METHOD:
Two unknown bacteria were provided that were labeled “12A” and “12B”. Both bacteria were then used individually following aseptic technique to create heat-fixed smears. Both bacteria were strategically placed on the slide in order to prevent cross contamination and inaccurate results. A total of three slides were made in order to ensure accuracy. The heat-fixed slides were then Gram stained using the procedure listed in the manual; followed by observation under a Leica Dm500 microscope. The slides were observed under 4X,10X,40X and oil immersion and both unknown reactions were double checked by Dr. McLaughlin.
Results from Gram staining procedures were provided a week later; hence, a dichotomous (a flowchart that is used to identify bacteria based on its taxonomic classification) was constructed based on a total of 10 possible unknown bacterium. The bacteria were then separated based on its Gram reaction; leading to two subcategories, including Gram positive and Gram negative. Both subcategories were then divided into two separate categories based on bacteria morphology, including rod and cocci shaped. A series of metabolic test were then selected strategically based on metabolic results provided by Dr. McLaughlin (See attached chart for possible results). The metabolic test for Gram positive rod
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It was then determined that unknown bacteria that could be categorized based on the mentioned gram reaction and morphology were either Cornybacterium xerosis or Bacillus subtilis; hence, a starch hydrolysis test, will be used to determine which bacterium could be the unknown. Metabolic test that are suitable for Gram positive cocci bacterium including, Micrococcus luteus, Staphylococcus aureus and Streptococcus faecalis were also included in the constructed dichotomous key; however, metabolic testing on GM+ cocci was not utilized due to gram staining
I identified the genus and species of an unknown bacterial culture, #16, and I applied the following knowledge of morphologic, cultural and metabolic characteristics of the unknown microorganism according to the laboratory manual as well as my class notes and power point print outs. I was given an incubated agar slant labeled #16 and a rack of different tests to either examine or perform myself; the tests are as follows: Gram Stain; Nutrient Gelatin Test; Carbohydrate Fermentation; Dextrose, Lactose and Sucrose; IMVIC tests; Citrate, Indole, Mythel-Red and Vogues Proskauer test; as well as a Urease and TSI Test. Materials and Methods/Results Upon receiving the Microorganism (M.O.) #16, I prepared a slide by cleaning and drying it. Then, using a bottle of water I placed a sterile drop of water on the slide and used an inoculating loop, flame sterilized, I took a small sample of the unknown growth in my agar slant and smeared it onto the slide in a dime sized circle and then heat fixed it for ten minutes.
The isolate possesses some enzymes required for hydrolytic reactions. Hydrolytic enzymes found to be secreted from the bacterium, are -amylase, casein, and PYRase. In the starch hydrolysis and casein tests, there was a zone of clearing around the bacterium, which was indicative of the secreted enzymes necessary to break down starch and casein. In the PYR test, the presence of PYRase was detected by a color change to red on the PYR disc after the addition of the PYR reagent (p-dimethylaminocinnamaldehyde). Hydrolytic enzymes for which the EI tested negative were urease, gelatinase, and DNAse. In the Urea Hydrolysis test, it was observed that the urea broth did not have a color change, indicating that there was no urease secreted to break down urea in the broth. Similarly, there was no gelatinase present to break down gelatin in the Gelatin Hydrolysis test, so the nutrient gelatin remained solid. It was concluded that the EI does not possess DNase because there was no clearing zone around the bacteria, indicating that DNA had not been
After 5 days of growth each slant was tested using the gram staining technique to confirm the complete isolation of the bacteria. Both isolations were completely successful. Then each sample of bacteria was subjected to a series of tests for identification.
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.
The results of the gram stain test were cocci and purple. This indicated that the unknown bacteria were gram positive. The gram stain test eliminated Escherichia coli, Klebsiella pneumonia, Salmonella enterica, and Yersinia enterocolitica as choices because these bacteria are gram negative. Next a Blood Agar plate was used because in order to do a MSA or a Catalase test there needs to be a colony of the bacteria. The result of the Blood Agar plate was nonhemolytic.
The purpose of this laboratory is to learn about cultural, morphological, and biochemical characteristics that are used in identifying bacterial isolates. Besides identifying the unknown culture, students also gain an understanding of the process of identification and the techniques and theory behind the process. Experiments such as gram stain, negative stain, endospore and other important tests in identifying unknown bacteria are performed. Various chemical tests were done and the results were carefully determined to identify the unknown bacteria. First session of lab started of by the selection of an unknown bacterium then inoculations of 2 tryptic soy gar (TSA) slants, 1 nutrient broth (TSB), 1 nutrient gelatin deep, 1 motility
In the last decade, the number of prescriptions for antibiotics has increases. Even though, antibiotics are helpful, an excess amount of antibiotics can be dangerous. Quite often antibiotics are wrongly prescribed to cure viruses when they are meant to target bacteria. Antibiotics are a type of medicine that is prone to kill microorganisms, or bacteria. By examining the PBS documentary Hunting the Nightmare Bacteria and the article “U.S. government taps GlaxoSmithKline for New Antibiotics” by Ben Hirschler as well as a few other articles can help depict the problem that is of doctors prescribing antibiotics wrongly or excessively, which can led to becoming harmful to the body.
The purpose of this project was to identify unknown bacteria species from a mixed culture. The two unknown species were initially plated onto Tryptic Soy Agar (TSA), Eosin Methylene Blue (EMB), Mannitol Salt Agar (MSA), and blood agar plates to distinguish between the two different bacteria using colony size, color, shape, and growth characteristics. By identifying and inoculating the differing types of colonies, the two unknown bacteria were purified and able to be tested
During this investigation to identify the unknown bacterium, seven different biochemical tests were performed. All biochemical tests were performed according to the Eastfield Microbiology Lab Manual. As with...
E. coli are bacteria that can cause an infection in various parts of your body, including your intestines. E. coli bacteria normally live in the intestines of people and animals. Most types of E. coli do not cause infections, but some produce a poison (toxin) that can cause diarrhea. Depending on the toxin, this can cause mild or severe diarrhea.
Bacterial cells, like plant cells, are surrounded by a cell wall. However, bacterial cell walls are made up of polysaccharide chains linked to amino acids, while plant cell walls are made up of cellulose, which contains no amino acids. Many bacteria secrete a slimy capsule around the outside of the cell wall. The capsule provides additional protection for the cell. Many of the bacteria that cause diseases in animals are surrounded by a capsule. The capsule prevents the white blood cells and antibodies from destroying the invading bacterium. Inside the capsule and the cell wall is the cell membrane. In aerobic bacteria, the reactions of cellular respiration take place on fingerlike infoldings of the cell membrane. Ribosomes are scattered throughout the cytoplasm, and the DNA is generally found in the center of the cell. Many bacilli and spirilla have flagella, which are used for locomotion in water. A few types of bacteria that lack flagella move by gliding on a surface. However, the mechanism of this gliding motion is unknown. Most bacteria are aerobic, they require free oxygen to carry on cellular respiration. Some bacteria, called facultatibe anaerobes can live in either the presence or absence of free oxygen. They obtain energy either by aerobic respiration when oxygen is present or by fermentation when oxygen is absent. Still other bacteria cannot live in the presence of oxygen. These are called obligate anaerobes. Such bacteria obtain energy only fermentation. Through fermentation, different groups of bacteria produce a wide variety of organic compounds. Besides ethyl alcohol and lactic acid, bacterial fermentation can produce acetic acid, acetone, butyl alcohol, glycol, butyric acid, propionic acid, and methane, the main component of natural gas. Most bacteria are heterotrophic bacteria are either saprophytes or parasites. Saprophytes feed on the remains of dead plants and animals, and ordinarily do not cause disease. They release digestive enzymes onto the organic matter. The enzymes breakdown the large food molecules into smaller molecules, which are absorbed by the bacterial cells. Parasites live on or in living organisms, and may cause disease. A few types of bacteria are Autotrophic, they can synthesize the organic nutrients they require from inorganic substances. Autotrophic bacteria are either photosynthetic or Chemosynthetic. The photosynthetic bacteria contain chlorophyll that are different from the plant chlorophyll. In bacterial photosynthesis, hydrogen is obtained by the splitting of compounds other than water.
Microbes are microscopic life forms, usually too small to be seen by the naked eye. Although many microbes are single-celled, there are also numerous multi-cellular organisms. The human body has 10-100 trillion microbes living on it, making it one giant super-organism. Since the first link between microbes and diseases was made, people have been advised to wash their hands. Scientists, however, have recently started to investigate more closely how the microbes that call the human body home affect our health. While some microbes cause disease, others are more beneficial, working with our bodies in many subtle ways.
Bacteria are small microorganisms that are found in all environments, they are the building blocks of all life. Bacteria are ubiquitous, they are present everywhere. Bacteria have been present on earth for approximately 3.5 billion years (1, p.1). Bacteria serve a variety of different functions that are ultimately vital to life but can also threaten human life. Bacteria may be small, but our life is dependent upon their activity and growth (1, p.1). Since bacteria are so small and our life is dependent upon their functions and activities, it is important that they are able to divide and reproduce. Bacteria cannot grow and divide unless it is in a favorable environment (2). Each bacterial species has its own specific, optimal requirements for maximal growth and reproduction. In order to deem an environment favorable, it must have the correct oxygen availability, temperature, nutrients, moisture, salinity, and osmotic pressure. Other factors, such as the presence of toxic products can stunt or harm the growth of bacteria.
The abnormal presence of bacterial growth can be inspected under a microscope. If the organism inspected is not the bacteria used in the experiment, it means that the growth of the bacterial culture investigated is absent. By using this method, contamination by foreign substances in the surrounding air can be ruled out and the results would be more accurate.