Antimicrobial Activity of Plant Extracts and the Resistance of Staphylococcus aureus and Escherichia coli to Tea Tree (Melaleuca alternifolia) Extract Ashley Lim, Department of Biology, York College of Pennsylvania The antimicrobial activity of plant extracts are of interest due to the increase in demand for more natural food and medicine products, and because many microbes have been able to build up a resistance to current antibiotics. Tea tree (Melaleuca alternifolia) is a natural antiseptic used in a wide variety of pharmaceutical and cosmetic products (Cox et al. 2000). Much of the research done on plant extracts have concluded that the extracts studied have potential to be used as antimicrobial substances, yet little testing has been done to see if microbes can build up a resistance to these extracts. The antimicrobial activity of commercially available extracts were tested in this study and compared to the results seen in previous studies. Test the antimicrobial activity of tea tree, cypress, bergamot, vetiver, and eucalyptus radiata extracts using an agar and broth assay against Staphylococcus aureus, Escherichia coli, and Candida albicans. Test if Staphylococcus aureus and Escherichia coli could build up a resistance to the tea tree extract. Disk Diffusion Assay Plates divided into quarters. Inoculated with 100 µl microbe. 6 mm disk dipped into extracts (5-10 µl). Blank disk used as negative control. Penicillin and Kanamycin used as positive controls. Incubated 24 hrs at 30°C. Zone of Inhibition (ZOI) measured in mm. Broth Dilution Assay 2-fold serial dilution in a 96-well plate. Final concentrations ranged from 5% (v/v) to 0.01% (v/v). 50 µl microbe inoculated into wells. Incubated for 24 hrs at 30°C. Minimum Inhibitory Concentration (MIC) was found by assessing the turbidity of the wells. Resistance Assay Agar plates inoculated with 100 µl bacteria. 6 mm disk inoculated with 5 µl tea tree extract. Incubated for 24 hrs at 30°C. ZOI was measured, then the most resistant bacteria was isolated. “Resistant” bacteria used to inoculate subsequent rounds (n=11). Mean ZOI measurements were statistically analyzed with a paired t-test. Tea tree had the largest ZOI of all the extracts. Bergamot and cypress showed no antimicrobial effect on E. coli during the disk diffusion assay. Only 3 extracts had MIC below 5% (v/v). Resistance Assay showed a lot of variability between rounds, but no change in resistance was observed. Commercial plant extracts of tea tree, cypress, bergamot, vetiver, and Eucalyptus radiata show antimicrobial activity and show potential for use in natural antimicrobial products. A universal methodology is needed for this area of research to check the efficacy of antimicrobial activity in some extracts. S. aureus and E. coli did not show any indication of being able to build up a resistance to the tea tree extract. Plant extracts combined with other plant extracts or antibiotics have showed synergistic effects that increased antimicrobial activity (Nanasombat and Wimuttigosol 2011, D’Arrigo et al. 2010). Future experiments can be done to test the synergistic effects of tea tree extract combined with Eucalyptus radiata extract. Introduction Objectives Methods Results Conclusion Future Studies Acknowledgment I’d like to thank Dr. Singleton for all the help and advice he provided throughout this experiment. I’d also like to thank Dr. Kleiner for providing a protocol I could use on the Resistance Assay. Literature Cited