Bacterial capsule

Many bacterial cells secrete some extracellular material in the form of a capsule or a slime layer. A slime layer is loosely associated with the bacterium and can be easily washed off, whereas a capsule is attached tightly to the bacterium and has definite boundaries. It is gelatinous layer covering the entire bacterium. It usually consists of polysaccharides, but can be composed of other materials (e.g., polypeptide (D-glutamic acid) in B. anthracis). The size and constituency of the capsule varies with species and strains

The capsule—which can be found in both Gram-negative bacteria and Gram-positive bacteria—should not be confused with the second lipid membrane (or bacterial outer membrane), which contains lipopolysaccharides and lipoproteins and is found only in Gram-negative bacteria.

Importance of Bacterial Capsule

•            Virulence determinants: Capsules are antiphagocytic. They limit the ability of phagocytes to engulf the bacteria. If a pathogenic bacteria lose capsule (by mutation), they won’t be able to cause disease (i.e. changes to nonpathogenic bacteria).
• .        Identification of bacteria: a. using specific antiserum against capsular polysaccharide.  E.g. Quellung reaction b. Colony characteristics in culture media: Capsulated organisms forms mucoid colonies
• .        Development of Vaccines: Capsular polysaccharides are used as the antigens in certain vaccines. E.g. the purified capsular polysaccharides of 23 types of S. pneumoniae are present in current vaccine.
• .        Initiation of infection: Capsules helps the organism to adhere to host cells.
• .        Most capsules are hydrophilic (“water-loving”) and may help the bacterium avoid desiccation (dehydration) by preventing water loss.
• .       Capsules can protect a bacterial cell from ingestion and destruction by white blood cells (phagocytosis). While the exact mechanism for escaping phagocytosis is unclear, it may occur because capsules make bacterial surface components more slippery, helping the bacterium to escape engulfment by phagocytic cells.

Examples of Capsulated bacteria:

The capsule is found most commonly among Gram-negative bacteria:

·         Escherichia coli (in some strains)

·         Neisseria meningitidis^[4]

·         Klebsiella pneumoniae^[5]

·         Haemophilus influenzae^[6]

·         Pseudomonas aeruginosa^[7]

·         Salmonella^{[citation needed]}

However, some Gram-positive bacteria may also have a capsule:

·         Bacillus megaterium for example, synthesizes a capsule composed of polypeptide and polysaccharides.

·         Streptococcus pyogenes synthesizes a hyaluronic acid capsule.

·         Streptococcus pneumoniae

·         Streptococcus agalactiae produces a polysaccharide capsule of nine antigenic types that all contain sialic acid (Ia, Ib, II, III, IV, V, VI, VII, VIII).

·         Staphylococcus epidermidis

The yeast Cryptococcus neoformans, though not a bacterium, has a similar capsule.

Capsules too small to be seen with an ordinary microscope, such as the M protein of Streptococcus pyogenes, are called microcapsules.

A common mnemonic used to remember some encapsulated pathogens is:

"Even Some Super Killers Have Pretty Nice Big Capsules"

Escherichia coli, Streptococcus pneumoniae, Salmonella, Klebsiella pneumoniae, Haemophilus influenzae, Pseudomonas aeruginosa, Neisseria meningitidis,Bacteroides fragilis, and the yeast Cryptococcus neoformans.

Capsule Staining Technique by Maneval’s Method.

Maneval’s staining method is a negative staining method.

Requirement

1.       1 % Congo red solution.

2.       Maneval’s stain.

3.       Capsulated cell culture.

Procedure

1.       Take a loopful of capsulated cell suspension on a clean grease free slide.

2.       Add a drop of 1 % Congo red solution in the suspension and then spread it gently on the slide to form a smear.

3.       Allow the suspension to air dry and do not heat fix it.

4.       Flood the slid with Maneval’s stain and keep it for 2 minutes.

5.       After 2 minutes discard excess stain and do not water wash the slide.

6.       Allow the slide to air dry and observe under oil immersion objective.

Mechanism

• As we know capsules are non-ionic in nature so they do not get stain easily.
• By using the negative staining method demonstration of capsule becomes easy.
• So in this method first we use Congo red stain as it is acidic stain it stain the background in red colour.
• Further we use Maneval’s stain this stain has four important components and they are-

1. 10 % FeCl3 – It acts as chemical fixative.
2. 5 % phenol – It increases penetration power of stain.
3. Acid fuchsin – It is a basic dye it stains the cell.
4. Acetic acid – It decreases the pH of smear to acidic side.

• Due to acetic acid pH is shifted to acidic side and due to which Congo red stain turns red to blue in colour that is why the colour of background changes from red to blue in colour.
• Due to acid fuchsin the cell get stained in pink colour.
• It creates a clear contrast in between cell, capsule and background and due to which capsule can be detected easily.

Note

Congo red stain is an indicator dye at acidic pH it appears blue in color while at neutral pH it appears red in color.