Final answer:
Observing blue E. coli after a gram-stain could be due to improper staining techniques or expired stains, as E. coli is a gram-negative bacteria that should appear pink, not blue or purple.
Step-by-step explanation:
One possible explanation for observing blue E. coli after gram-staining is a result of improper staining technique or expired stains that might cause E. coli, which is a gram-negative bacteria, to retain the crystal violet dye and appear blue or purple instead of the expected pink color. Gram-negative bacteria like E. coli usually appear pink because their cell walls are not able to retain the crystal violet dye after the decolorization step with alcohol; instead, they take up the counterstain, safranin. The color differentiation in the Gram stain process is due to differences in the cell wall structure of gram-positive and gram-negative bacteria.
The primary stain in the Gram stain process is crystal violet, which penetrates all bacterial cells. Then, Gram's iodine is used as a mordant to form a complex with the crystal violet, which is retained in gram-positive cells due to their thick peptidoglycan layer. The decolorization step involves alcohol, which dehydrates the peptidoglycan layer in gram-positive cells, trapping the dye within; however, in gram-negative cells, it dissolves the outer membrane and leaves the thinner peptidoglycan layer unable to retain the crystal violet, thus appearing pink after the counterstain.
Gram-positive bacteria, with their thick peptidoglycan layer, will appear purple after Gram staining, while gram-negative bacteria will appear pink due to the lack of retention of the crystal violet and absorption of the counterstain, safranin. Incorrect staining can lead to false results, such as blue E. coli, which might confuse the differentiation of bacterial species based on their cell wall characteristics.