Bacteria (Gram Negative Bacillus)
In a previous post I discussed the Quellung Reaction which is used to visualize capsules produced by the gram positive organism Streptococcus pneumoniae (pneumococcus), There I mentioned that organisms other than the pneumococcus are capable of producing a capsule that surrounds the bacterial cell. Bacteria strains of E.coli, Klebsiella pneumoniae, Haemophilus influenzae, Neisseria meningitidis, Pseudomonas aeruginosa and Cryptococcus neoformans are capable of capsular production.
Bacterial capsules, often referred to as the Glycocalyx, are generally composed of polysaccharides with a high percentage of water, however other materials such as polypeptides and hyaluronic acid may be components.
Capsules may be beneficial to the cell because;
- Capsules may provide a virulence factor by protecting the bacterial cell from destruction by phagocytes, thereby allowing it to survive and spread.
- The high water content may protect the cell from dehydration, from hydrophobic agents such as detergents and their own parasitization by bacteriophages (viruses). The high water content also makes the capsule difficult to stain with conventional microbiological stains.
Below is a routine gram stain of a patient’s sputum specimen. On examination the presence of numerous coliform-like gram negative bacilli were noted. Occasionally, where conditions were just right, the dark red (gram negative) colour of the bacterial cell and the varying shades of the sputum can be seen separated by a small zone. This is the bacterial capsule and its presence in this case, along with the size of the bacterial cell, would provide immediate suspicion that this organism may be Klebsiella pneumoniae. Subsequent culture confirmed the initial suspicion of the organism responsible for this respiratory infection,
Here the capsule is visualized without using antibodies as with the Quellung reaction with Streptococcus pneumoniae. Antibodies against K. pneumoniae could be produced to enhance appearance of the capsule. These may be manufactured and utilized in research however they offer little to the diagnosis and management of K. pneumoniae in the clinical setting, The cost would preclude their use in routine diagnosis.
Streptococcus pneumoniae’s capsule may also be seen in the gram stain where conditions are right however, quick and confident diagnosis justifies obtaining and using Quellung antisera.
(1) Negative staining is a technique where you don’t stain what you wish to visualize but rather you stain everything around it thereby enhancing the contrast and making the object more visible.
(2) An example of deliberate negative staining - using Nigrosin (or India Ink) to visualized the capsule surrounding the yeast Cryptococcus neoformans. In a darkly stained field, the yeast cells would be seen as a natural or neutral colour, with a definite clear zone seen around the outside of the cellular wall.