Differences Between Gram-Positive and Gram-Negative Bacteria

By Prof Mariam Mirambo

Gram Positive Bacteria

Gram-positive bacteria are bacteria with thick cell walls. In a Gram stain test, these organisms yield a positive result. The test, which involves a chemical dye, stains the bacterium’s cell wall purple.

Gram-positive bacteria take up the crystal violet stain used in the test, and then appear to be purple-coloured when seen through an optical microscope. This is because the thick peptidoglycan layer in the bacterial cell wall retains the stain after it is washed away from the rest of the sample, in the decolorization stage of the test.

The most common gram negative bacteria include:

  • Staphylococcus epidermidis
  • Streptococcus pneumoniae
  • Streptococcus pyogenes
  • Streptococcus agalactiae
  • Bacillus anthracis
  • Corynebacterium diphtheria 
  • Listeria monocytogenes

Gram Negative Bacteria

Gram-negative bacteria are bacteria that do not retain the crystal violet stain used in the gram-staining method of bacterial differentiation.They are characterized by their cell envelopes, which are composed of a thin peptidoglycan cell wall sandwiched between an inner cytoplasmic cell membrane and a bacterial outer membrane.

Gram-negative bacteria can cause many serious infections, such as pneumonia, peritonitis (inflammation of the membrane that lines the abdominal cavity), urinary tract infections, bloodstream infections, wound or surgical site infections, and meningitis

The most common Gram Negative Bacteria Include:  

  • Escherichia,
  • Klebsiella
  • Pseudomonas
  • Serratia 
  • Enterobacter

Cell Wall Anatomy

Key Differences

Thickness of cell wallCell wall of gram positive bacteria is 20 to 80 nanometer thickCell wall of gram negative bacteria is 8 to 10 nanometer thin
Cell Wall

Peptidoglycan layer
Smooth single layered cell wall

Peptidoglycan layer is thick up to 90% and are multilayered
Wavy double-layered cell wall

Peptidoglycan layer is thin and are often single layered
Teichoic acidTeichoic acid is present within the cell wallTeichoic acid is absent
Periplasmic spaceThere is no presence of periplasmic spacePeriplasmic space is present between the region of cytoplasmic and outer membrane
Outer membraneOuter membrane is absent but are surrounded by thick layers of peptidoglycanOuter membrane is mostly present with a unique component, lipopolysaccharide (LPS) with proteins and phospholipids
RigidityGram positive cell wall are more rigid due to cross linkage of peptidoglycan chainsGram negative cell wall are less rigid
ElasticityCell wall is less elasticCell wall is more elastic
Color observed on Gram’s stainingPurple or blue colored is observed when examined under the microscope after Gram’s staining which is due to the retaining of crystal violet dye by the cell wall of gram positive bacteria.
Pink or red colored observed when examined under the microscope after Gram’s staining which accepts the counterstain safranin by the cell wall of gram negative bacteria after decolourization step.
Morphology of bacteriaCocci or spore forming rods Gram positive cocci: Streptococcus pneumonia, Staphylococcus aureus, Enterococcus Gram positive rods: Corynebacterium diphtheria, Listeria monocytogenes, Bacillus anthracis, Erysipelothrix rhusiopathiaeCocci or non-spore forming rods Gram negative cocci: 
Neisseria gonorrhoeae, Neisseria meningitides, Moraxella catarrhalis, Haemophilus influenza Gram negative rods: Escherichia coli, Yersinia pestis, Salmonella Typhi/Paratyphi, Salmonella, Shigella, Proteus
PorinsNot presentPresent in outer membrane that helps in the diffusion of small hydrophilic molecules
Mycolic acidPresent in some bacteria like Mycobacterium tuberculosis that makes up to 60% of the cell wallAbsence of mycolic acid
MesosomeMesosomes are more prominentMesosomes are less prominent
Lipoprotein contentAcid fast bacteria have lipids connected to peptidoglycan layer therefore,  lipoprotein content is lowDue to presence of outer layer, lipoprotein content is high
Toxins producedExotoxinsEither exotoxins or endotoxins
Flagellar structureTwo rings in basal body, one in the peptidoglycan layer and one in the plasma membraneFour rings in basal body: L ring in the lipopolysaccharide layerP ring in the peptidoglycan layerM ring embedded in the plasma membraneS ring directly connected to the plasma membrane
Lipid contentVery low and are imbedded in the cytoplasmic membraneUp to 20 to 30% in the outer membrane
Endospore formationFormation of endospore during unfavourable conditions (eg: Bacillus cereus, Bacillus anthracis, Bacillus thuringiensis Clostridium botulinum, Clostridium tetani)Usually endospores are not formed
Cell wall disruption by lysozymeGram positive bacteria are more susceptible towards the lysozyme action because of their cell wall content up to 90% peptidoglycan layerGram negative bacteria are more resistant towards lysozyme due to its low content of peptidoglycan layer
Resistance to physical disruptionGram positive bacteria are more resistant to physical disruptionGram negative bacteria are less resistant
Resistance to antibioticsMost gram positive bacteria are sensitive to antibioticsThey are more resistant to antibiotics because of the presence of large impermeable cell wall
Antibiotics used to treat bacterial infectionsVancomycin, teicoplanin, Quinupristin, Oxazolidinones, Daptomycin, Telavancin, CeftarolineCephalosporins, fluoroquiolones, aminoglycosides, imipenem, broad spectrum penicillins with or without β-lactamase inhibitors
Resistance to dryingHighly resistant towards drying due to their thicker cell wallLow resistant towards drying
Resistance to disinfectantsMore susceptible to cleaning agents due to lack of an outer membraneLess susceptible to cleaning agents due to the reduced permeability of the double membrane
Nutritional RequirementsNutritional requirement is relatively complexNutritional requirement is relatively simple
Inhibition by basic dyes like salts, chloridesHighly inhibited by basic dyesLow inhibited by basic dyes
Susceptibility to anionic detergents like Sodium n-dodecyl benzene sulphonate and Sodium Lauryl sulphateHighly susceptible towards anionic detergentsLess susceptible towards anionic detergents
Murein contentMurein content in cell wall is high about 70 to 80%Murein content in cell wall is low about 10 to 20%
Variety of amino acids in cell wallFew variety of amino acids are found in the cell wall of gram positive bacteriaSeveral variety of amino acids are found in the cell wall of gram negative bacteria
RNA:DNA ratioIn gram positive bacteria, the ratio of RNA:DNA is 8:1In gram negative bacteria, the ratio of RNA:DNA is 1-3:1 which is almost 1
PathogensOnly six gram positive genera are pathogenic to humansMost gram negative bacteria are pathogenic to humans
DiseasesPharyngitis, cellulitis, scarlet fever, necrotizing fasciitis, glomerulonephritis, rheumatic fever.Brucellosis, Cholera. Plague, Typhoid fever, urinary tract infections (UTI), bloodstream infections, meningitis.