Read section II of chapter 26 in your lecture textbook for background on chemical antimicrobial control (for background)—pages 762-763.
Today you will perform a frequently used procedure called the Kirby-Bauer Antimicrobial Susceptibility test (disc diffusion technique). Each group will inoculate his/her own plate of Mueller-Hinton agar with an assigned culture. To that inoculated plate, you will then aseptically add sterile filter paper discs (using a disc dispenser), which contain a known concentration of antibiotics. As soon as the antibiotic discs touch the agar, the antibiotic will begin to diffuse into the surrounding agar. During incubation the bacteria you inoculated onto the agar will begin to grow. As the antibiotic diffuses, and the bacteria grow, organisms which are able to be inhibited by the antibiotic will fail to grow where the antibiotic is highest in concentration. This will leave a clear area, called the Zone of Inhibition, around the discs. During the next lab period, you will observe your plates and measure the diameter of the zone of inhibition, and use a standard interpretation table to interpret your results. Organisms may be susceptible, resistant or intermediate in resistance to the antibiotic. After you obtain results from your own plate, you will join the rest of your class to pool your data and discuss the results.
Day one. - Materials and Protocol
1. Your instructor will assign you a number. Look at table 1 for your assigned culture. Record the culture you have selected here: ______Group 1______________
1 | S. aureus (SA)|
2| E. coli (EC)|
3| P. aeruginosa (PA)|
2. Your group should obtain the following materials before you can proceed: * A container of sterile cotton swabs
* A Petri dish containing Mueller-Hinton agar (one per group; labeled MH)...