Unknown Lab Report

3. You test another new unknown bacterial sample, and find the G+C content is identical to one of the samples you have already identified, but the rRNA gene sequence contains one base that is different. What can you conclude: C.

A sample was taken, examination under the microscope did reveal some clue cells. An Affirm prep is pending.

An unknown test tubes labeled P and 11 was given out by the lab instructor. The goal was to determine two separate unknown bacteria, one gram- positive and one gram-negative. This technique was used by gram staining. The purpose of gram staining is to determine whether bacteria are present. Also, the structure of the organism’s cell wall determines whether the organism is gram positive or negative. My unknown test tube P was gram positive because the bacteria retained the primary stain, while my unknown test tube 11 was gram negative because it was decolorized and counterstained. Test tube P had small cocci’s in clusters and chains, while test tube 11 had small rods in clusters.

A Simmon’s citrate slant is needed for this test. This slant is a chemically defined medium of sodium citrate as a the carbon source, and the pH indicator bromothymol blue. Only microorganisms containing the enzyme citrate premease, that allow them to use citrate as the only source of carbon, will survive. They will produce alkaline byproducts which would turn the medium from green to blue. Using a sterilized inoculation needle, I collected the sample from Unknown 1B, then stabbed the needle all the way to the bottom of the tube. Then I gently pulled out the needle to the surface of the slant and streaked the surface before removing the needle from the tube.

The experiment was completed by first putting on safety goggles and an apron, then get a sharpie and draw a cross on the top of the petri dish that has agar in it, then label each quadrant with one control group. After that has been completed the partner will need to clamshell open the petri dish while the cotton swab dipped in Micrococcus luteus is being spread side to side from the top of the agar to the bottom. When that is completed the petri dish is going to need to be turned ninety degrees and the same procedure will need to be performed. Once the bacteria is spread across the agar it will need to be taken to the counter with six household products on it and put one household product in the first quadrant, another in the second quadrant,

This procedure describes the use of a mixed broth culture, where the culture contains many different bacterial species. The specimen streaked on a plate could come in a several of forms, such as solid or liquid samples and cotton or foam swabs. Material containing possibly infectious agents should be handled in the lab using proper bio safety procedure. Remove the test tube cap; Insert the loop into the culture tube and remove a loopful of broth. Replace the cap of the test tube and place it back into the test tube rack. The lid of the agar plate has to be opened just enough to streak the plate with the inoculation loop. Be mindful of the amount of agar and the length of time the agar is exposed to the environment during the streaking process. Also avoid talking over the plates, this keeps the plate from becoming contaminated. Loosen the cap of the bottle containing the inoculum. Hold an inoculation loop in your right hand and flame the loop and allow it to cool. Lift the test tube containing the inoculum with your left hand. Dip the loop into the broth and lightly touch a colony with the loop. Sightly lift the lid of the Petri dish containing the solid medium. Place a loopful of the culture on the agar surface. Flame the loop and cool it for 5 seconds by touching an unused part of the agar surface close to the ouster partition of the plate, and then drag it rapidly several times across the surface of area1. Remove the loop and close the Petri dish. Reflame and cool the loop, and turn the petri dish 90° then touch the loop to a corner of the culture in area1 and drag it several times across the agar in area 2, hitting the original streak only a few times. The loop should never enter into area 1 again. Remove the loop and close the Petri dish. Reflame and cool the loop and again turn the dish 90° anticlockwise. streak area 3 in the same way as area 2, hitting last area several times. Remove the

Figure 4: The 1/10 dilution is 10mL of Sewage water mixed in with 900mL of Turtle Creek water. The curves clearly show the closeness between positive control and DNA sample. Each sample has two repeats. Turquois is positive control, Red is the 1/10, and Green is negative

To obtain some organisms, I took four samples from four different environments and put them on a petri plate. The petri plate was then put in the incubator at 37°C for forty eight hours (Lammert, 2007). Out of four, samples A and D grew nothing and sample B had some growth but not much. As a result I had to choose two unknown organism, that were possibly different, from the third quadrant. This quadrant, which I refer to as sample C, was taken using a wet swab on the border of my right shoe. From a spot where the white border had an orange tint. In that quadrant, specimen Ca. came from a lone circular spot near the rim whereas specimen Cb. came from a fibrous colony near the tip of the quadrant.

I think that Pencil #2’s (Garlic was placed on specimen before bacteria started to grow) will show fewer bacteria growth than the other specimens.

The results for lab number 1 and 2 where that’s at the Agar in the plate shifted to the left side of the Agar plate. As a result, it was a failure. This was an indication to start the experiment portion of lab number 2 over again. The results for lab number 3 was successful. Look at figure number 1.0 on page 17. The plaques are distributed nicely throughout the Agar plates. The plaques are also opaque, large, and without any white spots known as contaminates.

Gram-positive can sometimes appear Gram-negative when Gram-positive lose their ability to keep their crystal violet when they are old and also when they are incubated longer. Another reason may occur when a lot of alcohol were used on Gram-positive organisms, and it would result in loss of crystal violet stain. The purpose for using a control was to be sure the known gram-negative and positive organisms stain correctly. If they were not correctly dye, the result would be incorrect. I improved on the mistake I made when I was doing the simple stain. The only area I think need to improve on would be to remember that I have to put the slide in focus in order to see the organism clearly. My unknown was Gram-positive and the shape was rod. This was different than the shape and the grouping that I saw on the simple stain experiment. My unknown was Bacillus subtilis. I excluded other organisms because E. coli, staphylococcus, streptococcus, and micrococcus all of them grew. I came up with this conclusion because my unknown never seem to show growth, even the media type lab that I did had no

The bacterium was then taken from the test tube. The loop and the test tube top were heated before and after the bacteria removed. One quadrant of the agar plate was streaked. The plater was closed, and the loop as flamed. The plate was rotated at 90° and the loop was cooled in an uninoculated area of the agar.

At the end of the experiment, the unknown substance was identified as stearic acid, which is found in animal fats. However, in order to reach my final conclusion each step in the experiment was to be done correctly and error free. For instance, when it came to solubility the substances that were mixed in could result being insoluble, soluble, or slightly soluble. The next procedural was density. My unknown was solid was the steps were to use a graduated cylinder and measure the initial and final volume and divide that by the mass; but we had to be careful when putting the water and the unknown together in the graduate cylinder. It could have resulted in air bubbles one of many errors that can occur in an experiment. If there was an error in

This is due to the unknown #12 being a gram negative bacteria with rod-shaped cells, etc. The gram staining gave more information on the cell arrangement as it being a single bacillus. Plate streaking at the NA plate was next and in so the microbes pigment and colony morphology was displayed as translucent to white creamy pigment and round convex. The streaking of the EMB, another plate gave more info as it displays a light purple, in so interpreting the organism is not inhibited by eosin and methylene blue. The IMVIC test is one the most recommended in ways that it differentiate gram negative microbes. It has four tests within it, but this test only focused on two. The outcome of the test was inaccurate in ways that the Methyl red test didn’t give any accurate result while the VP test remained accurate. In so moved to the final

A patient has come into the hospital with signs of a fever, vomiting, diarrhea, and fatigue. The doctor then takes a sample from the patient because he needs to know if there is a pathogenic species causing these signs in the patient. The doctor then gives me, the lab technician the sample he acquired from the patient. It is now my job to isolate the bacteria species and to determine if the species is pathogenic causing the patient to be sick. The first thing I am going to do is make sure all of my lab equipment is sterilized to make sure no outside bacteria are going to affect the results.