Bile solubility test is a biochemical experiment used to distinguish Streptococcus pneumoniae from other alpha-hemolytic streptococci. It is based on the idea that bile salts (e.g., sodium deoxycholate) lyse S. pneumoniae cells by activating an autolytic enzyme, while other alpha-hemolytic streptococci are unaffected and remain intact.
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What is Bile Solubility Test ?
Bile solubility test is a biochemical experiment used to distinguish Streptococcus pneumoniae from other alpha-hemolytic streptococci by determining their capacity to lyse in the presence of bile salts such as sodium deoxycholate. This test uses the fact that S. pneumoniae has an autolytic enzyme that is activated by bile salts, resulting in cell lysis, but other alpha-hemolytic streptococci do not lyse and stay intact.
Principle Bile Solubility Test
Bile Salts Action
The test takes use of the effect of bile salts on the bacterial cell wall. Bile salts reduce the surface tension of the bacterial cell wall, causing lysis in certain bacterial species.
Autolytic Enzymes
Streptococcus pneumoniae generates autolytic enzymes (autolysins), which are triggered by bile salts. These enzymes destroy bacterial cell walls, resulting in cell lysis.
Lysis Observation
The test involves evaluating if bacterial cells lyse (dissolve) when exposed to bile salts.
Procedure of Bile Solubility Test
Preparation
A suspension of the bacterial culture is made in a sterile saline solution.
Addition of Bile Salts
Sodium deoxycholate (bile salt solution) is added to the bacterial suspension.
Incubation
The mixture is incubated at 35-37°C for 10-30 minutes.
Observation
The tube is next tested for lysis. If the suspension clears, the test results are positive. If the suspension stays turbid, the test results are negative.
Types And Result of Bile Solubility Test
Tube Bile Solubility Test
Prepare Bacterial Suspension:
Using a sterile inoculating loop, transfer a few test organism colonies to a test tube containing 1 mL of sterile saline.
Mix carefully to get a uniform suspension.
Addition of Bile Salt:
Divide the bacterial suspension into two portions.
In a test tube, add an equal volume (0.5 mL) of 2% sodium deoxycholate solution to one part of the suspension.
Control Tube: Add an equivalent volume (0.5 mL) of sterile saline to the other half of the solution (control).
Incubation:
Incubate both tubes at 35-37°C for 10-30 minutes.
Observation:
Check the tubes for turbidity changes.
Positive Result: The test tube containing bile salt clears, demonstrating bacterial cell lysis (Streptococcus pneumoniae).
Negative Result: The test tube is still turbid, indicating no lysis.
Plate Bile Solubility Test
Procedure:
Prepare plate:
To create isolated colonies, streak the test organism onto a blood agar plate.
Application of Bile Salt:
Apply a few drops of the 2% sodium deoxycholate solution straight to a well-isolated colony on the blood agar plate.
Incubation:
Allow the dish to stand at room temperature for 15–30 minutes.
Observation:
Inspect the treated colonies for disintegration.
Positive Result: The bile salt-treated colony dissolves and is no longer visible (indicating Streptococcus pneumoniae).
Negative Result: The colony stays intact and visible (indicating that other alpha-hemolytic streptococci exist).
Comparison of Tube and Plate Tests
Tube Test:
More quantitative and exact.
Allows for better observation of turbidity variations.
Adds more stages and materials.
Plate Tests:
Simpler and speedier.
Suitable for direct application to culture plates.
Gives a visual assessment of colony breakup.
Applications and Considerations:
Both tests are primarily used to distinguish Streptococcus pneumoniae from other alpha-hemolytic streptococci.
It is critical to undertake quality control using recognized positive and negative control strains.
The decision between tube and plate tests is based on the laboratory’s resources, the nature of the samples, and the necessity for precision against speed.
Uses of Bile Solubility Test
It is the most typically used to distinguish Streptococcus pneumoniae from other alpha-hemolytic streptococci, notably those of the Viridans group.
Streptococcus pneumoniae is bile soluble, which means that it lyses in the presence of bile salts, whereas Viridans streptococci are bile insoluble.
Frequently Asked Question (FAQ)
What is Bile Solubility Test ?
It is a biochemical experiment used to distinguish Streptococcus pneumoniae from other alpha-hemolytic streptococci by determining their capacity to lyse in the presence of bile salts such as sodium deoxycholate
What are the uses of Bile Solubility Test?
It is most typically used to distinguish Streptococcus pneumoniae from other alpha-hemolytic streptococci, notably those of the Viridans group.
Streptococcus pneumoniae is bile soluble, which means that it lyses in the presence of bile salts, whereas Viridans streptococci are bile insoluble.
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