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Revision as of 12:04, 3 February 2014
Antibiotics are molecules that kill bacteria or inhibit their growth. Common antibiotics target important cellular functions in bacteria (such as cell wall remodeling or protein synthesis). Believe it or not, many microbes are naturally resistant to antibiotics, either because they produce them or because that resistance is advantageous in the habitat they occupy. One place in which such selection is currently taking place, is in the human microbiome. Unfortunately, due to the overuse of antibiotics (both in medicine and agriculture), widespread antibiotic resistance has become the norm. The figure below captures the trend and shows the increasing prevalence of antibiotic resistance in human pathogens.
Today you will be testing whether or not your honey bee isolates are resistant to antibiotics. Honey bees are agricultural organisms and are treated with the antibiotic tetracycline prophylactically, twice a year, by many bee keepers. Because of this practice, the honey bee microbiome has tetracycline resistance determinants. For a nice paper on these determinants, check this link out 
You will be using a soft agar overlay technique to test the degree of resistance in your bacterial isolates. First, you will take a cellulose disk, impregnated with antibiotics, and put it in the center of your agar plate. You will then take 2 mL of molten agar (45C water bath) and add 75 uL of an overnight culture of your isolate to that 2mL *very quickly* by inversion two times. Pour your soft agar over the surface of your agar plate, nutating the plate in order to spread the soft agar evenly. Don't take too long or the agar will solidify and you will not have time to pour it!
Set up an overnight culture the day before!