Difference between Gram positive and Gram negative bacteria


Gram positive and Gram negative bacteria are the two forms of bacteria based on the Gram’ staining technique, developed by Hans Christian Grams for the identification of bacteria.

In 1884, Hans Christian Gram developed a different type of staining process based on the reaction of the cell walls with certain chemicals or dyes for the characterization and identification of bacteria.

Gram-positive bacteria are the bacteria with thick cell walls (multi-layered peptidoglycan layer) that interact with gram stain retaining the crystal violet color and stain purple. Some examples of gram positive bacteria are Staphylococcus aureus, Streptococcus pyogenes, Bacillus subtilis, and Streptococcus pneumonia.

Gram-negative bacteria are bacteria with thin, single-layered cell walls that cannot retain the crystal violet color due to no or very weak interaction with gram’s stain, and hence appear as light red or pink when observed under the microscope. Pseudomonas, Klebsiella, Proteus, Salmonella, Providencia, Escherichia, Morganella, Citrobacter, etc. are some of the examples of gram-negative bacteria.

Mostly, they differ in their cell wall composition. The two main characteristics that influence the cell wall structure’s capacity to retain the crystal violet stain employed in the Gram staining method are the thickness of the peptidoglycan layer and the existence or absence of an outside lipid membrane.

Cell wall of bacteria gram positive and negative

It is thought that gram positive bacteria have considerably larger peptidoglycans or murein, which account for the difference between the two types of bacteria.


Gram negative bacteria only have a thin layer of peptidoglycans (around 20%), but they also have an extra membrane called the outer cytoplasmic membrane.

Figure: Cell wall of gram-positive and gram-negative bacteria

Similarity between gram positive and gram negative bacteria

Both bacteria; gram positive and gram negative, have basal rings on their flagella or have flagella and capsules. These bacteria even have a cytoplasmic membrane and pilus. The cell wall between them has a peptidoglycan layer and provides structural support to the bacteria.

Gram positive and gram negative color

Gram-positive bacteria have a distinct purple color when observed under the biological microscope due to the thick peptidoglycans layer and porous layer whereas gram-negative bacteria have a red or pink color stain.


Gram stain procedure

1. First, the bacteria were fixed on the slide. 2. Flood the bacterial smear with staining solution i.e crystal violet and leave for 1 min 3. Gently rinse with tap water or distilled water 4. Flood the smear with iodine solution for 1 minute 5. The smear will now appear purple. 6. Decolorize the smear using alcohol and leave for 30 sec and wash with water. 7. Gently flood with safranin counterstain and leave for 30 sec and rinse with tap water. 8. Blot the slide dry and then observe under the microscope with oil- immersion.


Due to the presence of peptidoglycans layer, in their differences and structure, some bacteria are gram-positive and others are gram-negative. Gram positive are more susceptible to antibiotics and gram negative are more resistant to antibiotics. Likewise, gram positive are purple or blue in color and gram negative is pink or red in color.

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