Why is it important to know the difference between Gram positive and Gram negative cell walls?

The terms Gram positive and Gram negative are commonly used to describe bacteria. The main difference between the two is the structure of their cell wall which changes their susceptibility to different antibiotics. The separation also loosely fits the location of these organisms in the body – Gram negative organisms predominate in the bowel (eg. E. coli and other coliforms, Bacteroides species etc) whilst Gram positives (Staphs. and streptococci) predominate on the skin, upper respiratory tract and oropharynx. Gram staining (see example Gram stains) was invented in the 1880s by Danish scientist Hans Christian Gram and takes several steps. A Gram positive organism holds onto the first stain and appears a purple-blue colour, whilst a Gram negative organism holds the second stain and appears pink. A bacterium’s ability to hold onto a stain is dependent on the structure of their cell wall.

Inhaltsverzeichnis Show

Pathogenesis in humans
Gram positive Cocci
Commercial uses of non-pathogenic Gram-positive bacteria
Gram-indeterminate and Gram-variable Bacteria

A Gram positive organism lacks an outer (LPS) membrane but has a thick layer of peptidoglycan and no LPS outer membrane. This facilitates access of cell-wall active antibiotics (eg. penicillin/betalactam or vancomycin-type antibiotics) to their site of action (the peptidoglycan).

In coliform-type Gram negative bacteria such as E. coli, these antibiotics have to traverse the LPS layer via porin (channel) proteins. The ability to transfer through these channels is dependent on the charge and shape of the antibiotic. For instance benzylpenicillin cannot cross whereas ampicillin can, causing intrinsic antibiotic resistance to the former drug. Porins may be modified or removed via genetic changes (via mutation usually) that occur which then potentially alters the resistance character of the organism. Efflux proteins may also be present in Gram negative cell walls. These are relevant to antibiotics that act intracellularly and pump them out of the cell, lowering the intracellular antibiotic concentration to a level where the antibiotic becomes inactive.

Image from: http://static.diffen.com/uploadz/0/0e/cell-wall-gram-bacteria.png

Image from: https://en.wikipedia.org/wiki/File:Gram_stain_01.jpg : Microscopic image of a Gram stain of mixed Gram-positive cocci (Staphylococcus aureus ATCC 25923, purple) and Gram-negative bacilli (Escherichia coli ATCC 11775, red). Magnification:1,000.

Medically Reviewed by Dan Brennan, MD on May 16, 2021

There are many ways of classifying bacteria. One method is based on the cell membrane. In 1884, a bacteriologist named Christian Gram created a test that could determine if a bacterium had a thick, mesh-like membrane called peptidoglycan. Bacteria with thick peptidoglycan are called gram positive. If the peptidoglycan layer is thin, it's classified as gram negative.

A gram-positive bacillus doesn't have an outer cell wall beyond the peptidoglycan membrane. This makes it more absorbent. Its peptidoglycan layer is much thicker than the peptidoglycan layer on gram-negative bacilli. Gram-positive bacilli are shaped like rods. They can also be characterized based on whether they form spores and whether they need oxygen to survive.

Resident flora. Most gram-positive bacilli live harmlessly on your body without causing problems. These are called resident flora. The can be found in the following places on your body:

Skin
Nose
Mouth
Throat
Vagina

Some types of gram-positive bacilli are pathogenic, which means they cause diseases in people. Some of the major gram-positive bacteria that cause diseases include:‌

Anthrax. This is a potentially fatal infection that usually affects the skin or lungs but can rarely infect the gastrointestinal tract as well. Anthrax is a spore-forming type of bacilli. It has the potential to be used as a biological weapon because its spores can be spread in the air and be inhaled. The spores can live for decades and are not easily killed.‌

Diphtheria. This is a contagious infection of the upper respiratory tract. Vaccination can prevent this infection. It's rare in developed countries.

Enterococcal Infections. These infections can be caused by bacteria known as enterococci. There are over 17 different species. They live in your intestines, where they don't normally cause problems. If they move to another part of your body, such as your blood, heart valves, or skin, they can cause serious infections.

Erysipelothricosis. This is a skin infection that usually happens when you get a scrape or puncture wound while handling infected animals. The main symptom is a hard, purplish-red rash.

Listeriosis. This is an infection caused by eating contaminated food. It causes fever, chills, nausea, vomiting, and diarrhea.

Gram-positive bacilli infections are treated with antibiotics. Penicillin, cloxacillin, and erythromycin treat over 90% of gram-positive bacteria.

‌However, antibiotic resistance is becoming a serious problem with gram-positive infections. New drugs are being developed to help with this problem. Antibiotics should only be used when you absolutely need them. Infection control standards need to be followed closely to prevent the development and spread of antibiotic-resistant infections.

Gram-negative bacteria have a hard, protective outer shell. Their peptidoglycan layer is much thinner than that of gram-positive bacilli. Gram-negative bacteria are harder to kill because of their harder cell wall. When their cell wall is disturbed, gram-negative bacteria release endotoxins that can make your symptoms worse.

Gram-negative bacteria can cause many serious infections, including:

Gram-negative infections are most common in hospitals. Your risk increases the longer you stay in the hospital. Other things that increase your risk of developing a gram-negative infection include:

Gram-negative bacteria have high resistance to antibiotics. They are one of the most serious public health issues in the world. Gram-negative bacteria have the ability to cause a lot of diseases in humans. They can reach almost all of the organ systems. Your doctor may need to try several antibiotics to beat the infection. Older antibiotics may work better.

Antibiotic resistance is made worse by the overuse of antibiotics. You can take the following steps to help control and prevent antibiotic resistance:

Only use antibiotics prescribed by your healthcare provider.
Follow your healthcare provider's advice about antibiotic use.
Don't use or share leftover antibiotics.
Wash your hands.
Practice safe sex.
Keep your vaccines up to date.
Avoid contact with people who are sick when you can.
Prepare your food safely.

Danish scientist Hans Christian Gram devised a method to differentiate two types of bacteria based on the structural differences in their cell walls. In his test, bacteria that retain the crystal violet dye do so because of a thick layer of peptidoglycan and are called Gram-positive bacteria. In contrast, Gram-negative bacteria do not retain the violet dye and are colored red or pink. Compared with Gram-positive bacteria, Gram-negative bacteria are more resistant against antibodies because of their impenetrable cell wall. These bacteria have a wide variety of applications ranging from medical treatment to industrial use and Swiss cheese production.

Microscopic view of dental plaque, showing Gram-positive (purple) and negative (red) bacteria

In a Gram stain test, bacteria are washed with a decolorizing solution after being dyed with crystal violet. On adding a counterstain such as safranin or fuchsine after washing, Gram-negative bacteria are stained red or pink while Gram-positive bacteria retain their crystal violet dye.

This is due to the difference in the structure of their bacterial cell wall. Gram-positive bacteria do not have an outer cell membrane found in Gram-negative bacteria. The cell wall of Gram-positive bacteria is high in peptidoglycan which is responsible for retaining the crystal violet dye.

Gram-positive and negative bacteria are chiefly differentiated by their cell wall structure

The following videos demonstrate the staining of Gram-positive and negative bacteria respectively.

Pathogenesis in humans

Both gram-positive and gram-negative bacteria can be pathogenic (see list of pathogenic bacteria). Six gram-positive genera of bacteria are known to cause disease in humans: Streptococcus, Staphylococcus, Corynebacterium, Listeria, Bacillus and Clostridium. Another 3 cause diseases in plants: Rathybacter, Leifsonia, and Clavibacter.

Many gram-negative bacteria are also pathogenic e.g., Pseudomonas aeruginosa, Neisseria gonorrhoeae, Chlamydia trachomatis, and Yersinia pestis. Gram-negative bacteria are also more resistant to antibiotics because their outer membrane comprises a complex lipopolysaccharide (LPS) whose lipid portion acts as an endotoxin. They also develop resistance sooner:

A lot of Gram-negative bacteria, they come out of the box, if you will, resistant to a number of important antibiotics that we might use to treat them. We’re talking about agents with names like Acinetobacter, Pseudomonas, E. coli. These are bacteria that have historically done a very good job of very quickly developing resistance to antibiotics. They have a lot of tricks up their sleeves for developing resistance to antibiotics, so they’re a group of agents that can quickly become resistant, can pose major challenges to resistance. And what we’ve seen over the past decade is these Gram-negative agents becoming very rapidly more and more resistant to all of the agents that we have available to treat them.

Greater resistance of gram-negative bacteria also applies to a newly discovered class of antibiotics that was announced in early 2015 after a decades-long drought in new antibiotics. These drugs are not likely to work on gram-negative bacteria.

Structure of a gram-positive bacterial cell.

Gram positive Cocci

Bacteria are classified based on their cell shape into bacilli (rod shaped) and cocci (sphere shaped).Typical Gram-positive cocci stains include (pictures):

Clusters: usually characteristic of Staphylococcus, such as S. aureus
Chain: usually characteristic of Streptococcus, such as S. pneumoniae, B group streptococci
Tetrad: usually characteristic of Micrococcus.

Gram-positive bacilli tend to be thick, thin or branching.

Commercial uses of non-pathogenic Gram-positive bacteria

Many streptococcal species are nonpathogenic, and form part of the commensal human microbiome of the mouth, skin, intestine, and upper respiratory tract. They are also a necessary ingredient in producing Emmentaler (Swiss) cheese.

Non-pathogenic species of corynebacterium are used in industrial production of amino acids, nucleotides, bioconversion of steroids, degradation of hydrocarbons, cheese ageing, production of enzymes etc.

Many Bacillus species are able to secrete large quantities of enzymes.

Bacillus amyloliquefaciens is the source of a natural antibiotic protein barnase (a ribonuclease), alpha amylase used in starch hydrolysis, the protease subtilisin used with detergents, and the BamH1 restriction enzyme used in DNA research.
C. thermocellum can utilize lignocellulose waste and generate ethanol, thus making it a possible candidate for use in production of ethanol fuel. It is anaerobic and is thermophilic, which reduces cooling cost.
C. acetobutylicum, also known as the Weizmann organism, was first used by Chaim Weizmann to produce acetone and biobutanol from starch in 1916 for the production of gunpowder and TNT.
C. botulinum produces a potentially lethal neurotoxin that is used in a diluted form in the drug Botox. It is also used to treat spasmodic torticollis and provides relief for approximately 12 to 16 weeks.

The anaerobic bacterium C. ljungdahlii can produce ethanol from single-carbon sources including synthesis gas, a mixture of carbon monoxide and hydrogen that can be generated from the partial combustion of either fossil fuels or biomass.