Gram Positive Bacteria

Bacteria can be classified broadly into two different groups: gram-positive and gram-negative bacteria. These classifications are based on their appearance after gram staining, a four-step procedure invented by scientist Hans Christian Gram in the 1880s. Gram-positive bacteria have certain characteristics that we must know and that scientists use to help develop medications that have gram-positive anti-bacterial activity.

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Table of contents
    • What are Gram-positive bacteria?
    • Gram-positive bacteria: colour
    • Difference between Gram-positive and Gram-negative bacteria
      • Gram-positive bacteria cell wall
    • Examples of Gram-positive bacteria
    • List of diseases caused by Gram-positive bacteria

    What are Gram-positive bacteria?

    Gram-positive bacteria are a type of bacteria with a thick cell wall made of peptidoglycans which turns purple after Gram staining.

    It's as simple as that: the Gram-positive/Gram-negative classification is just a way to differentiate bacteria in terms of their reaction to a really simple and common staining method, the Gram stain. That reaction, though, has a biological reason behind it.Gram-positive and Gram-negative bacteria are morphologically different: Gram-positive bacteria have a thick cell wall, while Gram-negative bacteria have a thin cell wall and a lipid outer membrane.

    Gram-positive bacteria colour

    Gram-positive bacteria appear purple after they go through the gram stain procedure. Gram staining involves four steps:

    1. Primary staining with crystal violet

    2. Fixing the sample

    3. Decolorizing with alcohol wash

    4. Counterstaining with safranin

    The four substrates that are used during each stage are, respectively:

    1. Crystal violet as the primary stain - this turns bacteria purple.

    2. Iodine as the fixing agent - this doesn't change the bacteria's colour.

    3. Ethyl alcohol as the decolourizer - this washes out the purple colour of gram-negative bacteria, but doesn't change the colour of gram-positive bacteria

    4. Safranin as the counterstain - this turns the now colourless gram-negative bacteria pink, but cannot overwhelm the purple colour of gram-positive bacteria, so they are unaffected by safranin and stay purple.

    Gram-positive bacteria remain purple following all four steps of gram staining because of certain unique properties of their cell walls. We will explain how and discuss these properties below.

    Gram-positive bacteria cell wall features

    Gram-positive bacteria have a cell membrane comprised of a phospholipid bilayer. External to this membrane there are a few additional features. In order of closeness to the cell membrane, they are:

    • the periplasmic space,
    • the thick peptidoglycan cell wall, and
    • the teichoic acids and lipoteichoic acids (Fig. 1).

    Thick peptidoglycan cell wall

    Gram-positive bacteria have a cell wall comprised of peptidoglycan. As the name suggests, peptidoglycan is comprised of both amino acids (peptides) and sugars (glycans).

    Specifically, NAG (N-acetylglucosamine) and NAM (N-acetylmuramic acid) are the two amino acid sugars that make up the backbone of peptidoglycan.

    NAM can carry a chain of four amino acids, called a tetra-peptide side chain. NAG and NAM are cross-linked together, and tetra-peptide side chains are linked to each other as well, giving peptidoglycan its strong but porous structure (Fig. 2).

    This basic peptidoglycan structure is present in both gram-positive and gram-negative bacteria, but what differentiates the two? Well, in gram-positive bacteria this peptidoglycan cell wall is very thick.

    The thickness of the cell wall allows iodine, the fixing agent (also called a mordant) to properly affix crystal violet to the bacteria during gram staining. Also, the pores within the cell wall, with the addition of ethyl alcohol, cause dehydration of the cell, as well as shrinking and closing of the pores. This helps trap the purple colour from the crystal violet dye and prevents gram-positive cells from being decolourized by the alcohol wash.

    Lipoteichoic acids (and teichoic acids)

    Lipoteichoic acids, unlike peptidoglycans, are only seen in Gram-positive bacteria. You will not find lipoteichoic acids in gram-negative bacteria.

    Lipoteichoic acids and teichoic acids are virulence factors. Their exact methods of action are not yet completely known to scientists, however, we do know they are analogous to lipopolysaccharides (LPS) in gram-negative bacteria.

    They arise from the plasma membrane (lipoteichoic acids) or the peptidoglycan wall (teichoic acids) and stick out above the cell wall. Here, they can activate enzymes or illness-inducing cytokines, that increase the virulence and pathogenicity of the bacteria that contain them.

    Periplasmic space

    Gram-positive bacteria have a small periplasmic space between their plasma membrane and their cell wall. The most important thing to note here is that the periplasmic space of gram-positive bacteria is much smaller than that of gram-negative bacteria. In fact, you may not see periplasmic spaces mentioned for gram-positive bacteria in all sources, but you should be aware that it is there.

    Difference between Gram-positive and Gram-negative bacteria

    Gram-positive and Gram-negative bacteria have quite a few differences in structure. These differences help decide what organisms and what organs within those organisms they can infect. We will outline the differences between gram-positive and gram-negative bacteria below.

    • Difference #1: Gram-positive bacteria have thick peptidoglycan cell walls. Gram-negative bacteria on the other hand have thin peptidoglycan cell walls.
    • Difference #2: Gram-positive bacteria have lipoteichoic acids and teichoic acids. Gram-negative bacteria do not have either.
    • Difference #3: Gram-positive bacteria do not have lipopolysaccharides (LPS) on the outside of their cell walls, while gram-negative bacteria do.
    • Difference #4: Gram-positive bacteria do not have an outer membrane. They only have a single cell membrane, beneath their cell wall. Gram-negative bacteria do have an outer membrane.
    • Difference #5: Gram-positive bacteria are defined by their purple colour after the process of gram staining. Gram-negative bacteria are defined by their pink colour after gram staining.
    CharacteristicGram-positive (+)Gram-negative (-)
    Peptidoglycan cell wallThickThin
    Lipoteichoic and teichoic acidsYesNo
    Outer membraneNoYes
    Gram stain colourPurplePink
    Table 1. Differences between Gram-positive and Gram-negative bacteria.

    Examples of Gram-positive bacteria

    There are a number of Gram-positive bacteria that are relevant medically. These are the ones that can cause illness in humans and non-human animals.

    Typically, bacterial agents that can cause disease are classified by two things. The first is their Gram stain, and the second is their shape. We will show here genuses of Gram-positive bacteria with two different shapes: rod-shaped (bacilli) or spherical (cocci). Listeria is the exception to this, because it has an intermediate shape, in between rod and sphere.

    You will notice that sometimes the shape of the bacteria is depicted in the name (some genus names include the words bacillus or coccus), but not always.

    Gram-positive bacilli

    • Bacillus

    • Corynebacterium

    • Clostridium

    Gram-positive cocci

    • Staphylococcus

    • Streptococci

    • Enterococci

    Gram-positive coccobacilli

    • Listeria

    List of diseases caused by Gram-positive bacteria

    The bacilli and cocci that we listed above can all be quite virulent pathogens and can cause many diseases in human hosts. We will make a sample list of some of the diseases that can be caused by these bacteria.

    Bacillus - this genus has three important species: B. anthracis, B. cereus, and B. subtilis.

    • B. anthracis = causes anthrax.

      • This can present in different ways - with black ulcers on the skin, diarrhoea, vomiting, or a severe lung infection.

    • B. cereus = causes acute food poisoning.

    • B. subtilis = this is a model organism and is the main organism used to study gram staining.

      • B. subtilis is an ideal model for scientists to observe how gram-positive rods, in general, look and behave.

    Clostridium - this genus has important species including C. botulinum, C. tetani, and C. perfringens.

    • C. botulinum = causes paralysis.

      • One can get infected by either food poisoning or by the bacilli entering a wound. Used to medically to make Botox!

    • C. tetani = causes tetanus.

      • You get immunizations to prevent this.

    • C. perfringens = causes massive wound necrosis.

    Streptococcus = this genus has important species, including S. pyogenes and S. pneumonia.

    • S. pyogenes = causes pharyngitis and skin infections.

      • The historically famous "scarlet fever" is caused by S. pyogenes.

    • S. pneumonia = causes pneumonia.

    Listeria = this genus has one very important species, capable of causing severe disease in human beings of all ages - L. monocytogenes.

    • L. monocytogenes can cause several illnesses, including:

      • Sepsis - this means widespread and highly dangerous accumulation of bacteria and bacterial products in the blood.

      • Pneumonia

      • Brain infections like meningitis and encephalitis.

      • Listeriosis - this is a food-borne listeria infection that happens to people mostly after consumption of affected meat and dairy products, like deli cold cuts or soft cheeses or milk.

        • Listeriosis symptoms include fever, muscle aches, vomiting, diarrhoea, fatigue, confusion, and more.

        • Pregnant women, elderly people, and people with weakened immune systems are at the highest risk.

    Gram-Positive Bacteria - Key takeaways

    • Gram-positive bacteria are purple after gram staining.
    • Gram-positive bacteria have thick peptidoglycan cell walls.
    • Gram-positive bacteria have lipoteichoic and teichoic acids, but do not have lipopolysaccharides.
    • Gram-positive bacteria do not have outer membranes.
    • There is a wide range of Gram-positive bacteria.
    Frequently Asked Questions about Gram Positive Bacteria

    What are Gram-positive bacteria?

    Gram-positive bacteria are bacteria that appear purple after they go through the Gram stain procedure.

    What colour are Gram-positive bacteria?

    Gram-positive bacteria do not have colour under the microscope. However, when they are stained through the Gram staining method, they appear purple under the microscope.

    Do Gram-positive bacteria have an outer membrane?

    No, Gram-positive bacteria do not have an outer membrane. They have a very thick cell wall made of peptidoglycans.

    What are the three Gram-positive bacteria?

    There are many Gram-positive bacteria. Some examples are some Staphylococci (S. aureus), some Streptococci (S. pneumoniae), or some Bacilli (B. subtilis).

    Why do Gram-positive bacteria stain purple?

    The crystal violet dye used for Gram staining can bind to the peptidoglycans present in the cell wall of bacteria. If the cell wall is thick enough, despite adding a decolourizer to the bacteria during the Gram staining, the crystal violet will remain in the cell wall, turning it purple to the human eye.

    Do Gram-positive bacteria produce endotoxins?

    No, Gram-positive bacteria do not produce endotoxins. This is because they do not have an outer membrane, where the endotoxins would be presented.

    How does penicillin work against gram-positive bacteria?

    Penicillin inhibits the synthesis of peptidoglycans, the molecules that form the cell wall of Gram-positive bacteria. Without the cell wall, the bacteria are much more fragile and are more easily accessible to the immune system.

    Do Gram-positive bacteria have flagella?

    Yes, Gram-positive bacteria have flagella to allow movement.

    Test your knowledge with multiple choice flashcards

    Are gram positive bacteria ever pink? Yes or No.

    True or False: Lipoteichoic acids are present on the outer surface of all kinds of bacteria.

    True or False: Lipopolysaccharides are the main source of virulence of gram-positive bacteria 


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