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The Presence of Fecal Contamination in Nearby Water Samples.
Brittany Stamper
03/21/2015
The University of Southern Mississippi
The Presence of Fecal Contamination in Nearby Water Samples.
Abstract
The presence of coliforms in a water supply can mean there is fecal contamination circulating in that water, which we tested for in this experiment. It is hypothesized that all of the water supplies will exhibit the presence of fecal coliforms, because they are all outside sources of water that have road runoff, sewage waste and animals defecating in them. We gathered several water samples and inoculated lactose broth with these water samples which tested for fermentation and helped determine if coliforms were present. Then if there was possible coliforms present, we placed them on either an Endo agar
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We observed the agar plate for colonies that had a metallic look, because these were likely E.coli colonies. We then recorded what we saw. After selecting what we believed to be an E. coli coliform colony, we inoculated a TSA slant, in addition to another lactose broth, and incubated these for another 24 hours at 37°C. After incubation, we checked the broth for the production gas and a color change from red to yellow which would indicate the presence of an acid. In order to perform a gram stain, we obtained a colony that we had grown on the TSA slant. Then we gathered one test tube containing tryptone broth, one containing citrate slant, and the last two containing MVRP broths. For the MVRP broths, one test tube should be labeled with MR and the next tube should be labeled VP. Next, we inoculated each test tube with the coliforms that we had confirmed as containing negative
Streak plate technique was used to isolate pure culture for each bacteria (2). The Gram stain was used to determine Gram reaction and morphology of each bacteria (2) Selective and differential media such as, salt agar, MacConkey agar and blood agar were used for bacterial identification (2). Gelatin deeps were inoculated to detect production of gelatinase (2). Starch Agar plate were inoculated to detect amylase (2). Ocular reticle used to determine bacteria size (2). Motility deeps were inoculated to detect motility on bacteria (2). Thioglycollate broth used to determine oxygen requirements (2). Carbohydrate fermentation
The purpose of this study is to identify an unknown bacterium from a mixed culture, by conducting different biochemical tests. Bacteria are an integral part of our ecosystem. They can be found anywhere and identifying them becomes crucial to understanding their characteristics and their effects on other living things, especially humans. Biochemical testing helps us identify the microorganism present with great accuracy. The tests used in this experiment are rudimentary but are fundamental starting points for tests used in medical labs and helps students attain a better understanding of how tests are conducted in a real lab setting. The first step in this process is to use gram-staining technique to narrow down the unknown bacteria into one of the two big domains; gram-negative and gram-positive. Once the gram type is identified, biochemical tests are conducted to narrow down the specific bacterial species. These biochemical tests are process of elimination that relies on the bacteria’s ability to breakdown certain kinds of food sources, their respiratory abilities and other biochemical conditions found in nature.
ABSTRACT: Water samples from local ponds and lakes and snow runoff were collected and tested for coliform as well as Escherichia coli. Humans as well as animals come into contact with these areas, some are used for recreational activities such as swimming and some are a source of drinking water for both animals and humans The main goal of this experiment was to see which lakes, snow run off and ponds tested positive for coliform or Escherichia coli and to come up with some reasoning as to why. It was found that the more remote pond with less contact contained the most Escherichia coli. However, another lake that many swim in and use as their drinking water indeed tested positive for a small amount of Escherichia coli. The two samples from the snow showed negative results for both coliform and Escherichia coli and the two more public ponds that aren’t as commonly used as a source of human drinking water but animal drinking water tested in the higher range for coliforms but in the little to no Escherichia coli range. It was concluded that the remote pond should be avoided as it’s not a safe source of drinking water for humans or animals. Other than that, the the other ponds are likely to be safe from Escherichia coli, but coliforms are a risk factor.
I also inoculated a tryptic soy broth (TSB), a nutrient gelatin deep, a motility agar deep, a fluid thioglycollate medium (FTM) tube, and a TSA plate with my unknown culture. All of these inoculated media were incubated until the next class period (about 48 hours). Then when I came to class most of my inoculated tubes and my streak plate appeared to have growth. The next step I took was making a gram stain to determine the gram reaction and cellular morphology of my unknown. I used my working slant to do this, after careful examination of the gram stain, I learned that my unknown was a gram-positive bacterium. I then preceded by making a negative stain to see the size of the cells of my unknown bacteria. The cell shape was cocci and the cells occurred in clusters of tetrads. After discovering that my unknown bacteria was gram-positive cocci, I turned to page 207 of the lab manual to narrow down my options, there was only four out of the gram-positive list that were
Escherichia coli is a member of the family Enterobacteriaceae. It is a bacterium with a cell wall that has many components. Escherichia coli can live without oxygen which means that it is a facultative anaerobe. It is also capable of fermenting lactose under anaerobic conditions, and in the absence of alternative electron acceptors. There are effects and various factors that limit its growth rate. Its morphology consists of a rod-shaped gram negative bacteria that is commonly found in soil, water, vegetation, human intestines, as well as the intestines of animals. Its presence can be good or bad.
First a control was established for E. coli in a 1.0x nutrient broth. This was
Sterile cotton swab was used to collect a sample from two objects, as well as leaving out a petri dish to open air. One object was unwashed hands and the other object was the back of a cell phone. The two samples were transferred to two separate sterile petri dishes, these petri dishes where all labeled with initials, lab section, and place collected from. All three petri dishes where given to TA who proceeded to place them in an incubator that was at 37° C and allowed to grow to form colonies.
The tubes of E. coli in the phenol red lactose broth one with and the other without mineral oil on top were reddish pink, meaning that the pH was alkaline. That would then mean E. coli can not utilize lactose. However, it is know that E. coli can utilize lactose (BioCoach Activity). The E. coli in a phenol red glucose broth with mineral oil on top produced a yellow color meaning it had a low pH and thus created an acid. There was also gas in the tube which meant that it produced a gas. The E. coli in a phenol red glucose broth without mineral oil was red, that would indicate that an acid was not produced. There was also no gas that was produced.
Coli. Each culture was grown in an M9 medium. One culture utilized glucose as a carbon source, while the other utilized succinate as a carbon source. Two other treatments of E. Coli were also tested, one without succinate and one without glucose. These two treatments were added as a baseline to compare how much succinate and how much glucose actually helped the E. coli grow. The two treatments were covered with parafilm and for the purposes of this experiment, will be called blanks. These cultures remained within their assigned group all day to measure the growth of E. Coli. The following process was repeated by all groups throughout the day. A cuvette was labeled with the sample that was being tested. The writing was at the top of the cuvette to prevent light from being disturbed and affecting results. 3 mL of the tested sample were placed in a flask using a sterilized 1 mL pipet. The spectrophotometer was then rezeroed with the corresponding blank inside. This was so that only growth would be measured. After recording measurements the flasks were returned to the incubator and the pipets were disposed of in a red biohazard bag. The contents of the cuvette were poured into 50% bleach to kill any E. coli. The cuvette was rinsed with distilled water. This process was repeated every 30 minutes over the course of eight and a half hours. Measurements at 12:00, 12:30, and 15:30 were missed due
This lab experiment was conducted in order to identify and confirm the presence of three types of organism within an unknown broth culture. The organism that were being tested fall under the categories of gram-positive, gram-negative paracolons, and gram-negative coliform. Gram positive organism are known to have thick cell wall made up of peptidoglycan, which are cross-link sugar chains with peptide bonds (Carson 13). Gram-positive bacteria are found within the phylum Firmicuites (Slonczewski & Foster 94). While gram-negative, have thin layers of one or two peptidoglycan cell wall, this type of bacteria are found within the phylum Proteobacteria (Slonczewaki & Foster). The gram positive and negative characteristic derived from a staining technique that was developed by Hans
The purpose of this experiment is to measure the effect of flow rates on distilled water by recording its volume every second.
Olowe, S. A., Ahmed, I., Lawal, S. F., Ransome-Kuti, S. (1987). Bacteriological quality of raw human milk: effect of storage in a refrigerator. Annals of Tropical Paediatrics, 7(4), 233-237. Retrieved from http://europepmc.org/abstract/MED/2449844/reload=0;jsessionid=fqrxwgkMoHG1MUPMgzpm.38
The mystery microbe T4 first results were from my gram stain that showed a purple color, rod shape bacteria, that demonstrated the stain was positive. The next test preformed was spore stain, it was positive for spores by taking up on the counterstain that appeared green. The capsule stain was negative the microbes did not have clear halos around them. Next up was the Acid-fast stain that appeared purple/bluish color meaning it was negative.
Fecal coliform, the human and animal wastes, carried to stream systems are sources of pathogenic or disease-causing, bacteria and viruses. (Divya,
Nester, E. W., Anderson, D. G., Roberrs, E. j., & Nester, M. T. (2007). Microbiology . New York: The McGraw-Hill Companies .