Gram-Positive Bacteria

The following tests according to the lab manual were performed: gram stain, fermentation tubes, methyl red, vogues proskauer, sulfur, indole, motility and growing it up on MacConkey agar. The gram stain was performed incorrectly the first time. This is because the decolorizer was not on the bacterium slide for long enough, giving a false outcome.

Bacteria can exist almost everywhere, some are harmless, and some are harmful. There are thousands of different types of bacteria and they fall

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

Gram negative and gram positive bacteria differ from each other in many ways especially in the composition and size of their cell walls. Unlike Gram-positive bacteria, Gram-negative bacteria have a thin peptidoglycan layer surround by an outer membrane. This outer membrane contains many proteins one of them being lipopolysaccharides (LPS), which contributes to the bacteria’s negative charge. One part of this protein is a lipid, called Lipid A, which is considered an endotoxin because this lipid triggers an immune response stimulating fever

Only the Gram positive organisms were put through the starch test, as testing the Gram negative organisms would not have helped with identifying the unknown organism. Proper inoculation techniques were used to transfer the Gram positive organisms onto the starch plate in a straight line, and then the plate was incubated. The following week, after incubation, the plate was flooded with Gram’s iodine and was left to sit for ten minutes. Gram’s iodine indicated the presence of starch by forming a starch iodine complex that turns blue-brown or black. After pouring the excess iodine off, the plate was set on a white paper to analyze. A positive test with amylase present would present itself with a halo of clearing around the cell

Bacteria is a single celled organism, bacteria have evolved to thrive in almost any environment and can be found in almost any substance/surface and also in the human body, only 1% of bacteria is actually harmful.

Name one endogenous source of contamination and discuss the modes of transmission from the source to the new host.

For the Gram stain, I used a microscope slide, wash bottle of water, clothespin, and the reagents crystal violet, Gram’s iodine, ethyl alcohol, and safranin to observe whether the organism was Gram negative or positve. The method was placing and heat fixing a loopful of the unknown organism on the slide. The organisms were then stained with crystal violet for a minute, rinsed off, flooded with iodine for a minute, rinsed off, decolorized with alcohol for 30 seconds, and then finally stained with the counter-stain, safranin, for a minute. I streaked the unknown onto a TSA plate and incubated at 35 C. With a pure colony, we performed a second gram stain procedure and inoculated it into a TSA slant. I used BCP Lactose broth and a loop to perform a BCP Lactose test. The broth was inoculated with a loopful of the unknown and incubated

The morphology of Klebsiella pneumoniae is a bacillus shaped bacterium, which means it looks like a rod. This bacterium is also a non-motile and Gram-negative. This means that wherever the bacterium was inoculated into an agar, it only grew in that spot and did not grow outwards. Being gram-negative means that the bacterium has a thin layer of peptidoglycan and stains red during the Gram-staining process. Klebsiella pneumoniae is an encapsulated bacterium, where the capsule

　　I did a Fermentation Test where I inoculated 1 tube of phenol red sucrose, 1 tube of phenol red glucose, and 1 tube of phenol red lactose. A positive result for fermentation would be a change of the color (original color is a red color to a yellow color) of the sucrose, lactose and glucose as well as if gas was produced, gas bubble present within the Durman tube inside the test tube.

Scientists classify bacteria according to shape. Most bacteria fall into three different categories of shape: cocci(round), spirilla(spiral-shaped), and bacilli(rod-shaped). Another way scientists distinguish bacteria is by staining them. Because gram-negative bacteria have thinner cell walls than gram-positive bacteria, when stained with purple dye, gram-negative bacteria will not retain the purple color the dye, but will appear pink or red. Gram-positive bacteria will appear purple.

The Gram-positive cell wall is composed of peptidoglycans, a thick layer of protein-sugar complexes taking up 60-90% of their cell wall. Peptidoglycan is composed of two glucose derivatives, N-acetylmuramic acid and N-acetylglucosamine alternated and cross-linked by tetrapeptides that is composed of L-alanine, D-glutamine, L-lysine

Lee and Bishop (2016) have stated, microbes are microscopic, living single-celled organisms such as bacteria, fungi and virus which is non-living. The structure of bacteria that causes disease are the fimbriae, flagella and toxins. The cell wall provides structural support and the cell membrane supports the functions that occur in the subcellular structure of higher organisms. Bacteria are also known as pathogens which cause diseases in plants, insects, animals and humans. Flagella are the long tail-like structures that enable the bacteria to swim or move and it also help the bacteria to migrate to its site of infection and survive. Toxins are very harmful which are responsible for the symptoms and complication of the disease. One of the chronic disease caused by bacteria is tuberculosis (TB). A favourable environment such as time, temperature and oxygen are important to facilitate the bacterial growth.

Gram-negative and Gram-positive bacteria can be distinguished by conducting biochemical tests. Its growth can be examined on selected mediums containing metabolic inhibitors such as antibiotics. The medias utilized in this project are: media containing the antibiotic vancomycin, Eosin Methylene Blue-lactose (EMB-lactose) agar, and Phenylethyl alcohol (PEA) agar (Holbrook and Leicht 2017). Vancomycin is most effective at inhibiting the growth of Gram-positive cocci (Rotschafer et al. 2005). EMB-lactose agar inhibits the growth of many Gram-positive bacteria and the lactose component of the medium allows for lactose-fermenting bacteria to cause color changes within the medium (Reynolds 2011). The presence of lactose fermenters in the EMB-lactose agar will cause a color change in the resulting colonies based on the strength of their acidity. A metallic green colony indicates that that active fermenters are present. A pink colony indicates that weak fermenters are present. If a bacterial species does not ferment lactose, it will appear white or colorless (Holbrook and Leicht 2017). PEA agar is a selective medium that inhibits the growth of most Gram-negative bacteria species. It does so by interrupting the lipids within the Gram-negative cell membrane, which will affect its permeability to certain molecules. This could result in the intake of materials that are supposed to be blocked and the release of potassium from within the cell, which is necessary for DNA synthesis (Lal and