An Ethnobotanical Approach to Finding An2microbial Compounds in Wooly Blue Curls (Trichostema lanatum) Using a Kirby- Bauer Disc Diffusion Assay
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1 An Ethnobotanical Approach to Finding Anmicrobial Compounds in Wooly Blue Curls (Trichostema lanatum) Using a Kirby- Bauer Disc Diffusion Assay By: MaDhew C. Fleming and P. MaDhew Joyner
2 Ethnobotany is the scienfic study of characterizing why cultures use plants the way they do. hdp://en.m.wikipedia.org/wiki/chumash_(tribe) hdp:// monica- mountains
3 Goals of my project. Determine whether a wooly blue curls extract exhibits anmicrobial characteriscs.. Determine what compound in the wooly blue curls extract works as an anmicrobial. 3. Determine how these anmicrobials effect different bacteria.
4 Preparaon of WBC extract Soak in MeOH for - 4 hrs. Re- dissolve in DMSO hdp://cen.acs.org/arcles/9/ web/3/6/nontoxic- Solvents- Extract- Compounds- Plants.html Strain out solids Evaporate solvent with a Rotevap.
5 The technique used to determine anmicrobial acvity was the Kirbey- Bauer disc diffusion assay. Swab onto agar Check Density hdp:// world.com/producnfo/ 3_43_87_683/49/M- Medium- x- sterile- liquid.html hdp://openwetware.org/wiki/ Luckau_Protocols:NanoDrop Incubate at 37 C for 8hrs. hdp:// Grainger/incubators/lab- ovens- heang- and- refrigeraon/lab- supplies/ecatalog/n- ktb Inoculate discs with ul of extract Introduce discs
6 Ager incubaon, I quanfied my results by measuring the diameters of inhibion. negave control ampicillin (. mg) wooly blue curl extract
7 Diameter of All Gram + bacteria had a response to the wooly blue curls extract, but there was some variaon among responses. Staphylococcus epidermidis Diameter of Staphylococcus aureus Corynebacterium xerosis Diameter of.5 = No Inhibion NT= Not Tested
8 Diameter of 3.5 Though some Gram + bacteria varied in response, others reacted very similarly. Bacillus sub8lis Diameter of 3.5 Bacillus megaterium Enterococcus faecalis Diameter of 3.5 = No Inhibion
9 The E.coli ΔtolC strain, which does not produce the tolc drug efflux pump, was suscepble to the wooly blue curls extract! Escherichia coli WT Diameter of.5mg mg.mg.mg ampicillin DMSO Escherichia coli ΔtolC Diameter of = No Inhibion.5mg mg.mg.mg ampicillin DMSO
10 .5 Gram- posive bacteria (blue) show greater sensivity to growth inhibion than gram- negave bacteria (orange). Diameter of No inhibi on No inhibi on
11 All the Gram + bacteria showed posive results, whereas the Gram had no response regardless of the niche they most commonly occupy. Bacterium Niche Gram (+/- ) Diameter of Staphylococcus epidermidis Skin Staphylococcus aureus Skin Corynebacterium xerosis Skin Enterococcus faecalis Gut Bacillus sub<lis Gut Bacillus megaterium Env Salmonella typhimurium Gut Escherichia coli WT Gut Escherichia coli ΔtolC Gut
12 Conclusions of goals. The Chumash medicinal plant wooly blue curls (Trichostema lanatum) does exhibit anbacterial properes.. Gram - and Gram + bacteria show different suscepbility to growth inhibion by wooly blue curls extract. 3. Resistance seen of Gram - bacteria may be due to drug efflux pumps and an extra outer membrane. *We were unable to determine the compound working as an anmicrobial.
13 Further Study We would like to fraconate the extract to determine the acve compound and test how effecve it actually is. Test for synergisc properes of compounds within the wooly blue curls extract 6. Screen other Chumash plants for efflux pump Anagalis Arvensis Scarlet Pimpernel inhibitors. hdps://upload.wikimedia.org/wikipedia/ commons/b/b8/anagallis_arvensis_.jpg
14 Acknowledgments This research was funded by the Naonal Science Foundaon, Research Experience for Undergraduates, REU- Site Grant, #DBI- 67 and the Natural Science Division of Pepperdine. We would also like to thank Victoria Hester for her help with plant collecon and extract preparaon.
15 References. Tegos, G., Stermitz, F. R., Lomovskaya, O., and Lewis, K. An<microb. Agents Chemother., 46 (), Timbrook, J. Chumash Ethnobotany: Plant Knowledge Among the Chumash People of Southern California. 7, Heyday Books. 3. The Human Microbiome Project Consorum. Nature, 486, Cox, P. A. Science 994, 87, Davis, W. W. and Stout, T. R. Applied Microbiology 97, (4), Stavri, M., Piddock, L. J. V. & Gibbons. J. An<microb. Chemother. 7, 59, 47 6.
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