a)Bacterial inoculum preparation (isolation and culturing) (95,97): - A pure standard Enterococcus Faecalis strain (ATCG 29212) was isolated and sub- cultured in blood agar plates and incubated at 370c for 24hours. - A pure single E.faecalis colony was isolated from the cultured plates and gram stained to confirm the colony growth under microscope.(Fig.12) - The isolated colony was then inoculated with nutrient broth and incubated again at 370c for 24hours in anaerobic conditions. The bacterial growth was checked by changes in the broth turbidity. - A standardized suspension of turbidity adjusted spectrophotometrically to 0.5 Mc- Farland units , equivalent to 1.5x108 colony forming units / milliliter (CFU/ml).(Fig.13) b)Infection of dentin
During the purification section of this lab, the LB/amp/ara agar plate was examined for well-isolated green colonies and the LB/amp plate was observed for white colonies with space between each other. These colonies were circled on the outside of the plates using a marker. Next, two 15 milliliter culture tubes containing 2 milliliters of nutrient growth media were obtained and labeled “+” and “-“. Using a new inoculation tube, the circled colonies from each plate were scooped out and immersed in their respective culture tubes. Once the bacteria was mixed into the solution, the tubes were sealed and placed horizontally into the 32⁰ incubator for 24 hours.
The purpose of this lab was to identify two unknown bacteria cultures using various differential tests. The identification of these unknown cultures was accomplished by separating and differentiating possible bacteria based on specific biochemical characteristics. Whether the tests performed identified specific enzymatic reactions or metabolic pathways, each was used in a way to help recognize those specifics and identify the unknown cultures. The differential tests used to identify the unknown cultures were oxidase, catalase, lactose and sucrose fermentation, Kugler/iron agar, nitrate reduction, gelatin hydrolysis, starch hydrolysis, manitol salt, MR-VP, citrate, bile esculin,
The possible organisms in this lab included: Serratia marcescens, Escherichia coli, Bacillus subtilis, Sarcina lutea, Micrococcus luteus, Pseudomonas fragi, Staphylococcus epidermidis, Alcaligenes faecalis, Micrococcus roseus, and Clostridium sporogenes.1 In start of the elimination process, the two nutrient agar plates that were made by unknown tube number two were observed and characterized by their colonies. Gram positive: Plate A was incubated at 37 C and had a colony appearance of bright yellow dots on the nutrient agar. This eliminated the possibilities of S. marcescens, E. coli, B. subtilis, P. fragi, S. epidermidis, A. faecalis, M. roseus, and C. sporogenes. S. marcescens colonies are red; M. roseus colonies are orange; E. coli and A. faecalis colonies are beige; B. subtilis, C. sporogenes, and S. epidermidis colonies appear in white.2 This left
An unknown bacterium was handed out by the lab instructor. The methods that have been learned so far in identifying bacteria were applied to this unknown. Procedures were followed as stated in the lab manual and biochemical test handouts. The first procedure that was done was a gram stain followed by a streak of the unknown on a TSA plate in order to determine the gram reaction and observe the colony morphology. After that, specific biochemical tests were performed for gram positive, since unknown number five was determined to be gram positive rod. The other tests were performed in this order: Mannitol Salt (MSA) streak, Blood Agar streak, Catalase test, Nitrate Reduction test, and Phenyl
Abstract: The objective of this experiment is to find out what the optimal growth environment is for V. natriegens out of three different environments. The first is a Brain Heart Infusion broth (BHI) that contains 50 mM of NaCl. The second is BHI containing 250 mM of NaCl. The third is BHI containing 1000 mM NaCl. In order to obtain this, a growth curve graph must be constructed for each different environment, and the generation time of the bacteria in each environment must be calculated. The optimal environment for V. natriegens was found to be the BHI with 250 mM of NaCl.
The first result of importance was the result of the Gram stain. The observations of the unknown bacteria from the slant culture after Gram staining showed that the unknown bacteria were Gram negative bacilli (Image 1). After determining the unknown bacteria was Gram negative, an oxidase test was conducted on a sample from the slant culture. The cotton swap with the sample of bacteria did not change color when the oxidase reagent was applied, thus providing a negative result. With a negative oxidase test, further tests were conducted to determine various characteristics of the unknown bacteria. A MR-VP broth was inoculated with a sample from a slant culture of unknown bacteria. After incubation, the methyl red reagent was added to the broth, and the broth turned red, providing a positive result (Image 2). An EMB agar streak plate was inoculated with a sample from a slant culture of the unknown bacteria, and after incubation, growth was found on the plate, providing a positive result (Image 3). A Citrate agar slant was inoculated, and after incubation, growth was found on the media, providing a positive result (Image 4). A Urea agar slant was inoculated, and after incubation, the agar had changed from a peach color to a bright pink color, providing a positive result (Image 5). Using the flowchart (Figure 1) developed from the Table of Expected Results, the lab partners started at the oxidase test. Given the negative result of the oxidase test, the flowchart is
When reflecting back to experiment 3, Aseptic Technique and Culturing Microbes, I realized the large amount of microorganisms that can be found in everyday life. Many different types are found with in the human body. Theses experiments focused on two types of bacteria. First was Staphylococcus epidermidis, found on the skin, and second was Lactobacillus acidophilus, found in the gastrointestinal tract. Both have similar needs for growth when it comes to temperature, however, different growth environments are used.
The purpose of this lab was to identify two unknown bacteria cultures using various differential tests. The identification of these unknown cultures was accomplished by separating and differentiating possible
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 purpose of the following study is to determine where the two unknown bacteria acquired in Microbiology lab should be classified in regards to temperature, pH level, and osmoregularity. It is important to classify bacteria in order to identify them. Identification of bacteria is important because they are not only useful but potentially dangerous as well. The identification of bacteria can lead to breakthroughs in healthcare regarding treatment of old and new diseases alike. Identifying bacteria can also be used in many other areas from better crop production through microbial pesticides to biological warfare. Their uses are endless as are their abilities to evolve and adapt to changing environments. That is why it is so important
This laboratory experiment’s objective was to take a pure culture and isolate it from a mixed culture. The other part of the objective was to ascertain what species of bacteria that the pure culture was. The hypothesis made stated that so long as lab protocol was followed, the unidentified culture would be positively recognized/identified. An isolated pure colony of the unknown culture was obtained using the quadrant streak plate method. Afterward, the culture was Gram stained, and the results showed that it was Gram positive. Motility tests were done on the unknown using a filter paper bridge on a petri dish that contained TTC with agar. The unknown was revealed to not be motile, which meant that it did not possess flagella. The last test done was to learn the metabolic capabilities of the unknown bacteria. There were tests done for citrate utilization, the mixed fermentation pathway, catalase presence, carbohydrate fermentation in mannitol, lactose and glucose, urease production and the butanediol fermentation pathway in order to better identify the unknown bacteria. The results from each of the metabolic tests in conjunction with the motility and Gram staining tests were ultimately compared to results from database containing many different kinds of results from various bacteria. The unknown from the mixed culture was identified as Staphylococcus
A mixed culture of two unknown bacteria was provided by the instructor. The methods used for
In the beginning of lab, we were advised to obtain a nutrient agar petri plate, which is used for the cultivation of microbes supporting growth of non-fastidious organisms. Since it contains many nutrients, a wide variety of bacteria and fungi can grow. Taking the plate,
2. Introduction: Each student was given unknown bacteria and was instructed to perform a variety of experimental tests that would help to identify their bacteria. During the process of identification, the unknown bacteria was added to many different testing medias using aseptic technique. They are as follows: lactose fermentation on eosin methylene blue (EMB), TSI (Triple Sugar Iron agar), Phenol red sucrose, the SIM test, H2S by SIM, IMViC (indole, motility, voges-proskauer, and citrate), Urease (urea broth), PDase (Phenylalanine Deaminase), Lysine Decarboxylase, and Ornithine Decarboxylase. Colonial morphology on EMB was used to
Each mixed culture that was tested had one gram positive and one gram negative bacterial species. The possible species of bacteria that could have been isolated from the mixtures included the following: Bacillus subtilis, Corynebacterium glutamicum, Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, Enterobacter aerogenes, Salmonella enterica, and Pseudomonas aeruginosa. The identities of the unknown species were determined through comparing the experimental data against data acquired from earlier experimentation.