True statement about Gram staining?

• A. Gram-positive bacteria retain crystal violet and stain purple due to thick peptidoglycan.
• B. Gram-positive bacteria stain pink.
• C. Gram-negative bacteria stain purple due to thick peptidoglycan.
• D. Gram staining cannot differentiate bacteria.

Answer: (A)

Gram staining differentiates bacteria by their cell wall structure. Gram-positive bacteria have a thick peptidoglycan layer that locks in the crystal violet–iodine complex during the decolorization step, so these cells appear purple under the microscope. This is why the statement is true: the thick peptidoglycan lets the stain stay, giving a purple color.

In contrast, Gram-negative bacteria have a thinner peptidoglycan layer and an outer membrane; during the decolorization step they lose the crystal violet, and then they take up the counterstain (usually pink or red), so they appear pink. Saying Gram-positive stain pink or that Gram-negative stain purple contradicts the observed results. And Gram staining does differentiate between these groups, so the idea that it cannot differentiate is incorrect.