Nt1310 Unit 3 Pathogens

1. Make sure you label both plates on the bottom, with your name, date and name of pathogen you have chosen (listed above under materials) Then label numbers 1-5 on one plate and 6-10 on the other as there are 10 disks that need to be placed. Spread out in a circle formation like pictured below (This is the final product; look at this for reference on how to label)

Now you are ready to start culturing. Start with one of your plates you can do both at the same time but to avoid making any errors I recommend just doing one at a time. Using aseptic technique take the tube containing the pathogen you have chosen and sterilize it. Next, take your sterile pipette (dropper) and pick up a small amount of your sampled pathogen. Using aseptic technique again flame the tube and close it, set aside till you are ready to use for the next plate. Open the lid of the first plate (I used the one labeled 1-5 first)
away for you, remember to always use aseptic technique.
Then drop 3 drops of the sampled pathogen from the pipette into the plate. Close the lid and discard the pipette into the clear bag with the red label on it. Then take one of the sterile spreaders (looks like a hockey stick) open the same plate aseptically and spread the three drops you put in all around the plate. Be careful not the press and damage the broth. After you have spread the pathogen around close the lid and throw away the hockey stick in the biohazard bag. Repeat for the next plate you labeled 6-10. The final step of the first day you will place the disks containing various antibiotics (known concentrations). Aseptic technique is used when picking up each disk and putting it into the plate. Meaning you need to flame the forceps before and after you pick up and put in the disk in while also opening the plates away from you. Each plate that contains the concentrated disks is labeled with a number match that number with the number you have labeled on the bottom of your
plate. I started with number one, I flamed the forceps opened the plate that was labeled 1, picked up the disk and then opened my plate that was labeled 1-5. Place the disk over your label #1 do not press down hard when placing the disk, this may damage the broth. Repeat for each disk through number 10. Your professor will place them to inoculate when you have left the lab. One the second day you will be able to determine if the drug is sensitive or resistant to ten different antibiotics/drugs. Flip plate over where you have labeled to see the zone of inhibition better. Using a metric ruler, measure the zones diameter, make sure you measure were you can see a distinct line on the circle. Do this for all of the zones of inhibition. Make sure you are recording the measurements on a piece of paper as you measure. When you are finished measuring and recorded your data refer to the chart your professor has provided for you. Find the name of your disk located on the disk itself and find the name on the chart. When you have found the name on the chart look for the pathogen that you cultured on the plate if you can’t remember look on the back were you labeled the name! If you have P. vulgaris and E.coli use Entereobacteriacae or Enterococci Refer your measurement of that zone and compare it to what the chart has as a resistant, intermediate or sensitive measurement. Record if it is resistant, intermediate or sensitive. Do this for all of the measurements you have. When done share data with the people at your table.