After being assigned a broth containing two unknown bacteria on 3/17/15, I inoculated a sterilized wire loop with the bacteria and streaked it across a TSA plate. This was to separate the two bacteria. After two rounds of streaking, I discovered one of my bacteria was dead and had to acquire it separately from Dr. Christmann.
Once my bacteria were separated, I began Gram staining. To Gram stain, I took a slide and spread each bacterium with a droplet of water onto two different slides. After heat-fixing, I stained with Crystal Violet for 1 min, rinsed with distilled water, stained with Gram Iodine for 1 minute, rinsed with distilled water, gently rinsed with 95% ethanol, rinsed with distilled water again, stained with Safranin for 1 minute, rinsed with water, and dried the slides with bibulous paper. I checked the bacteria with Microscope 90. For the rest of the lab, I performed different tests on the two different bacteria.
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On 4/20/15, I set up a TSA stab test for this unknown bacterium. I checked the TSA tube on 4/21/15. On 4/23/15, I performed an oxidase test and an endospore stain. The oxidase test required the bacteria to be spread on an oxidase dry slide. Results take about 20 seconds to appear and be confirmed. For the endospore test, I used the Schaffer-Fulton spore staining method. I spread the bacteria on a slide and covered it with a piece of bibulous paper. I applied malachite green stain on top of the paper and steamed the slide for 6 minutes, reapplying drops of malachite green stain periodically. After steaming, I removed the paper, rinsed with distilled water, counterstained with Safranin for 1 minute, rinsed with distilled water again, and dried the slide with bibulous paper. I checked the endospore stain with Microscope
An unknown bacterium was handed out by Dr. Honer. The appropriate tests were prepared and applied. The first procedure that was done was the gram stain. Under a microscope, if the gram stain is purple, the bacterium is gram positive, if the stain is red, it is gram negative. The next test was the fermentation tests for glucose, sucrose and
The first step toward identifying this unknown organism was to perform a Gram Stain to differentiate between gram positive and gram negative bacteria. This is an important step because it directs what the next tests will be. My Gram Stain on sample #12 showed that the bacteria was gram negative, however, after receiving the results of the OF glucose, H2S, Citrate, Urease and Motility tests, it was apparent that my Gram Stain was contaminated. I then performed a catalase test which came back negative, so I ordered a Bacitracin disc, Optochin disc and a CAMP test which had to be incubated overnight. After receipt of those test results,
The unknown bacteria plate chosen was plate #2. It was identified to be Micrococcus luteus. It is a gram positive, Coccus bacteria that is commonly found in dust, water, soil, and the air. M. luteus also thrives in the human mouth and upper respiratory tract. Sir Alexander Fleming discovered it in 1928 before he identified penicillin. It is part of the normal flora on human skin as well as other mammals. Since it is part of normal flora it is normally not pathogenic, but can become opportunistic in an immune-deficient person. It has been known to cause septic shock, UTI’s, and even pneumonia. Micrococcus luteus is both urease and catalase positive. It does not utilize tryptophan for indole production. It is a facultative anaerobe. Mobility is not present for this bacterium. Starch is also not hydrolyzed and oxidase is not present.
1) Apply the stain to your first unknown slide and examine it under the microscope.
This report discusses the procedures and equipment needed to determine a bacterium knowing and having nothing but the sample bacteria given by the instructor. The importance of this lab is to test your ability to think clearly about what each test will tell you about your unknown organism and selecting tests that will save time and give you the most information. The objective of this lab is to determine what species level and genus level your bacterium is categorized as. The first step taken to determine the bacteria was to do a sub-culture of my stock broth to have a backup of my bacteria just in case my broth became contaminated. Doing a T-streak on a TSA plate is when the plate is divided into three sections and using an inoculating loop, grab some
The main idea of this experiment was to correctly identify the unknown bacteria, #3. Identification of unknown bacteria yields multiple benefits in many different areas in the research of microorganisms. In this experiment I performed many different test dealing with things such as the presence of enzymes, fermentation abilities and different chemical reactions. Observations made from the tests were then compared to a gram negative unknown chart in order to identify the bacteria. Based off of my results and the chart, I concluded the bacteria #3 was the bacteria Escherichia coli. E. coli is most commonly found in the intestines of warm blooded organisms. Most E. coli strands are non pathogenic however, there are strands
I identified Citrobacter freundii, the gram negative rod, by running a series of tests. I began with the Phenol Red Lactose tests, which tests if the organism contains various enzymes that determine if it can ferment lactose. The broth turned yellow after it was incubated, indicating that the lactose was fermented to acid, and there was also gas present in the Durham tube. Since the Phenol Red Lactose Test was positive, I then ran the Phenol Red Sucrose test, which tests if the bacteria contain different enzymes that determine if sucrose can be fermented. After incubation, the broth was yellow, indicating that sugar was fermented to acid, and there was also gas present in the Durham tube. Next, I ran the Sulfide Production, Indole Formation, Motility test, but I was only testing for Hydrogen Sulfide Production to differentiate between the organisms Citrobacter freundii and Enterobacter aerogenes. This test detects if the organisms can metabolize sulfur into hydrogen sulfide, which is revealed by the formation of ferrous sulfide that causes blackening around the growth. The test also reveals if the organism can break tryptophan into indole or migrate away from initial stab area. After incubation, the agar slant was completely black, indicating that the organism produces hydrogen sulfide and is motile proving that it was Citrobacter
Unknown lab report# 24 Introduction Microbiology is a study of organisms that surrounds us every day. It requires an educational understanding to identify organisms, and the ability to distinguish different bacteria’s. In applying the learning process of the different bacteria’s, unknown bacteria samples are given to be studied and identified.
The bacteria that was contained within Unknown tube #12 is believed to be Pseudomonas aeruginosa, Figure 1. The bacteria tested to be Gram Stain negative, producing a pink, red color retained from the staining process. When the species of bacteria was plated on nutrient media, the cells produced an irregular and spreading configuration as shown in Figure 2. This same plating test provided the margins and elevation, lobate and hilly, respectively. The specimen was stabbed in a Fluid Thioglycollate Medium (FTM) tube using an inoculated loop of the bacteria. The results of this experimentation indicate the type of oxygen requirement of the bacteria. The test found the bacteria to be aerobic as colonies of the bacteria began to form along the top of the FTM tube (Manual 2017).
The biochemical tests, the Gram stain, the Bartholomew and Mittwer’s stain, the microscopic and macroscopic observations, and chemical sensitivity tests helped to identify the unknown bacteria as Citrobacter Freundii. Each biochemical test result maintained consistency with the
To start off unknown #3, I picked out O17 from rack A and wrote down my name, date, class, and I first noticed the broth was a light yellow that was quite cloudy or milky. Soon later I streaked my organism onto a TSA plate and incubated it at 37oC for 24 hours, t was almost a perfect plate. There was no contamination; I could tell it there was only one species on the plate and that it was okay for me to move on. There were many isolated colonies in quadrants 3 and 4. The colonies were small and white in color, it was not translucent. I then began to make my 2 slants, inoculating a straight line on each slant, I incubated it at 37oC for 24 hours. The next day, I made sure my slants were pure and also there was a thick line of organism where I had inoculated with a yellow-white color. Now I was ready to begin making my slides and start staining. I used E. Coli and S. Aureus next to my unknown to help me
Two smears of the unknown bacterium #5 were inoculated while the second smear was used for a back up. The unknown bacterium dried for at least forty minutes. After the smears dried, the slides were heat fixed two times to ensure the stability of the organism. The slide was placed on top of the staining rack then over the small sink.
Purpose: is to determine the unknown bacteria with a variety of biochemical tests. There are many reasons that contribute to why it is so important to test patients for both high and low risks diseases. The most important reason would be to know the identity of microorganism and how it can be treated .This study was performed in microbiology laboratory class by applying the microorganism to the tests that have been performed in the class prior to the identification of the unknown. First, the lab professor handed out a bacteria that was on the unknown streak plate labeled B5 that consisted of an unknown gram positive or gram negative bacteria.
The lab manual confirmed the experiments findings to be correct and each one were pink colored on the slide under a microscope. If the result had varied from gram negative a reason could include leaving the decolorizer on the slide for one minute instead of ten seconds. Doing this could make the slide appear any color other than pink. The results could also have varied
The Unknown Bacteria 36/Bacteria # 2 on a TSA plate was examined by the naked eye and under a dissecting microscope. Bacteria # 2 was approximately 3 - 4 mm in diameter. They were circular in form with an entire margin and a flat elevation. The colonies were rough (granular), translucent, and white brownish color with black granules. The Gram stain resulted in a Gram negative rod. After the Gram stain was completed, the bacteria were streaked on an Eosin -Methylene Blue Agar plate and an Enterotube II was inoculated.