Enterococci are bacteria that are included in the intestinal flora of humans and are relatively harmless. One method Enterococci maintain a symbiotic relationship is by hydrolyzing esculin in the presence of bile (Susan L. Fraser, 2012). However Enterococci are in the top three-nosocomial infections. When infected, enterococci can cause numerous infections such as urinary tract infection coupled with fever (Susan L. Fraser, 2012). More specifically, Enterococcus Faecalis causes urinary tract infections. Enterococcus Faecalis (E. Faecalis) is a gram-positive bacteria that is difficult to eliminate.
With an increasing dependency on antibiotics, E. Faecalis is becoming feared due to its intrinsic antibiotic resistance (Wesley Glick, 2000). This strain is naturally immune to weaker levels of penicillin because of its ability to synthesize cell wall components in the presence of penicillin. Instead of allowing this antibiotic to destroy the cell wall, E. Faecalis produces penicillin-binding proteins that inhibit penicillin from disintegrating its cell wall (Susan L. Fraser, 2012). Thus a more effective antibiotic must be used.
For the purposes of this study, Penicillin, Tetracycline, and Ampicillin have been chosen to counter E. Faecalis. Based on a previous study completed by Wesley Glick, E. Faecalis is intrinsically resistant to weak Penicillin and Tetracycline (Wesley Glick, 2000). Therefore a hypothesis can be generated stating that Penicillin and Tetracycline will be partially ineffective towards eliminating E. Faecalis. However, since the Penicillin and Tetracycline used in this experiment are more potent, it is predicted that all three antibiotics will be effective against E. Faecalis, with Ampicillin exhibiting the largest zone of inhibition. Materials Enterococcus Faecalis was taken from a stock sample and was spread across a blood agar plate. The plate was divided into four quadrants: three for antibiotics and one for control. A small disk was chosen, each containing one antibiotic of Penicillin, Tetracycline, or Ampicillin and placed at the center of each quadrant on the blood agar plate. The plate was sealed with tape, and was left to incubate at 37°C for twenty-four hours.
Then the plate was left in a refrigerator for one week. After, the agar plates were taken out for observation. The zone of inhibition, or region around the antibiotic disk absent of bacteria, was measured and recorded for each quadrant (General Biology Laboratory Manual, 2010). To determine if the bacteria was gram-positive or gram-negative, a slide must be prepared. A sample of E. Faecalis was swabbed onto a slide and then first stained with crystal violet. After about one minute, the slide was rinsed with water and then gram iodine was poured onto the slide.
Once again the smear was rinsed with water only this time a 95% alcohol/acetone solution was placed onto the slide briefly and then rinsed off with water. Finally Safranin was dropped onto the slide for one minute and then the slide was rinsed with water yet again. The slide was finally blotted dry (General Biology Laboratory Manual, 2010). Results The incubated blood agar plate revealed noticeable differences between the four quadrants. Quadrants one through three exhibited a zone of inhibition because of the presence of antibiotics, but quadrant four was control and allowed normal growth.
Based on the experiment performed, Ampicillin proved to be the strongest antibiotic followed by weaker antibiotics Penicillin and Tetracycline. Ampicillin’s zone of inhibition was roughly one hundred and thirty percent that of Ampicillin and Penicillin. If we consider that a zone of inhibition greater than 10mm is effective, then the results achieved disproved the hypothesis because Penicillin and Tetracycline had zones of inhibition both larger than 10mm. However they were not as potent as Ampicillin. Ampicillin managed to eliminate thirty percent more of the E. Faecalis.
These results are supported by a similar experiment performed by Butaye, Devriese, and Haesebrouck. When they performed the experiment, it was found that E. Faecalis had no resistance to Ampicillin. Not only was Ampicillin extremely effective, but E. Faecalis was found to have high resistance to Tetracycline (Patrick Butaye et al. , 2001). In another study completed by Serror, eighty-one percent of E. Faecalis was still present after a dose of antibiotics. The antibiotic E. Faecalis was most resistant to was Tetracycline (Emmanuel Jamet, 2011).
Again the data that Tetracycline was not a popular choice to treat E. Faecalis was supported. Based on M. D Susan L Fraser, Ampicillin is the antibiotic of choice to treat E. Faecalis over Penicillin because it is more sensitive (Susan L. Fraser, 2012). Congruent with this clinical choice, Ampicillin proved to have the larger zone of inhibition, 20. 8 mm, compared to Tetracycline’s zone of inhibition, 15. 3mm. Though these results countered the hypothesis, the prediction held true. On a large term, Ampicillin should be used over Penicillin and Tetracycline because E. Faecalis is more sensitive to Ampicillin. However, based on this experiment alone, this data cannot be applicable because of the small sample size. Only four other copies of this experiment were performed, limiting the validity of this data. This exact lab would have to be performed at least thirty more times to be considered plausible. Also, the E. Faecalis used was drawn from one sample. The bacteria from this culture could have been weaker than other E. Faecalis, skewing the data. To minimize error, multiple samples of E. Faecalis cultures would have to be utilized.
Additionally, this strain of bacteria was cultured in a lab and not obtained from a hospital environment. Therefore the results cannot be used in clinical situations because the experiment is not representative of an actual predicament. An abnormality did occur. Based on previous studies, tetracycline eliminated little to no E. Faecalis. But in this particular lab, the data revealed otherwise. This error could be the result of a faulty sample. The E. Faecalis could have been a particular colony that was sensitive to tetracycline instead of the norm of being resistant.
Regardless of the errors and limitations, this data does provide viable information. These three antibiotics are all effective, but ampicillin would be the best choice. Knowing that there are multiple antibiotics provides another solution incase the patient reacts to ampicillin. Even though it may not be as effective, it can hopefully help suppress E. Faecalis long enough for the body’s natural immune system to take over. To address the limitations and make this experiment more relevant to clinical situations, E. Faecalis should be obtained from patients and then cultured. The experiment should be performed and repeated numerous times. Then the results from these experiments would create a more accurate picture for the use of either Ampicillin, Tetracycline, or Penicillin to combat Enterococcus Faecalis. References Emmanuel Jamet, Elodie Akary, Marie-Ange Poisson, Jean-Francois Chamba, Xavier Bertrand, Pascale Serror. (2011). Prevalence and characterization of antibiotic resistant Enterococcus faecalis in French cheeses. Food Microbiology, Volume 31, Issue 2 191-198.
General Biology Laboratory Manual. (2010). Lab Topic 4: Bacteria. Boston, MA: Pearson Learning Solutions. 41-57. Patrick Butaye, Luc A. Devriese, and Freddy Haesebrouck. (2001). Differences in Antibiotic Resistance Patterns of Enterococcus Faecalis and Enterococcus Faecium Strains Isolated from Farm and Pet Animals. Antimicrob. Agents Chemother. vol. 45 no. 5. 1374-1378. Susan L Fraser, MD. (2012). Enterococcal Infections Treatment and Management. 1-5. Wesley Glick. (2000) Enterococcus Faecalis. Revised August 2000.