Throat Spray VS. Home Remedy: Which is More Effective on Bacteria in the Human Throat
Prathyush Adapa, Joel Estrada, Julianne Zapanta, Brandon Folts
Biological Sciences 302
California State University, Fullerton
10 December, 2018
Abstract
The healthy throat microbiota consists of coryneform and streptococci families. During times of sickness there are a variety of remedies available to alleviate the patient of a sore throat. One of those methods is to use over the counter throat spray manufactured by prescription drug companies in order to get rid of the sore throat. Another method is to use home remedies passed through generations which could be a combination of vinegar, water, salt, and lime juice. Although both of these methods have been effective in the past, one must test them in a controlled environment in order to see which treatment is more effective for specific bacteria. During experimentation it was expected that the throat spray would be more effective than the home remedy liquid being tested. As well as bacteria from Cornyeform bacteria, Streptococcus, and Staphylococcus genus would found in throat culture. The effectiveness of the throat spray was prevalent as its zone of inhibition when tested on a Kirby-Bauer plate was 11.0mm compared to holistic remedy 9.0mm. As for bacteria identification only Staphylococcus aureus could be identified in throat swab culture.
Introduction
During times of illness in particular cultures, the use of home remedy treatments are used in ways of relief and possibly to cure the actual ailment. An example of this is the use of a liquid Mexican home remedy with ingredients of vinegar, hot water, lime and salt is used to treat sore throats. This liquid supposedly, kills bacteria that are in the throat causing inflammation to decrease, resulting in the patient being cured. Treatments like these have been used to help relief in patients with strep throat, while infected with the flu, or even in singers where their throats become inflamed and in need of relief (4). However, such treatments can be misleading, are not effective, and in ways harmful as they substitute for actual treatments that have been tested to work against particular infections as well have certain characteristics that help prevent future bacterial infections (3). Sprays can also be more effective as they are designated to attack unwanted bacteria while not attacking actual host cells or even the types of bacteria that are helpful and needed in the body such as Bacteroidetes, Coryneform bacteria, and Proteobacteria (5).
Active ingredients in throat sprays and lozenges aim to provide symptomatic relief of pain. Some have disinfectant properties to decrease the likelihood of further infection
Now for the throat it contains one of the richest and diverse microbiota environments in the body. Because the throat is part of the primary response to digestion, a multitude of bacteria is needed in order to digest the various food. Not only that, the mucus of the throat is also an important factor in trapping bacteria and neutralizing the harmful strains before it gets digested by the stomach. However, during times of sickness, the aid of any throat relieving device is needed. This can come in the form of throat sprays or holistic medicines. But the true question is, which method is the most effective in clearing the throat. For this project, a general CVS brand throat spray was tested against a throat relief home remedy consisting of hot water, vinegar, salt, and the citric juices from a lime. Therefore, if we are to test this, we believe that the throat spray will be more effective than the home remedy at killing off the bacteria in the throat. As for identification of bacteria, throat swab culture will contain Cornyeform Bacteria, specimen from Streptococcus genus, and Staphylococcus aureus.
Materials and Methods
General: Salmonella Typhimurium (Ames Test), CVS – Branded Throat Spray,
Name of Method Used: Ames Test
In the first experiment, we had had used a variety of microbiology techniques in order to determine which one was more effective, the throat spray or the home remedy. We first performed an Ames test which allowed us to see if the throat spray or the home remedy was effective against the Salmonella Typhimurium. There was no change in this procedure from the procedure listen in the Biology 302 Lab Manual provided by the California State University, Fullerton. We then performed a Kirby Bauer to test with the same procedure listed in the Biology 302 Lab Manual provided by the California State University, Fullerton except we substituted the antibiotic with the throat spray and home remedy. These two tests allowed us to identify which remedy would be most effective.
General:
Name of Method Used: Biochemical Tests
In the second experiment, a multitude of various microbiol techniques were used to identify the various bacteria species present in the the throat of the sample subject. In order to create cultures of bacteria, we first swabbed the throat of our lab partner, Joel Estrada, and plated that on a TSA plate in order to have a culture that we could study. We then tested our first hypothesis to understand which method was more effective at killing bacteria, the throat spray or the home remedy.After growing the bacteria in TSA cultures for 48 hours, we then performed an Ames test was well as a modified Kirby-bauer test in order to After growing the TSA culture for 48 hours, we had decided to inoculate that culture in TSB broth so that we could perform the various biochemical reactions whose procedures are found within the lab manual for Biology 302L at CSU Fullerton. After inoculating the bacteria in TSB broth, we followed the procedures in the lab manual in order to conduct a glucose, sucrose, and mannitol test as well as inoculate our bacteria on SIM (Sulfur, Indole, Motility) and TSI (Triple Sugar Iron Agar) media, which tests for the fermentation in order to determine what species are present. We then also inoculated the sample on MAC and Blood Agar and did the glucose, sucrose, mannitol, SIM, and TSI tests in order to see what bacteria may have been in the sample. The Mac plate is a differential medium that tests to to see if the bacteria is an Enterobacteriaceae or is gram positive, since it inhibits the growth of gram positive bacteria. The MSA plate is differential media that tests for the growth of Staphylococci, and inhibits most other bacteria species. The Blood Agar plate is a different media that tests for α, β, and γ hemolysis. After these tests were conducted, we had also done a gram stain on the bacteria that were grown on each media to again isolate the bacteria and correctly identify what was present.
Results
Grown samples were taken from TSA plate that had grown bacteria from throat swab. Specimens were plated on MSA, blood agar, EMB, and MAC. Both MSA and blood agar had strong growth in each plate. Along with growth, specimen 1 on MSA plate changed agar color from faded dark pink to bright yellow. As for blood agar the color of growth was white and lysis of blood agar around colonies grown suggests that specimen is β-hemolysis. Gram stains were then performed on specimens that had shown significant growth in plates. Gram stains were performed on blood agar specimen, MSA specimen, and original TSA specimen. For specimens TSA and MSA, gram stains were positive while for blood agar result was gram inconclusive as it had both gram positive and negative bacteria.. All three samples, however, had cocci morphology.
Figure 1. Gram stains of (from left to right) TSA specimen, MSA specimen, and blood agar specimen; all from original throat swab grown in TSA. Specimens from TSA and MSA are gram positive; blood agar was inconclusive. All specimen have cocci morphology.
Table 1: Results of recorded growth of specimen 1 on TSA plate from throat swab. Plates used were MSA, blood agar, EMB, and MAC.
Specimen 1 (TSA)
MSA
Growth
Blood Agar
β-hemolysis
EMB
Negative (Little to no growth)
MAC
Negative (Little to no growth)
Multiple biochemical tests were conducted on samples taken from TSA, blood agar, and MSA plates to further identify bacteria in grown samples. Fermentation reactions of sugars were conducted on the three bacteria samples; specimen 1 from TSA plate produced positive growth and fermentation for glucose, sucrose, as well as mannitol; each broth produced gas as well. Specimen 2 from blood agar produced growth in each sugar broth but none in mannitol broth; as for fermentation production, it did not ferment in mannitol broth but did ferment in glucose and sucrose sugar broths; only produced gas in glucose broth. MSA bacteria sample produced positive results for growth and fermentation in all sugar broths; gas was produced in all broths but glucose. When conducting SIM testing, results in sulfur reduction and indole production produced negative results; however, positive results were seen for motility in all SIM agars for the three samples being tested. Performing TSI (triple sugar iron agar) testing all samples had same results where each TSI slants had yellow slants and butts indicating fermentation of glucose, sucrose/lactose, but no fermentation was seen for H2S.
Table 2: Table showing results of all tests done on all three specimens. Test were performed over three different lab periods. Results are recorded qualitatively in the chart below.
Specimen 1 (TSA)
Specimen 2 (Blood Agar)
Specimen 3 (MSA)
Glucose
(+) Growth (+) Fermentation
(+) Gas
(+) Growth (+) Fermentation
(+) Gas
(+) Growth (+) Fermentation
(-) Gas
Sucrose
(+) Growth (+) Fermentation
(+) Gas
(+) Growth (+) Fermentation
(-) Gas
(+) Growth (+) Fermentation
(+) Gas
Mannitol
(+) Growth (+) Fermentation
(+) Gas
(-) Growth (-) Fermentation
(-) Gas
(+) Growth (+) Fermentation
(+) Gas
SIM
Sulfur (-)
Indole: (-)
Motility: Motile
Sulfur (-)
Indole: (-)
Motility: Motile
Sulfur (-)
Indole: (-)
Motility: Motile
TSI
Glucose: (+)
Sucrose/Lactose: (+)
H2S:(-)
Glucose: (+)
Sucrose/Lactose: (+)
H2S:(-)
Glucose: (+)
Sucrose/Lactose: (+)
H2S:(-)
More experimentation was performed to test mutagenicity of both throat spray and home remedy liquid, as well as testing each liquids strength of inhibition on bacteria sample grown on TSA plate. Ames test was carried out to produce results on whether each liquid had mutagenic traits as well as testing their toxicity when introduced to a cultured sample of Salmonella typhimurium his-, uvrB-, rfa-. Minimal growth was observed on minimal control plate on the side of sodium azide filter disc but no growth was seen on both minimal plates for home remedy and throat spray. When observing complete media plate for both liquids a small zone clearing surrounded the two filter discs in their respective complete media plates. To test the strength of inhibition for the CVS throat spray and the home remedy, Kirby-Bauer test was performed. When examining the dipped filter disks of both liquids, CVS throat spray had a zone of inhibition of 11.0mm while the home remedy had one of 9.0mm.
Discussion
After performing the Ames Test for Method 1, we were able to determine that the throat spray was more effective than the home remedy. First, we established that the throat spray and home remedy were toxic as we analyzed the results of our ames test. This was a result of the complete plates having clearing zone when inoculated with throat spray and home remedy. This clearing zone shows that both substances were toxic to the bacteria and that mutagenicity could not be determined by the Ames Test. When analyzing the Kirby Bauer results, we were able to see that there was zone of inhibitions for both the throat spray and the home remedy. This was expected as we knew from our Ames test that both were toxic. The Kirby Bauer therefore allowed us to analyze how effective each treatment was on the bacteria. Our results showed that the zone of inhibition of throat spray was considerably larger at 11.0mm whereas the home remedy had a zone of inhibition of 9.0mm. Therefore, we can conclude that the throat spray is more effective than the home remedy. This can be the result of the throat spray containing chemicals like phenol which are more specific at targeting and killing bacteria compared to the home remedy which is less specific and contains high concentrations of salt which can affect the ability of bacteria to proliferate.
After studying which medicinal therapy would be more effective, we analyzed the data pertaining to the identification of the bacteria present in our sample. All media were inoculated with bacteria grown on our initial TSA plate. When analyzing our Blood Agar plate, we saw that the specimen clearing around the colonies. This in turn suggests β-hemolysis and could be either Staphylococcus aureus and Streptococcus pyogenes. When observing the MAC and EMB plate, we received a negative result as there was little to no growth which indicates that gram positive bacteria were inhibited from growing. This in turn further provides evidence that the bacteria in question is either Staphylococcus aureus and Streptococcus pyogenes as they are both gram-positive bacteria. When observing the the MSA plate which is a selective differential media with high saline levels that selects for the growth of Staphylococcus aureus, we were able to observe growth and fermentation, which alludes to the bacteria being Staphylococcus aureus. We could not conclude if Coryneform Bacteria as a result of the tests that we were able to perform. We may have had some in our blood agar plate as a result of some different results that occured when inoculating the blood agar colonies into various biochemical tests however, other bacteria may have contaminated the agar as well.
Since it was observed that the throat spray was more effective against the bacteria that grew on the ames plate it could be inferred that the pharmaceutical medicine is more effective in the treatment of illnesses compared to home remedies. Although the throat spray is more effective, the argument can be made that the home remedy is still a viable option for relieving symptoms and killing some bacteria due to its zone of inhibition being only 2mm smaller than the the the throat spray. This difference is due to the CVS throat spray being more specific for certain the bacteria, and the home remedy only relieves symptoms as a result of its over the counter ingredients that have less specificity for killing certain bacteria. Overall, this could lead to further research being done testing multiple different scenarios throughout the human body to see if this is true for other microbiota.
References
Graevenitz, Alexander. “Cornyeform Bacteria in Throat Cultures of Healthy Individuals.†Current Neurology and Neuroscience Reports., U.S. National Library of Medicine, www.ncbi.nlm.nih.gov/ pmc/articles.
Lauer , Brian. Effect of Atmosphere and Duration of Incubation on Primary Isolation of Group A Streptococci from Throat Culture. Journal of Clinical Microbiology, Feb. 1983.
Farrer, F. “Sprays and Lozenges for Sore Throats.†South African Family Practice, vol. 54, no. 2, 2012, pp. 120–122.,
Abaza, M. M., & Sataloff, T. (1999). Sore throats in singers. Journal of Singing – the Official Journal of the National Association of Teachers of Singing, 56(1), 33-38. Retrieved from https://search-proquest- com.lib-proxy.fullerton.edu/docview/1400794?accountid=9840
American Society for Microbiology. "Bacteria in the nose and throat: Most comprehensive comparative analysis." ScienceDaily. ScienceDaily, 22 June 2010.