User:Mary L. Pavia/Notebook/Biology 210 at AU

1.27.2015: Lab 2- Identifying Algae and Protists: Hay Infusion Culture Observations; Preparing and Plating Serial Dilutions (Date of Entry was 1.27.2015, but the date in which we gathered observations and collected a sample from the transect was 1.21.2015)

Purpose: In this lab, our purpose was to examine algae and protists from our transect through observations and the use of a dichotomous key. Our goal was to determine the identity of organisms using the key, and try to identify as many different organisms as possible from different niches within our Hay Infusion Culture, which is even considered an ecosystem (Bentley, 2015). My hypothesis is that there will be few differences between organisms found in different niches within the culture. If we use a dichotomous key to identify organisms from different niches, then we will observe and identify similar organisms in both niches.

Materials & Methods: Firstly, we took visual and olfactory observations of our Hay Infusion Culture that had been sitting undisturbed in an area in the lab for a week. After observing the culture, we took samples for microscopic observation from two different niches: on the very top surface and within the very bottom, mucky layer of the jar. Next, we made wet mount slides from the two samples drawn from the culture. Using the dichotomous key, we identified organisms (protists and algae) by making a series of two morphological choices. The organisms needed to also be measured using an ocular micrometer, and their sizes recorded.

In the second part of lab, we prepared and plated serial dilutions for the next week’s microbiology lab. To do this, we labeled four tubes of 10 mL sterile broth with 10-2, 10-4, 10-6, and 10-8. Using a micropippettor set at 100 microliters and clean tips, we transferred 100μL from the recently gently swirled culture to the 10 mL sterile broth tube labeled 10-2. This created a 1:100 dilution. Next, we swirled the tube, used a clean tip to transfer 100μL from the 10-2 tube to inoculate tube 10-4. This process of serial dilution was repeated two more times to create a 10-6 and 10-8 dilution. We plated the serial dilutions on four nutrient agar plates and four nutrient agar plates plus tetracycline. To do this, each plate was labeled. One of each type of agar plate was labeled 10-3, 10-5, 10-7, and 10-9, and the four plates with tetracycline were also labeled with “tet”. After the plates were labeled, we pipetted 100μL from the 10-2 tube onto the two nutrient agar plates labeled “10-3”. The sample was then spread carefully across the agar. This procedure of plating was repeated with the 10-4 dilution on the 10-5 plates, the 10-6 dilution on the 10-7 plates, and the 10-8 dilution on the 10-9 plates. After each agar plate was prepared, they were set-aside upside down in a rack so they would safely incubate at room temperature for a week.

Data & Observations: The culture smelled moist, rancid, and pungent. As seen in image 1, the culture was semi-transparent, with plant matter floating at the top of the sample. At the bottom of the sample (seen in images 2 and 3), there was a darker muck layer composed of mostly soft, wet soil. There was mold on top of the culture and was growing up the sides of the jar. On a decomposing leaf floating on top of the liquid, white specks of mold grew. There was more mold growing on a lot of the plant matter. The pine needle in the culture was fairly in tact, while the Brussels sprout was shriveled up and smaller in size than when it was initially added to the Hay Infusion Culture.

Image 1

Image 2

Image 3

From the bottom layer niche, organisms observed were Paramecium Aurelia (130 μm, motile), Gonium (colony 90 μm in diameter, non-motile), and Colpidium sp. (60 μm, motile).

From the bottom layer niche, organisms observed were Pelomyxa (2mm, motile, pseudopodia), Paramecium Aurelia (140 μm, motile) and Pandorina (colony 85 μm, non-motile).

All organisms observed are protozoa and are not photosynthesizing organisms.

Image 4 shows the Paramecium Aurelia observed through a microscope from the bottom layer niche

Figure 1 is a dilution table that illustrates the serial dilution procedure used in the lab. The reason the nutrient agar plates are labeled 10-3, 10-5, 10-7, and 10-9 is because the 100 microliters used to inoculate the surface of the agar is considered another ten-fold dilution.

Conclusions & Future Directions: While my data mostly refutes my hypothesis that there will not be drastic differences between the niches, there were some organisms observed in both niches, as expected. Paramecium, for example, was present in the top layer and bottom layer niche.

The organisms might differ close to verses away from the plant matter because the plants could attract various organisms to them as food or shelter. However, other organisms may not need nutrients from the plants or even prefer to live in different environments, such as a less oxygen present area like the one that exists in the bottom layer amidst the wet soil.

If the Hay Infusion Culture “grew” for another two months, I predict that all of the plant matter would decompose, and mold would infest the culture so much that the carrying capacity of the culture could not have space for many other organisms besides the mold. Or, stronger individuals with higher fitness based on the ecosystem will survive and repopulate and inhabit the culture.

References: Bentley, M., Walters-Conte, K., and Nancy K. Zeller. 2015. A Laboratory Manual to Accompany: General Biology II. Department of Biology, American University: Washington D.C.. 22-23.

-MLP

1.25.2015: Lab 1- Biological Life at AU: Observing a Niche at AU (Date of Entry was 1.25.2015, but the date in which we gathered observations and collected a sample from the transect was 1.14.2015)

Purpose: I am analyzing a particular transect (group 4) on the American University main campus to better understand the organisms that inhabit the land, the biotic and abiotic factors that affect the niche, and how all of these factors interact within the different niches in the transect (Bentley, 2015). On the 14th of January, we went to our location to gather observations about it and collected a sample of soil/ground vegetation from the ecosystem that became a Hay Infusion Culture. My hypothesis is that we will observe fewer organisms and factors than are truly present. If we only complete visual observations of the area, then we will record fewer biotic (and abiotic) factors than we will find when carrying out closer microscopic observations of our Hay Infusion Culture.

Materials & Methods: After embarking to the group 4 designated transect and documenting the biotic and abiotic factors observed, we collected a sample of soil and ground vegetation in a sterile 50 mL conical tube. We were sure to collect a diverse group of plant matter so that our sample was representative of our specific transect. After returning to lab, we placed 10-12 grams of the sample into a clean, plastic jar with 500 mLs of Deer Park® water and 0.1 gm of dried milk. Then we put a lid on the jar so that we could carefully mix the contents of it for 10 seconds. Finally, we removed the lid of the jar and placed the open jar in the lab and let it remain there for one week.

Data & Observations:

Figure 1. An Aerial-View Diagram of Group 4’s 20 by 20 meter Transect

Figure 1 is an aerial-view Diagram of my group’s (group 4) 20 by 20 meter ecosystem. The transect was in part of a small farm garden located behind the tennis courts on the western side of the American University main campus. The four smaller rectangles represent the wooden planter boxes in the garden where 4 types of vegetation grow: Brussels sprouts, lettuce, kale and cucumber. The main features in between and outside of the boxes on the ground are woodchips, soil, decomposing plant matter, and some weeds. The compass is provided to indicate navigation of the transect.

Figure 2: Observations about the Biotic and Abiotic Characteristics of Group 4’s Transect

Figure 2 is a table listing the biotic and abiotic factors within the transect in the farm garden observed on Wednesday, January 14, 2015 during the late afternoon lab period.

Image 1 shows the Brussels sprout planter box, as well as the decomposing plant matter on top of the soil within the box. The image also displays the wood chipped pathway in between the boxes within the fenced in farm garden.

Image 2 depicts the kale and cucumber planter boxes. The kale planter box has over 15 sprouted kale plants and an irrigation hose is present within the box. Also, a small scarecrow is lying in the far southwestern corner of the box. In the cucumber planter box, there are no apparent signs of cucumber plant grown above the surface of the soil.

Image 3 is of the lettuce planter box. The northwestern corner of the box is home to one lettuce plant. Snow is inside of the planter box as well.

Image 4 shows the south and southeastern side of the transect, where the 5 shrubs are present. Plant matter surrounds each individual plant.

Conclusions & Future Directions: As previously noted within the Data & Observations section, the transect was within a fenced off area with a gated entryway. This could be important, as it possibly reduces foot traffic of humans and animals, limits quantity of pests in the garden, and contamination of area in general. The animal waste found within the ecosystem indicates that, while no animals were seen at that time, there is clear evidence of animal interaction with transect (ie. Rabbit). Lastly, the small shrubs near the southeastern side of the transect against the fence were surrounded by decomposing plant matter. This looked intentional and was probably laid down for insulation of the plants during the cold winter months. According to the BBC, cold weather can have damaging effects on the cellular make-up of plants. Because of this, layering plant matter around the bases of plants can minimize the damage of cold temperatures (“Cold weather”, 2014).

My hypothesis was that we will observe fewer organisms and factors than are truly present. I predicted that if we only complete visual observations of the area, then we will record fewer biotic (and abiotic) factors than we will find when carrying out closer microscopic observations of our Hay Infusion Culture. While the hypothesis did not truly address the original purpose of the lab, the hypothesis is necessary and will still be applicable for the second part of the lab. Next lab period, we will use the Hay Infusion Culture created in this lab to test my hypothesis and attempt to identify different organisms and protists in the sample by using a dichotomous key. We will also observe the cultures after the week incubation period and find different niches within the one culture. To finish lab, we will prepare and plate serial dilutions to prepare for the next week’s microbiology lab.

References: Bentley, M., Walters-Conte, K., and Nancy K. Zeller. 2015. A Laboratory Manual to Accompany: General Biology II. Department of Biology, American University: Washington D.C.. 16.

“Cold weather”. Gardening guides. 2014. BBC. (25 Jan 2015) <http://www.bbc.co.uk/gardening/basics/weather_coldweather.shtml>

-MLP

1.20.2015: I'm am able to access OpenWetWare and submit content for Biology 210.

-MLP