Biology

5/17/13

If I were given a sputum sample to analyze in order to find out what is wrong with my patient I would look at each bacterium individually. The three types we will be identifying are, and Mycoplasma, Bacillus, Escherichia. To identify the bacteria I would first determine the staining protocol to be used and then proceed to the staining process.

To identify gram stain you would follow certain steps. A heat smear is covered with a basic purple dye, usually crystal violet. Because purple stain gives its color to all cells it is referred to as the primary stain. After a short period the purple dye is washed off and then the smear is covered with iodine, a mordant, which simply means it helps keep the dye in place once it’s bound to wherever it binds. When the iodine is washed off, both gram-positive and gram-negative, bacteria appear dark purple or violet. Then the slide is washed with alcohol or alcohol-acetone solution. The solution is a decolorizing agent which means it removes the color from some of the species but not all of them. The alcohol is then rinsed off and the slide is stained with safranin, a basic red dye. The smear is washed again and blotted dry and looked at under a microscope. The purple dye and iodine combine in the cytoplasm of each bacterium and color it dark violet or purple. Bacteria that retain their color after decolorizing are gram-positive; bacteria that lose their purple color are considered gram-negative.

Staining is important because it helps correctly identify the type of bacteria and provide valuable information for treatment of the disease. Gram-positive bacteria tend to be killed easier than gram-negative ones because the antibiotics cannot penetrate the lipopolysaccharide layer, which exists outside the cell wall.

Mycoplasma are unlike bacillus because they have little to no cell walls and are said to be the smallest known bacteria that can grow outside the living host cells. At first...