User:Madison Ballacchino/Notebook/Biology 210 at AU

Description of Transect 1/13/16 The purpose of this lab is to observe and record all different kinds of life and non life that is surrounds the environment, by describing all different aspects of life and non life in a 20x20 transect. My group's transect was located right outside of Hurst's side doors and immediately to the right, next to Ward. The direction was about 30 degrees NE. In the transect, the soil was hard with sparse patches of grass and crinkled leaves were scattered throughout. Grass was found towards the southern direction of the popsicles when looking from an aerial view. There were acorns that were laying there, along with twigs branches. Brown tree shrubs with no leaves, only branches, left were more towards the north direction at the top. If any grass was found, it was near the outsides, towards the popsicle sticks, or surrounding the roots of the branches of the shrubs.There was cement behind the area with a sewer entry leading to a building, and that is where the popsicles cut off. The abiotic things we found in this transect were candy wrappers, tree branches, fallen leaves, cement, and a sewer grate. The biotic things found were short grass, tall grass, clovers, shrubs, and acorns.

M.B.

Protists and Algae 1/27/16 Hay Infusion setup and initial observations: Originally, the Hay Infusion smelled like musty, moldy mud. There was a thin layer of mold at the top. There was a layer of soil at the bottom and water in the middle with leaves and grass floating at the top. If this Hay Infusion continued to grow for another two months, then there would be a thicker film of mold at the top with even more bacteria growing. Different niches would probably develop as bacteria can grow fast and they would create different roles for the varying bacteria. As it grew, it would probably smell worse because the bacteria secrete a gross smell. One organism we identified was Eudorina. It meets all the needs of life because it obtains energy, grows, reproduces, and is made up of cells.

Indication of where protists samples came from in the Hay Infusion: The first and second protist found came from a niche right above the bottom layer of the Hay Infusion. This is where the dirt sank to the bottom and combined with the water to make a muddy/sand-like area. The second protist also came from this area. The third protist was found in an area at the top of the Hay Infusion. It was taking going through the thin film of mold at the top, near a dead leaf. This changed the options of what protists were found because of the different food sources and living environment that was available.

Description of the protists samples from the Hay infusion: The first protist found was an organism that was irregularly-shaped and had one big circle near the top, along with small little circles found throughout the "body" of the organism. It was clear and appeared to be non-motile. In this sample, this was the only organism found for this niche. It was ten micrometers at 40x.

For the top layer niche, two protists were found. The first one was thought to be Eudorina. It was a large purple circle with little green circles on the inside of it. It also appeared to not be moving. It was difficult to classify as it did not match up with anything on the dichotomous key. It was 12 micrometers at 40x.

The second protist found was clear and moved fast. It had no visible flagellum. The size of it was 3 micrometers at 40x. It was difficult to classify as it did not match up with anything on the dichotomous key.

A serial dilution was then set up for the next lab meeting. The serial dilution was made of agar plates, half with tetracycline and half without. Different dilutions of the Hay Infusion were added to these agar plates.

M.B.

Microbiology 2/3/16

Now the Hay Infusion's smell was way more potent. The layer of mold at top was now more like a opaque film of mold that was more thick. The hypothesis for the change in appearance and smell is that more bacteria are growing and grouping together. This then releases a stronger smell.

The results from Table one showed that the dilution of 10^-3 and 10^-5 plates with nutrient and tet were the only ones to grow colonies. This indicates that the bacteria was resistant to the bacteria for these two sets of dilution plates. 10^-3 had 112000 colonies/mL and 10^-5 had 400000 colonies/mL. They can survive more easily in 10^-3 because there were more of them, making it easier for them to spread out to find their own food, instead of having to group together. The tetracycline killed off the bacteria more easily when there weren't as many bacteria per mL.

Mechanisms for tetracycline: The mechanism of action for tetracycline is to invade a cell of bacteria and disturb the process of protein synthesis or they are destructive to the membrane. Bacteria that is sensitive to this tetracycline are gram-positive and gram-negative bacteria, chlamydiae, mycoplasmas, rickettsiae, and protozoan parasites. Chopra, I., & Roberts, M. (2001). Tetracycline Antibiotics: Mode of Action, Applications, Molecular Biology, and Epidemiology of Bacterial Resistance. Microbiology and Molecular Biology Reviews, 65(2), 232–260. http://doi.org/10.1128/MMBR.65.2.232-260.2001 Schanppinger D, Hillen W. Tetracyclines: antibiotic action, uptake, and resistance mechanisms. Arch Microbiol 1996; 165:359.

Characterization Table:

Materials and Methods for gram stain: Growth on the agar plate was put onto a wet mount slide and held over heat to dry. The bacterial smear was covered with crystal violet for a minute and rinsed with water. Then iodine was added on for one minute and the rinsed off. Then 95% alcohol was added for 20 seconds and rinsed. Safranin stain was added for 30 seconds and rinsed off after. The mount was then air dried for microscope observations.

Materials and Methods for PCR amplification: 25 microliters of primer/water were added to the two separate PCR tubes and shaken around. Bacteria from the agar plate with tetracycline with 10^-3 dilution was added to a PCR tube. Bacteria from the agar plate without tetracycline with a 10^-5 dilution was added to a second tube.

Colony Label: 10^-3(with tet): It was pure, 10 micrometers on 4x magnification, irregular shaped, and wrinkled. It did not move and is shaped like an actual human heart. It is gram positive. Identification is Blepharisma.

At 40x.

10^-5(with tet): It was purple, rod shaped, irregular, over 100 micrometers on 4x, and wrinkled. It did not move, and the surface was raised. It is gram positive. Identification is Blepharisma.

 At 4x.

10^-3 (no tet): It was purple, rough, linear, and over 100 micrometers on 4x. It did not move and you could not zoom in close enough to see the cell shape.Identification is Stentor.

At 4x.

10^-5 (no tet): It was filamentous, purple, and wrinkled and was 50 micrometers at 10x. It did not move and was chromosomal looking. Identification is Stentor.

At 40x.

M.B.