Question

what can cause a false outcome with gram-positive bacteria? method? technique? the age? thr orgsnism itself? explain

Answer 1

• Gram staining procedure uses stains to distinguish Gram positive bacteria from Gram negative.
• The thicker peptidoglycan layer forms complex with primary stain crystal violet and mordant Gram’s Iodine, along with heat fixing. This stain is retained even during the decolorization phase with alcohol.
• While cells with thinner peptidoglycan layer cannot retain primary stain, after decolorization and take up the secondary stain Safranin.

However false results may be obtained in certain cases:

1. Method:

• Methodical mistakes may lead to false result.
• Like lomger heat fixing may cause damage to cell wall structure causing primary stain to be lost during destaining.

2. Technique:

• Certain techniques may cause destaining of Gram positive cells.
• Like using alcohol for longer period or using stronger decolorising agents.

3. Age:

• Certain bacteria may loose the integrity of the peptidoglycan layer with age.
• This may lead to false Gram staining results.

4. BActerial nature.

• Some bacteria may have structure around cell walls, like capsule, that may lead to variation in Gram staining.
• Bacteria like Mycobacterium cell has a waxy coat, containing mycolic acid, which make it impervious to Gram staining. Their cell wall contains high proportion of high-molecular-weight lipids. They may unpredictably acquire Gram positive or negative characteristics.