User talk:Jordan Lenkin
July 14, 2015 Invertebrates and Vertebrates Intro
In continuing the study of levels of biodiversity present in the transect, the next organisms that were observed were invertebrates, followed by a brief examination of the vertebrates present in and around the transect. Despite their size, invertebrates are a highly diverse and complex phyla of organisms. It is believed that the more simplistic systems present in invertebrates eventually evolved into the complex organs systems of vertebrates today. Using the Berlese Funnel system set up during the previous lab, a number of invertebrates were identified and examined in detail.
The Berlese Funnel was broken down by pouring 10-15 mL of liquid and organisms each into two petri dishes. Invertebrates were separated from the debris present using a combination of observations under the microscope and with the naked eye. Each organism found was identified using a dichotomous key. Upon completing the study of invertebrates, a final trip to the transect was made to observe and identify larger animals in order to complete a food web.
The table below details the specifics of the four invertebrates that were observed once the Berlese Funnel was broken down.
- It should be noted that many bees were observed in the transect during the various visits, however there were none in the sample collected for the Berlese Funnel.
As Table 1 demonstrates, the invertebrates collected ranged in size from about seven millimeters to approximately 15 with the bees being the largest and the beetle and pill bugs being the smallest. Because no bees were collected, there is no count for the number present in the sample. There were two pill bugs and one of each of the other invertebrates (see images below). Additionally, it should be noted that while instructions called for identification of five invertebrates, only three were present in the Berlese Funnel.
Image 1: Pill Bug
Image 2: Cedar Beetle
Image 3: Earwig
Table 2 includes a list of five vertebrates that might inhabit the transect.
Of the five vertebrates listed above, three are species that have been observed with frequency on American University’s campus and that are likely to inhabit an area akin to the transect chosen for this study. Two of the species (the house sparrow and the grey squirrel) were actually present in the transect during observations.
Before considering how each of the observed species present in the transect are interconnected, it is important to consider how the general abiotic and biotic characteristics may benefit each of these organisms. As previously listed, the abiotic features present in the transect include soil, rocks, sprinklers, but also rainfall, sunlight, and dead organisms. Each of these provides a benefit in some way or another. For example, the soil provides a habitat for many of the microorganisms as well at some of the invertebrates. The water, whether it is coming from the sprinklers or from rainfall, promotes vegetative growth, which in turn promotes habitation by a range of the organisms observed. And the sunlight promotes photosynthesis in the plant life present, which is a food source for many, if not most, of the other organisms present in the transect. Diagram 1 below is a food web based on all of the organisms observed in the transect. As noted in the Results section, only four invertebrates were identified from the transect and included in the food web. This may be an indication that the transect is lacking in high levels of diversity.
Diagram 1: Transect Food Web
As the diagram demonstrates, although not especially diverse, there is a functioning community of organisms present within the transect. Beginning at the microbiotic level and following through to the vertebral level, the organisms present interact with each other in this common location. For example, the sparrows feed on the pill bugs that feed on the dead plant matter. If there were no plants to produce the dead plant matter, the pill bugs would not have a food source. Although the sparrows feed on a variety of other invertebrates, pill bugs seemed to be the invertebrates with the greatest population, so the sparrows may go elsewhere to find a more abundant food source if the population of pill bugs came into rapid decline. Additionally, the sparrows represent a good example of the carrying capacity within the transect. Of all the vertebrates observed, the sparrows appeared to have the largest population. This would suggest that the transect provides a proper habitat, reliable food sources, and enough water to sustain the population. However, in order to determine if the sparrow population is actually at it’s carrying capacity or if it is simply in equilibrium, further tests and observations would have to be carried out. JL
July 9, 2015 Identifying and Studying Plants
In the study of land plants it is generally agreed upon that evolution began with aquatic green algae. Initially these plants were thought to resemble modern Bryophytes, and eventually they became the plethora of species present today. As an evolutionary step, modern Bryophytes provide key insight into several developments plants took on when they moved from an aquatic environment to a dry one. These developments include: adaptation to fluctuating temperatures on land, efficient access to water, access to gasses (such as CO2) and other nutrients, and reproductive interactions. By observing plants from the transects, a small survey of land plants was conducted in order to identify the aforementioned key evolutionary features that have allowed plants to succeed as they are today.
Before looking for specific plants to sample, a gallon sized Ziploc bag was filled with leaf litter (about 500 grams) to be used for the Berlese funnel. Samples from five different plants were taken from the transect. The location of each plant was detailed, and photographs were taken. The samples and the leaf litter were returned to lab for identification. The Berlese funnel was set up by pouring 25 mL of a 50:50 ethanol/water solution into a tube. The tube was taped to the bottom of a funnel, which had a piece of screening material fit into the bottom. The leaf litter was placed into the funnel. The funnel and ethanol filled tube were set up on a ring stand underneath a 40-watt lamp. All of this was covered with foil and allowed to sit. No seeds were collected.
The table below details the specifics of the five plant samples taken from the transect.
As Table 1 demonstrates, all but one of the samples were dicots, they all shared similar specialized structures such as stomata and waxy leaves, and they all reproduced using pollen. Most of the plants were also perennial, flowering plants. The diagram below shows the location where each plant sample was taken from.
The images below show the plants as they were found in the transect.
Image 1: Broad leafed plantain
Image 2: White clover
Image 3: Japanese pagoda tree
Image 4: Flat weed
Image 5: Crabgrass
Although only two of the plants had flowers, all but the crabgrass are known to flower at certain times of year. Thus, their reproductive methods were identified. All of the plants possessed waxy leaves, or a cuticle, which indicates they all have stomata and guard cells to control water and gas exchange. Finally, while the root structures were not unearthed, it is safe to assume that they all have efficient arrangements that allow for nutrient uptake. In taking a close look at five land plants, not only were shared characteristics identified, a range of diversity was also taken into account. Because the transect in question is relatively homogenous, it was not surprising that many of the plants had similar vascularization, specialized structures, and methods of reproduction. It is also not surprising that many of them were classified as weeds. JL
July 7, 2015 Identifying and Studying Bacteria
In keeping with the study of prokaryotic, unicellular organisms, the focus of this lab was to observe members of the Domain Bacteria, to look at their morphological characteristics, and to observe naturally occurring antibiotic resistance in bacteria. In addition to bacteria, organisms from the Domain Archaea are also prokaryotic. However, these organisms tend to grow in extreme environments such as in Yellowstone National Park. It is not likely that there will be any organisms from Archaea present in this study. The bacteria that will be studied during this lab were cultured from the hay infusion using a series of serial dilutions that were plated on either an antibiotic positive or an antibiotic negative nutrient agar plate. The hay infusion was also allowed to continue its own culturing process. It may be hypothesized that the hay infusion will change in appearance and smell as time goes one and more microorganisms are allowed to grow.
Two serial dilutions were prepared and plated. One set was plated on nutrient agar containing tetracycline. One was plated on nutrient agar with no antibiotic. The dilutions were incubated at room temperature for five days. After the incubation period, the plates were observed and dilution results were recorded. Further analysis was conducted by isolating four bacterial samples from the 10-9 dilution (two from the tetracycline positive plates and two from the tetracycline negative plates). Wet mount and gram staining procedures were conducted on each sample. Set up for a PCR 16S sequence was then prepared by transferring a single colony of bacteria into 25μL of master mix with 0.5 μL of primer. The primer-mix solution was then placed in a water bath.
Before beginning observations of the bacteria, it was noted that the hay infusion culture was not as murky as it had been previously. There seemed to be more growth on the top, and there was still a thick layer of sediment at the bottom. It had also taken on a stronger odor of rust and sewer. The two tables below detail the results from the observations of the nutrient agar plates as well as the gram staining procedure.
Table 1 details the observations from the serial dilution of the hay infusion. The dilutions were plated on either a nutrient agar plate that also had the antibiotic tetracycline, or on a nutrient agar plate that did not have the antibiotic. There was more consistent growth on the tetracycline negative plates, resulting in bacterial lawns. The lawns were also primarily white in color with sporadic yellow colonies. The growth on the tetracycline positive plates was still high, but less so than on the tetracycline negative plates. The colonies were primarily yellow in color, although there were a handful of white colonies. Additionally, it should be noted that the number of colonies decreased as the solutions became more dilute.
Image 1:Serial Dilutions
Image one shows the cultured serial dilutions.
Table 2 is a record of the descriptions of the cell and colony morphologies from each wet mount created. All cells were non-motile and had a spherical shape. The cells collected from the white colonies were gram negative, brownish in color, and their arrangements were not very dense (see Images 3 and 4). The cells collected from the yellow colonies were gram positive, pinkish in color (from the stain), they were smaller than the cells from the white colonies, and their arrangements were dense (see Images 2 and 5).
Image 2: Slide 1 (tet+, yellow)
Image 3: Slide 2 (tet+, white)
Image 4: Slide 3 (tet-, white)
Image 5: Slide 4 (tet+, yellow)
It should be noted that the Image of Slide 4 does not appear very pink. This is due to the image and not the actual coloring that was present under the microscope.
The procedures conducted during this lab allowed for observations of the diversity of morphological characteristics of bacteria, as well observations of naturally occurring antibiotic resistance. It should also be noted that a simultaneous study of the hay infusion as a whole was conducted in order to see the changes these microorganisms have on a larger scale. There was a clear distinction between the plates with versus those without the antibiotic, which indicates the presence of antibiotic resistant bacteria within the hay infusion. Over all, the tetracycline did seem to decrease the number of bacterial colonies, but not by much. Additionally, there was a notable difference in the type of colony. Tetracycline decreased the number of white colonies while, which allowed for increased growth of the yellow. Given this observation, it may be assumed the yellow colonies are resistant to tetracycline. There are a number of bacteria that are tetracycline resistant. Nelson and Levy detail three types of tetracycline resistance present in bacteria in their paper “The history of the tetracyclines”. First, some cells have plasmids that show a decreased uptake of the antibiotic. Second, some have a protein-based ribosomal mechanism that serves as a protection against tetracycline. And third, some bacteria chemically inactivate tetracycline using an enzyme in the tet(X) gene (Nelson and Levy, 2011). All of these resistance methods stem from the pretense of a tetracycline repressor protein in the DNA of some bacteria, but not in others.
Works Cited Nelson, M. L. and Levy, S. B. (2011), The history of the tetracyclines. Annals of the New York Academy of Sciences, 1241: 17–32. doi: 10.1111/j.1749-6632.2011.06354.x JL
July 2, 2015 Hay Infusion Culture Observation: Identifying Algae and Protists
In the previous lab, a 500mL hay infusion was created using a soil sample from a transect of land. Despite the size, a culture of this size may be considered an ecosystem with many different niches. Using a dichotomous key, organisms from two different niches within the hay infusion will be identified under a microscope in order to continue the study of biodiversity of the chosen transect. Upon completing these initial observations, a serial dilution will be prepared in order to afford more in depth investigation next week.
The hay infusion was allowed to sit for two days. Four wet mounts were created for microscopic observation from two different niches within the hay infusion. Samples for the first niche were taken from the top of the infusion. Samples for the second niche were taken from the bottom of the infusion. All samples were observed under a microscope. Two protists and algae were identified from each niche using a dichotomous key. Once these steps were completed, four tubes of 10 mL sterile broth were labeled 10-2, 10-4, 10-6, and 10-8. Additionally, eight nutrient agar plates, four with and four without the antibiotic tetracycline were labeled as follows: one each of 10-3, 10-5, 10-7, 10-9. The hay infusion was agitated, and 100 μL from the culture was added to the 10mL broth in the tube labeled 10-2. The tube was stirred thoroughly before 100 μL from this solution was added to the tube labeled 10-4. These steps were repeated twice more to make the 10-6, and 10-8 dilutions. The dilutions were then plated (the 10-2 on the 10-3 plate, and so on) on the nutrient agar plates and spread using a glass spreader. The dilutions were spread most to least dilute on the nutrient agar plates without the tetracycline first and then most to least dilute on the plates with the tetracycline (see Diagram 1).
Diagram 1: Serial Dilution
After sitting for two days, the hay infusion began to smell of manure. Image 1: Hay Infusion
As the image demonstrates, it had separated with most of the sediment sinking to the bottom of the jar. There was plant-like material and possible mold growth floating at the top, all of which was brown in color. The liquid in the middle was a murky grey-brown color. Table 1 Niche Organism Size Classification Top Spirostomum 1-3 mm Ciliate protist Top Peranema 30 mm Flagellate protist Bottom Stentor 1-2 mm Ciliate protist Bottom Colpidium 50-70 mm Ciliate protist The table above details the organisms observed from each niche. They were all motile protists. None were photosynthesizing.
Image 2: Surface Niche
Image 3: Bottom Niche
The observations conducted in this lab were meant to give a preliminary look into the organisms present in two different niches that exists within the transect of land chosen during the last lab. Because each niche had different biotic and abiotic features, it is expected that organisms differ close to versus away from the surface where there appears to be more plant matter. There was a difference in the organisms observed in the niche from the surface of the infusion compared to the organisms observed from the niche at the bottom of the infusion. The majority of these organisms were ciliate protists; heterotrophic organisms that tend to live anywhere there is water. To the extent that the five fundamental characteristics of life (energy, cells, information, replication, and evolution) are visible, the organisms present appear to meet all of them. Consider stentor as observed from the lower niche. Stentor is a unicellular organisms that acquires energy through a filter-feeding method. While not observed, it is known that they reproduce and transfer information asexually (given the number of stentor protists present in the sample, it may be safe to assume reproduction did occur). A two-day growth period is not a lot of time to allow for growth and diversification. If allowed to sit for longer, especially in the presence of sunlight, the infusion would probably support wider range of organisms. More photosynthetic organisms would arise, and the number of heterotrophic organisms would fluctuate with the increase plant life. Setting up the serial dilutions should allow for a glimpse into what may grow given a longer gestation period. JL
June 30, 2015 Observing A Transect at AU
A transect is an ecosystem that contains both biotic and abiotic components. Within a single ecosystem there may be many variable niches, or a specific set of environmental requirements necessary for certain organisms to thrive. The biotic organisms within these ecosystems and niches are further broken down into communities, species, and populations of those species. During this lab, a study of a 20 by 20 foot transect of land on American University’s campus will be conducted in order to observe the biodiversity, complexities, and balance that may exist within a single ecosystem.
The transect of land chosen was just south of Clark Hall and north of McCabe on American University’s campus. Once the transect had been measured out, several handfuls of soil were placed in a plastic bag and returned to the lab. 10 grams of the soil was placed in a plastic jar. 500 mL of Deer Park water and 0.2 grams of condensed milk were added and agitated. The jar was stored with the cap off.
The transect is a grassy area with no shade cover (i.e. no trees or other obstructions to the sunlight) and receives high sun exposure almost all day. It is a low foot traffic area that does not appear to be closely manicured and is covered in different types of grass and weeds. The weeds appear to cover more area than the grass. The growth pattern is patchy. The soil seems to by clay heavy; it is very moist, there is moss growth on the soil in spots, and there are small rocks present in the soil. The ground is uneven with a slight downward slope towards the west, and planted and landscaped mounds surround it. There are several sprinklers on the perimeter of the transect that are close to the landscaped areas, so there is a chance that the transect receives be period, artificial water. The abiotic components include: rocks, soil, sprinklers (on perimeter of transects). The biotic organisms include: grass, clovers, several species of weeds, gnats, tree (on perimeter of transect), grasshoppers, and moss.
The purpose of the procedures detailed above is to begin a study of the intricacies that are typical of any communities within an ecosystem. Eventually, the steps taken to create the hay infusion will go so far as to demonstrate how small changes in two different niches affect the biodiversity of each. The transect that has been chosen appears to be very simple on first glance. Ideally, there would be the time and resources to create several hay infusions from various locations on the transect. This would allow for a more in depth comparison of different ecosystems. However, a transect that seems to have a consistency to the quality of the soil as well as the plant life was chosen. This may provide either a very simple into the biodiversity of the ecosystem, or it may afford an unforeseen amount of variety. JL