Thursday, September 12th 2013

*This blog does not reflect Franciscan University in any way*

Class date: Thursday, September 12th 2013

Dear Blog and Blog readers,

Today, we first observed our streak plates!

Previously on "Max and Sam's Adventures in Bio" . . . we placed the streak plate that had been in the 30 degree incubator (red pigment) in the 25 degree incubator and the streak plate that had been in the 25 degree incubator (cream pigment) in the 30 degree incubator. At this point we hypothesized that when they switched incubators they would switch pigments as well. We did this in an attempt to see if this was truly the same bacteria.

Here were our results:

The streak plate that had been in the 30 degree incubator and was placed in the 25 degree incubator has almost no growth, and the growth that did result had cream/no pigmentation. Keep in mind that this is what the same streak plate looked like before: 

Quite a difference!

The streak plate that had been placed in the 25 degree incubator and then transferred to the 30 degree incubator appeared the same in both cases: cream/no pigmentation. 

Our conclusion was that these were two different types of bacteria. The cream/non-pigmented bacteria is able to grow at least between the temperature range of 25 degree Celsius and 30 degree Celsius. However, the red pigmented bacteria is only able to grow at 30 degree Celsius (there may be some variation here- all we were able to determine is that it cannot grow at 25 degrees Celsius and cannot grow at 37 degrees Celsius).

To test this further, we placed the streak plate that went from the 30 degree incubator to the 25 degree incubator and placed in back in the 30 degree incubator to see what would happen. We believed that the red pigment would return, because we still had some sort of hope that this was the same type of bacteria.

                   

We were also given a agar slant test tube with a culture of (different) red bacteria and placed in the the 25 degree incubator and labeled it "S". 

Later, we took a gram stain smear of the bacterium slide from last class.

The purpose of this procedure was to determine whether the bacteria examined was gram positive or gram negative.

First, we prepared a fixed smear of bacteria.

Max sterilizing the loop. 

Max putting distilled water into the sterilized loop. 

Max putting the water on the sterilized glass slide. 

Sterilizing the loop once again. 

Obtaining a pure sample of bacteria

Smearing the bacteria on the water droplet on the slide. 

Allowing the sample to air dry. 

Heat-fixing the sample. 

We covered the smear with crystal violet for  twenty seconds.

Rinsing off the crystal violet with an indirect stream of DI water.

Then we covered the smear with Gram's Iodine for one minute.

We rinsed the slide with indirect DI water. 

Next, we rinsed the slide at a 45 degree angle with 95% ethanol drop by drop until the color stopped running. 

Following that procedure, we rinsed the slide again with DI water,

Next we covered the smear with Gram's Safranin for one minute. 

Once again, we rinsed the slide with DI water.

Finally, we blotted the water from the slide with bibulous paper.

Our slide was now ready to be used under the microscope!

We examined this slide under the oil immersion lense.

Placing the oil on the slide.

Focusing the microscope. 

Our gram-stained red pigmented bacteria from the soil around the Ohio River in Steubenville, OH under the oil immersion lens of the microscope. 

(image taken from Techniques in Microbiology: A Student Handbook by John. M. Lammert) 

*We do not own this image*

We determined that the bacteria was gram-negative bacteria. Gram-negative bacteria is in the shape of rods and has a thinner cell wall than Gram-positive bacteria. Gram-negative bacteria has a Peptidoglycan layer sandwiched between two phospholipid bilayers and has a lipopolysaccharide on it's surface which is an endotoxin. Through this we are able to tell that this bacteria is not denatured by boiling, is antigenic, does not form a toxoid, has a low potency, has a low degree of specificity, has no enzymatic activity, and is pyrogenic (fever-causing).