After performing a Gram stain, why do Gram-positive bacteria appear purple under the microscope?

• A. They lack peptidoglycan, allowing the crystal violet to bind directly to the cell membrane.
• B. They retain the crystal violet stain due to their thick peptidoglycan layer.
• C. Their outer membrane prevents the crystal violet from being washed away.
• D. They absorb the safranin counterstain more strongly than Gram-negative bacteria.

Answer: (B)

Verified step by step guidance

1. Understand the basic principle of the Gram stain: it differentiates bacteria based on the structure of their cell walls, particularly the thickness of the peptidoglycan layer.
2. Recall that Gram-positive bacteria have a thick peptidoglycan layer, which is a mesh-like polymer that provides rigidity and retains certain stains.
3. During the Gram staining process, crystal violet dye is applied first, which penetrates all bacterial cells and stains them purple.
4. Next, iodine is added as a mordant, forming a crystal violet-iodine complex that gets trapped within the thick peptidoglycan layer of Gram-positive bacteria.
5. When the decolorizing agent (usually alcohol or acetone) is applied, it dehydrates the thick peptidoglycan layer in Gram-positive bacteria, trapping the crystal violet-iodine complex inside, so these cells remain purple under the microscope.