Why gram positive bacteria do not counter stain as with gram negative?
Gram-positive bacteria do not counterstain as with Gram-negative bacteria because they have a much thicker peptidoglycan layer in their cell wall.
To understand why this is the case, let's first discuss the Gram staining technique. It was developed by a Danish bacteriologist named Hans Christian Gram in the 1880s. The Gram stain is a differential staining method used to categorize bacteria into two groups: Gram-positive and Gram-negative.
In the Gram staining procedure, a bacterial smear is first treated with crystal violet, a purple-colored dye. This dye binds to the peptidoglycan layer, which is present in both Gram-positive and Gram-negative bacteria. Then, the bacteria are treated with iodine, which forms a crystal violet-iodine complex, helping to retain the dye in the cell wall.
Next, a decolorizing agent, typically alcohol or acetone, is applied. This agent dehydrates the peptidoglycan layer, causing it to shrink and become more compact in Gram-positive bacteria. As a result, the crystal violet-iodine complex is trapped within the thick peptidoglycan layer, making the bacteria appear purple.
On the other hand, the decolorizing agent dissolves the outer membrane of Gram-negative bacteria, making the peptidoglycan layer more porous. This leads to the loss of the crystal violet-iodine complex, resulting in the bacteria becoming colorless.
Finally, a counterstain, usually safranin, is applied to the bacterial smear. The safranin stains the decolorized Gram-negative bacteria, giving them a pink or red color. However, the thick peptidoglycan layer of Gram-positive bacteria acts as a barrier, preventing the counterstain from entering and coloring the cells.
In summary, the reason Gram-positive bacteria do not counterstain as with Gram-negative bacteria is due to their significantly thicker peptidoglycan layer. This layer prevents the counterstain from penetrating the cells, resulting in Gram-positive bacteria retaining the initial purple color obtained from the crystal violet dye.