Gram-positive agar formulation

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Authors
Lemen, Laura
Advisor
McKillip, John
Issue Date
2023-05
Keyword
Degree
Thesis (B.?)
Department
Honors College
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Abstract

Selective media which allows for the growth of specific bacteria while inhibiting the growth of others is an important tool used daily by microbiologists. There exists excellent media for the selective growth of Gram-negative bacteria, yet there are minimal options for the selective growth of Gram-positive bacteria. While there are Gram-positive selective media, they may contain antibiotics, be species specific, or allow the growth of some Gram-negative organisms. This poses an issue to students in microbiology classes who may struggle to isolate Gram-positive bacteria from a mixed or contaminated sample potentially due to improper streaking techniques. An effective Gram-positive selective media would also be beneficial in the research or clinical setting where it could improve the efficiency of isolation. The aim of this study was to create a new formulation for Gram-positive selective agar (GPA) which does not contain antibiotics and allows for the growth of most Gram-positive bacteria while inhibiting the growth of most Gram-negatives. To do this, a formulation previously created by McKillip and Drake was used as an initial guideline. This GPA utilized 2-phenylethanol as the main selective ingredient due to its documented inhibition of Gram-negative growth. The formulation was mixed, and plates were inoculated with different species of Gram-positive or Gram-negative bacteria. The plates were statically incubated at 37℃ for 96 hours and were checked every 24 hours for growth. Based on the presence or absence of growth, the components and their concentrations were adjusted to address undesired results. This process was repeated for different GPA formulations. The study was ceased before optimal results were achieved due to time constraints. The final formulation contained 33.6 g/L sodium chloride, 15 g/L sodium glycerophosphate, 12 g/L lithium chloride, 0.31% 2-phenylethanol, 15 g/L glucose, 10 g/L tryptone, 10 g/L yeast extract, 19 g/L agar, and 5% sheep’s blood. While a perfect formula has not yet been achieved, considerable progress has been made. Further work on GPA is required to create a media which addresses the needs of scientists in a suitable manner.