The genus Streptococcus represents a heterogeneous group of Gram-positive bacteria.
Many streptococci exist as part of the normal microbial flora of both animals and humans.
Under certain favorable conditions, some species can cause diseases, ranging from sub-acute infections to acute or even chronic diseases/infections.
Streptococci are spherical in shape and grow in chains due to incomplete separation following cell division.
Historically, hemolysis—defined as the lysis of erythrocytes on blood agar medium caused by enzymes (toxins) released by the organism—was considered a valuable identifying characteristic for streptococci.
Based on hemolytic characteristics, streptococci were initially classified into β-hemolytic and non-β-hemolytic groups.
In 1934, the classification was expanded through Lancefield grouping, which is based on the presence of group-specific wall polysaccharides located on the bacterial surface.
Thirteen distinct serological groups have been identified to date.
Among these, groups A, B, C, and G, along with Streptococcus pneumoniae and the viridans group streptococci, are considered most significant for human health.
Hemolysis refers to the breakdown of red blood cells (RBCs).
Organisms capable of producing a toxin known as hemolysin contribute to the hemolysis process.
In 1919, Brown introduced the terms alpha, beta, and gamma to describe three types of streptococci, distinguished by their hemolytic reactions on blood agar plates.
Beta-hemolytic streptococci exhibit complete lysis of hemoglobin in red blood cells surrounding the colony on blood agar.
This reaction produces a clear zone around the colony with transparency of the underlying medium when viewed under light.
Beta hemolysis is caused by two hemolysins: hemolysin O and hemolysin S.
Hemolysin O is oxygen-labile, meaning it becomes inactive in the presence of oxygen.
Hemolysin S is an oxygen-stable cytotoxin, remaining active even in oxygen-rich conditions.
Beta hemolysis is more pronounced when blood agar plates are incubated anaerobically.
The organism also produces various toxic by-products that contribute to the destruction of red blood cells or the lysis of hemoglobin.
Examples: Streptococcus pyogenes, or group A streptococci (GAS). Weakly beta-hemolytic species: Streptococcus agalactiae or group B Streptococcus (GBS).
Examples of beta-hemolytic streptococci include Streptococcus pyogenes (Group A Streptococci, GAS).
A weakly beta-hemolytic species is Streptococcus agalactiae (Group B Streptococcus, GBS).
In certain cases, hemolytic reactions display synergy, where the combination of two reactions results in a stronger effect than either reaction alone.
Streptococcus agalactiae typically shows weak beta-hemolysis, but when located near a strain of Staphylococcus species, the beta-hemolysins from both organisms combine to produce an intense beta-hemolytic reaction with increased visibility and clarity.
2. Alpha-hemolytic Streptococci (incomplete hemolysis or partial hemolysis)
Alpha-hemolytic streptococci are Gram-positive, spherical, and non-motile organisms, with cells arranged in pairs or short chains.
They are facultative anaerobes and produce a characteristic α-hemolytic zone on blood agar plates.
Alpha hemolysis is a partial or smaller zone of hemolysis, presenting as a narrow grass-green zone around colonies on blood agar.
This greenish coloration is due to the oxidation of iron in hemoglobin within red blood cells, converting it into methemoglobin.
The organism causes partial hemolysis of hemoglobin but leaves the red blood cell membrane intact.
Microscopic observation of alpha-hemolyzed red blood cells confirms that the cell membranes remain unbroken, meaning it is not true lysis.
Examples: Streptococcus pneumoniae, viridans streptococci (Streptococcus viridans, S. mutans, S. sanguinis)
Examples of alpha-hemolytic streptococci include Streptococcus pneumoniae and viridans streptococci such as Streptococcus viridans, S. mutans, and S. sanguinis.
It is important not to confuse “partial” or “incomplete” hemolysis (alpha hemolysis) with the “weak” or “subtle” lysis seen in Group B streptococci (Streptococcus agalactiae).
In beta hemolysis, there is no brown or green discoloration of the cells in the surrounding medium.
With prolonged incubation, many alpha-hemolytic reactions may become more vivid and distinct.
If the surrounding medium shows any shades of brown or green, the hemolysis is still classified as alpha, regardless of intensity.
3. Gamma-hemolytic (Non-haemolytic) Streptococci
Gamma hemolysis indicates no hemolysis or absence of hemolytic activity around the bacterial colony.
Colonies grown on blood agar do not exhibit the typical characteristics of either alpha or beta hemolysis.
In some cases, there may be slight discoloration of the medium.
Blood agar plates showing gamma hemolysis often appear brownish, which results from the normal reaction of the blood under the growth conditions used (37 °C in the presence of CO₂).
Examples: Formerly Group D Streptococci, re-classified as Enterococcus faecalis and Enterococcus faecium.
References
Tille, P. M. (2014). Bailey & Scott’s Diagnostic Microbiology (13th ed.). Mosby, Inc., an affiliate of Elsevier Inc., 3251 Riverport Lane, St. Louis, Missouri 63043.
Buxton, R. (2005). Blood Agar Plates and Hemolysis Protocols. American Society for Microbiology.
Patterson, M. J. (1996). Streptococcus. In S. Baron (Ed.), Medical Microbiology (4th ed.). Galveston, TX: University of Texas Medical Branch at Galveston. Chapter 13. Retrieved from https://www.ncbi.nlm.nih.gov/books/NBK7611/
%20Test%20Principle,%20Procedure,%20Uses,%20Interpretation%20&%20Limitations.webp)
