Broth Tubes, Streak Plates, Stains and More!

Today was a fun-filled day in lab!  We accomplished so many things, executing both new and old techniques.

First, we made broth tubes of both our environmental sample and our unknown sample.  To do this, we used aseptic technique.  To reiterate, this consists of sterilizing the inoculating loop between each step and sterilizing the tubes containing the broth and the sample.

 

Our bacteria will grow in the broth for next class. We placed both samples  in the proper incubator after marking them with a China Marker. (The unknown at 37 degrees Celsius and the environmental sample at room temperature.)

After this, we made a streak plate of "F," our unknown.  To do this we followed the same procedure found in "Examination of Bacteria and Creating a Pure Culture."  The purpose for this was to create a pure culture of the bacteria to further examine the details of the colony and morphology of the unknown specimen.

This is "F." We placed this sample in the 37 degree incubator as well so it can grow for our next class.

Also, we repeated the procedure to make a simple stain on a microscope slide for both samples (the unknown and the environmental sample).  The charge of the stain is very important!  There are two types of stains used in a simple staining procedure.  The first is Acidic, or anionic, and it is used to bind to proteins with a positive charge.  However, this stain won't work with bacteria because the bacteria's negative charge would repel the stain.  When staining bacteria, it is important to use a basic stain, or cationic stain, with a positive nature so the stain will reveal the mysteries of the bacteria!

We used methylene blue stain on both of the samples.  The procedure for creating a simple stain can be found in our previous blog, "Staining Bacteria: Preparing a Smear and Preparing an Unknown Sample."  However, to reiterate, "heat-fixing" the bacteria is very important.  To receive better results this time, we waited for the smear to dry thoroughly, and heat-fixed it for three quick motions.  We let the stain rest on our sample for one minute, and then we wiped it dry.  This slide reveals our wonderful bacteria!  The results will have to wait! We stored the slides in the drawer until next class.

Now it's time to dive into a new procedure, literally!  This is a new test that we find really exciting. We are now going to test the motility of our bacteria!  To do this, we used a sterilized inoculating needle to capture a small amount of bacteria.

After capturing the small amount of bacteria, we stuck the inoculating needle into a tube of 0.4% agar (this means it is semi-solid).  It was important that we stuck the needle straight down three-quarters of the way and straight back up the same hole.  After retrieving it from the incubator next class, there should be cloudiness.  Cloudiness is a result of bacterial growth.  If the cloudiness is about the whole tube, our bacteria is considered motile!  We did this for both the unknown and environmental sample.

After, we got a throat swab from Dr. Joesph!  He made a culture of his bacteria on an agar plate to grow for next class, but added something very unique to it, TWO ANTIBIOTICS!  The reason he used two, Bactitracin and Penicillin, was to discover if he had strep throat (caused by Streptococcus pyogenes).  He placed them on opposite ends of the agar plate.  Next class, by examining the growth on the plate, we will be able to diagnose our professor.  If the Penicillin kills bacteria, it doesn't necessarily mean that he as Strep.  It only means that he has bacteria in his throat, but that's a good thing, WE ALL DO!  However, if Bactitracin kills bacteria, he does have Strep and should go to the doctor because this antibiotic only kills the bacteria that causes Strep.

Next, we took a new unknown bacteria and made a pure culture of it on an agar plate.  However, to this sample we added a T-4 bacteriophage (a virus that kills bacteria).

We will not know the exact results until retrieve the dish next class. However, as a class, we hypothesized that there will be clear pockets on the plate where the T-4 bacteriophage was added to the agar.  A bacteriophage kills bacteria, so none will be present there.

So many exciting results are awaiting next class! Can our bacteria swim around?  What will our broth samples reveal?  DOES DR. JOSEPH HAVE STREP? Tune in next time!