0%) None (0%) Day 3 (Wednesday) Discreet growth (<5%) Light Growth (<20) Medium growth (25-40%) Light Growth (<20) Day 4 (Thursday) Light growth (<20%) Medium growth (25-40%) Medium growth (25-40%) Light Growth (<20) Day 5 (Friday) Medium growth (25-40%) Medium growth (25-40%) Intermediate growth (50-75%) Medium growth     (25-40%)          Thesepictures were taken on day #3, and starts from person #1 on the left and endson person #4 on the right. We can see that the three mouthwashes that containalcohol (1,2,4) reduced the growth of bacteria the most, with person #3 whoused nonalcoholic mouthwash, bacteria growth was noticeable and covered atleast 50% of the agar plate. Thesepictures were taken on day #5, and starts from person #1 on the left and endson person #4 on the right.

We can see that at the end of the trial, bacteria growthslowed down even more after being left in an incubator to grow for four days.Except person #3, who used Crest nonalcoholic mouthwash. We can determine fromthis test that Listerine was the best at reducing the growth of bacteria, withCrest nonalcoholic mouthwash being the worst.      Conclusion: Thehypothesis stated in the beginning does support the conclusions that wereproposed from my data, The Listerine mouthwash was the most effective inkilling the bacteria due to its key ingredient alcohol, while the Crestnonalcoholic mouthwash was the least effective due it not having alcohol. Thegraphs showed the number of colonies of bacteria present in the agar plate, andit is evident that after day 3, the colonies stopped growing or reduced afterbeing left in the incubator during the trial. The graph also shows that person#3 who did not use alcohol, the colonies of bacteria on that agar plate had ahigh number of colonies of bacteria growing, and that the mouthwash offered minimalresistance.  One ofthe limitations of this experiment was that there were not multiple trials andbecause of that, I was unable in calculating the standard deviation of thecolonies of bacteria. Another limitation, is that I did not have access to anincubator or more people to make my trial more accurate.

I would have to checkon the agar plates at an appropriate time and sometimes, the room would belocked for testing. My experiment was conducted at two different places, onewas at home, so I could swab my family members mouth and another was checkingthe incubator at school, which was only at certain times. Also, some of myfamily members ate different foods which could’ve potentially affected my data,such as during work or at the gym.  The agarplates that were used during the trial were purchased from Amazon and were setup to my specifications and included sterile swabs. My first order, the agarplates were damaged and had to request another order be shipped. This delayed myexperiment and reduced the number of trials that could be done to one week.  In orderto combat these errors if i were to redo the trial again, the way the trialsand experiments were conducted would need to be standardized to make theresults more reliable.

Such as have all the materials and equipment set up beforestarting the experiment. Overall, the experiment was organized but the delaysand low number of trials that occurred could’ve been eliminated if all materialwas present before starting and had access to an incubator or a heat lamp in myhouse.