Exploring Gram Positive vs Gram Negative Bacteria Differences

Gram Positive vs Gram Negative: An Overview

Gram positive and Gram negative refers to the results of a laboratory test called a Gram stain or Gram staining. This test identifies harmful bacteria in samples of bodily fluids, such as sputum, blood, and urine, which are obtained from people with a suspected infection.

The Gram stain is a quick test done in the microbiology laboratory. The results of this test can tell healthcare providers whether bacteria are present, and if so, what are the broad types.

This information can help your healthcare provider decide on further tests and treatment options. Certain antibiotics are more effective in treating Gram-negative infections, while others work better for Gram-positive infections.

History and Development of the Gram Stain

The Gram stain is named after Hans Christian Gram, a Danish microbiologist who first used the method in 1882 to identify organisms causing pneumonia. Today, Gram staining is a widely used technique in microbiology laboratories and is often the first test done on a patient’s sample.

Relevance in Medical and Scientific Contexts

Whether bacteria are “Gram-negative” or “Gram-positive” depends on the physical and chemical properties of their cell walls.

Gram staining is performed using methylene blue or crystal violet as the primary colors. When these stains or dyes are applied, different types of bacteria develop one of two color changes, either purple/blue or pink/red. Some bacteria retain the primary color (dye or stain) and appear purple/blue when examined under a microscope. They are called Gram-positive organisms. Some bacteria do not take up the primary color (dye or stain) and appear pink/red under a microscope. They are called Gram-negative organisms.

Note:

The Gram test does not tell healthcare providers which individual bacterial cells are present. Other tests such as a bacterial culture must be done to identify the specific type of bacterium present. In other words, Gram stains alone do not provide a diagnosis. They only provide a clue about the general type of bacteria present.
Some bacteria cannot be tested using Gram staining.
Most bacteria are Gram-positive or Gram-negative, but there are also Gram-variable bacteria that give irregular results on Gram staining (a mix of pink/red and purple/blue colors).
Gram-negative does not mean bacteria are absent. Gram-positive does not mean bacteria are present.
Gram-negative does not mean bad bacteria. Gram-positive does not mean good bacteria.

Differences in Cell Wall Structure

Role of Peptidoglycan Layer

Bacteria are single-celled organisms (round-shaped cocci or rod-shaped bacilli) with a protective cell wall. Peptidoglycan is a complex polymer (net-like structure of glycan chains cross-linked by short peptides). It is a key component of the bacterial cell wall. The amount of peptidoglycan in the bacterial cell wall determines the results of the Gram staining procedure.

Composition of Gram-Positive Cell Wall

In Gram-positive cells, the cell wall is about 90% peptidoglycan. The peptidoglycan takes up the Gram stain and causes the bacteria to appear blue/purple under the microscope.

Composition of Gram-Negative Cell Wall

In Gram-negative cells, the cell wall is only about 10% peptidoglycan, and the rest is lipid (fatty acids). As a result, these bacteria do not retain the Gram stain and appear pink/red under the microscope. However, Gram-negative bacteria have additional thin layers forming a protective outer membrane.

Staining Techniques and Results

The Gram Staining Process Explained

Gram stain tests are done on samples such as blood, urine, sputum, as well as fluid obtained from the abdominal cavity (ascitic fluid), lungs (pleural fluid), joints (synovial fluid), or brain and spinal cord (cerebrospinal fluid). A Gram stain test can also be done on swabs from the nose, throat, cervix, rectum, or wounds. The sample is collected in a sterile container to prevent contamination.

The underlying principle of Gram staining, as mentioned above, is that Gram-positive bacteria retain the crystal violet dye due to their high peptidoglycan content, whereas Gram-negative bacteria do not retain the stain due to their high lipid content in the cell wall.

The steps of the Gram stain procedure are listed below:

First, apply a crystal violet stain to the specimen.
Following the application of the primary stain, it is “fixed” using Gram’s iodine solution. This prevents easy dislodgement of the dye.
Next, a decolorizer (usually a solvent such as acetone or ethanol) is used to remove the dye.
All bacteria initially take up the dye or stain. However, the solvent is able to dissolve the dye in the lipid layer of Gram-negative microorganisms, causing them to lose the stain. In contrast, the solvent dehydrates Gram-positive microorganisms and causes the pores in the cell wall to close, causing them to retain the stain. The decolorization step is, therefore, critical in the Gram staining procedure because prolonged exposure to the solvent can cause both types of bacteria to lose the stain.
The final step in the Gram stain test is a counterstain with a fuchsin or safranin stain. This gives the decolorized Gram-negative bacteria a pink color and makes them easier to identify on Gram stain results.

Gram Positive Stain Characteristics

Gram-positive bacteria appear blue to purple under a microscope.

Gram Negative Stain Characteristics

Gram-negative bacteria appear pink to red under a microscope.

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Implications for Health and Disease

Antibiotic Treatment of Bacterial Infections

Gram-positive bacteria have a thick peptidoglycan cell wall that absorbs antibiotics, making them easier to kill.

Gram-negative bacteria have a thin peptidoglycan layer and a protective outer membrane that makes them more difficult to kill.

Antibiotic Resistance in Bacteria

Gram-negative bacteria are more likely to develop antibiotic resistance due to the presence of an outer membrane. Mutations or changes in this outer membrane help Gram-negative bacteria escape being killed by antibiotics, thus creating antibiotic resistance. Gram-positive bacteria lack an outer layer, which makes them less resistant to antibiotics compared to Gram-negative organisms.

Risks and Contagiousness

As mentioned, Gram-negative bacteria have a protective outer membrane that makes them harder to kill. They are also more likely to develop antibiotic resistance compared to Gram-positive bacilli.

Worldwide, Gram-negative infections cause serious infections with high morbidity and mortality rates. Gram-negative bacterial infections are a significant public health problem. Patients admitted to hospital are at highest risk of serious bacterial infections caused by Gram-negative microorganisms.

Examples and Classification

Common Types of Gram-Positive Bacteria

Staphylococcus species: Methicillin-resistant staphylococcus aureus (MRSA) infections, pneumonia, and toxic shock syndrome
Streptococcus pneumoniae: Pneumonia.
Group A Streptococcus: Strep throat.
Clostridium species: Food poisoning
Clostridium botulinum: Botulism (severe food poisoning).
Listeria species: Food poisoning.
Corynebacterium species: Diphtheria.
Enterococci: UTIs, blood infections, infective endocarditis, abdominal infections, and meningitis.

Common Types of Gram-Negative Bacteria

Escherichia coli: Urinary tract infections (UTIs) and food poisoning.
Salmonella: Typhoid fever and food poisoning.
Klebsiella: Wound infections, surgical site infections, bloodstream infections, bacterial meningitis, and pneumonia.
Morganella morganii: UTIs, pneumonia, sepsis, wound infections, musculoskeletal infections, pericarditis, and brain and spinal cord.
Pseudomonas species: Blood infections, ear infections, eye infections, lung infections, skin infections, and urinary tract infections.
Aeromonas: Bacterial meningitis, osteomyelitis, eye infections, joint infections, and respiratory infections after "near drownings."
Legionella pneumoniae: Legionnaires’ disease.
Neisseria gonorrheae: Gonorrhea.
Proteus species: UTIs.

Clinical Relevance and Implications

Both Gram-positive and Gram-negative bacterial infections are treated with antibiotics. Antibiotics cannot treat a viral or fungal infection.

Penicillin, cloxacillin, and erythromycin can treat 90% of Gram-positive infections.

A bacterial infection caused by Gram-negative bacilli can be more challenging to treat due to antibiotic resistance, including multidrug resistance (bacteria that are resistant to many antibiotics). Some of the antibiotics that may be used to treat a Gram-negative infection include polymyxins, aminoglycosides, carbapenems, and fosfomycin.

Frequently Asked Questions

What's The Difference Between Gram-Negative and Gram-Positive?

Two key features that differentiate between Gram-negative and Gram-positive bacterial cells are in the cell wall. The physical and chemical features of the bacterial cell wall cause the bacteria to either retain or lose the Gram stain (a special dye) during a Gram stain test. Gram-positive bacteria retain the dye and appear blue/purple. Gram-negative bacteria lose the dye and appear pink/red under the microscope. In addition, Gram-negative bacteria have an outer membrane that makes them harder to kill.

Is Gram-positive or Gram-Negative More Harmful?

The Centers for Disease Control and Prevention (CDC) and World Health Organization (WHO) categorize Gram-negative as more harmful. This is because although the cell wall thickness of Gram-positive bacteria is thicker, Gram-negative bacteria have an outer protective cell membrane made of thin layers of phospholipids and lipopolysaccharides. This outer cell wall makes Gram-negative bacteria harder to kill and more resistant to antibiotics.

Why Is Gram-Negative More Resistant To Antibiotics?

Gram negative bacteria are more resistant to antibiotics because they have a protective lipopolysaccharide-containing outer cell wall. Changes in this outer membrane allow these bacteria to develop antibiotic resistance. Many Gram negative microorganisms have developed multidrug resistance (inability to be killed by the multiple commonly prescribed antibiotics).

Is Gram-Negative Bacteria Contagious?

Both Gram negative and Gram positive bacteria are contagious. Bacterial infections spread through respiratory droplets, skin-to-skin contact, sexual contact, contaminated objects and surfaces, contaminated food and water, and insect bites, and other methods of transmission.

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