Streptococci- Overview of Detection, Identification, Differentiation and Cultivation Techniques

By: By Jvo Siegrist, Product Manager Microbiology,, AnalytiX Volume 7 Article 3

Streptococci are non-motile, microaerophilic, Grampositive spherical bacteria (cocci). They often occur as chains or pairs and are facultative or strict anaerobes. Streptococci give a negative catalase test, while staphylococci are catalase-positive. The cell division of Streptococci species involves two separate biosynthetic events: peripheral cell-wall elongation and septal-wall synthesis.1 Because they are unable to synthesize cytochromes, Streptococci cannot carry out oxidative phosphorylation. They are able to ferment sugars, but the end product is always lactic acid. Therefore, Streptococci are very acid tolerant and count among the lactic acid bacteria order.

There are many natural sources of Streptococci, including humans and diverse animals where they often colonize the mucosal surfaces of mouth, intestinal tract, nasal passages and pharynx. The presence of Streptococci  in drinking water indicates fecal contamination. Food sources with high risk of Streptococcal contamination include milk and dairy products, eggs, steamed lobster, ground ham, potato salad, custard, rice pudding and shrimp salad. In most Streptococcal food poisoning cases, the food was allowed to stand at room temperature for several hours between preparation and consumption. The contamination of the food is most often the result of poor hygiene, handling of the food by infected people, or the use of raw (unpasteurized) milk. Although they can be potent pathogens, some Streptococci are commercially important for the production of cheese and yogurt. These include S. lactis, S. cremoris, S. diacelillactis and S. thermophilus, the latter being the most well-known.

For detection, identification, differentiation, enumeration and cultivation of Streptococci, Sigma-Aldrich provides a broad range of specific agars and broths (Table 1), Streptococci Diagnostic Tests (Table 2) and a Gram staining kit and component solutions (Table 3).


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Scientific classification of Streptococcus

Kingdom: Eubacteria
Phylum: Firmicutes
Class: Bacilli
Order: Lactobacillales
Family: Streptococcaceae
Genus: Streptococcus

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Grouping of Streptococci based on hemolytic activity

Streptococci are divided into three groups based on their hemolytic (red blood cell lysing) activity. The hemolytic reaction can be visualized on blood agar plates, such as the non-selective Agars for Differentiation that are listed in Table 1.

Table 1. Media for Streptococci

Non-selective Enrichment Broths
Cat. No. Brand Description
53286 Sigma-Aldrich Brain Heart Broth
B2551 Sigma-Aldrich Brewer thioglycollate medium
22089 Sigma-Aldrich Casein peptone Lecithin Polysorbate Broth
22098 Sigma-Aldrich CASO Broth
60865 Sigma-Aldrich Cooked Meat Broth
17123 Sigma-Aldrich Elliker Broth
05121 Sigma-Aldrich Heart Infusion Broth
70122 Sigma-Aldrich Nutrient Broth No. 1
70149 Sigma-Aldrich Nutrient Broth No. 3
40893 Sigma-Aldrich Peptone Water, phosphate-buffered, Vegitone
70179 Sigma-Aldrich Peptone Water
77187 Sigma-Aldrich Peptone Water, phosphate-buffered
S4681 Sigma-Aldrich Standard Nutrient Broth No. 1
85905 Sigma-Aldrich Stuart Ringertz Medium
90404 Sigma-Aldrich Thioglycolate Broth with Resazurine
T1438 Sigma-Aldrich Todd Hewitt Broth
22092 Sigma-Aldrich Tryptic Soy Broth
51228 Sigma-Aldrich Tryptic Soy Broth No. 2
41298 Sigma-Aldrich Tryptic Soy Broth, Vegitone
T3938 Sigma-Aldrich Tryptone Soya Broth without Dextrose
T4407 Sigma-Aldrich Tryptose Broth
V5262 Sigma-Aldrich Veal Infusion Broth


Selective Enrichment Broths
Cat. No. Brand Description
17157 Sigma-Aldrich Glucose Azide Broth


Non-selective Agars for Cultivation, Enumeration and Isolation
Cat. No. Brand Description
A3340 Sigma-Aldrich AC Agar
70138 Sigma-Aldrich Brain Heart Infusion Agar
22095 Sigma-Aldrich CASO Agar
27688 Sigma-Aldrich Columbia Agar
D3060 Sigma-Aldrich Dextrose starch agar
D8184 Sigma-Aldrich Diagnostic sensitivity test agar
70147 Sigma-Aldrich Milk Agar
70148 Sigma-Aldrich Nutrient Agar
17197 Sigma-Aldrich Peptonized Milk Agar
70152 Sigma-Aldrich Plate Count Agar
88588 Sigma-Aldrich Plate Count Agar according to Buchbinder et al.
19718 Sigma-Aldrich Plate Count Agar, Vegitone
17257 Sigma-Aldrich Streptococcus thermophilus Isolation Agar
22091 Sigma-Aldrich Tryptic Soy Agar
14432 Sigma-Aldrich Tryptic Soy Agar, Vegitone
51414 Sigma-Aldrich Tryptic Soya Agar with Polysorbate 80 and Lecithin
70159 Sigma-Aldrich Tryptone Glucose Extract Agar
T2313 Sigma-Aldrich Tryptose Agar


Non-selective Agars for Differentiation
Cat. No. Brand Description
70133 Sigma-Aldrich Blood Agar (Base)
B1676 Sigma-Aldrich Blood Agar Base No. 2
55420 Sigma-Aldrich CLED Agar
70136 Sigma-Aldrich Deoxyribonuclease Test Agar
D2560 Sigma-Aldrich DNase Test Agar with Toluidine Blue
17153 Sigma-Aldrich LS Differential Agar


Selective Agars with Differential System for Differentiation, Detection and Isolation
Cat. No. Brand Description
06105 Sigma-Aldrich Bile Esculin Azide Agar
17151 Sigma-Aldrich Kanamycin Esculin Azide Agar
01337 Sigma-Aldrich Mitis Salivarius Agar


Media for Sensitivity Testing
Cat. No. Brand Description
97580 Sigma-Aldrich Mueller-Hinton Agar 2

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Alpha-Hemolytic Streptococci

In alpha hemolysis, the red blood cells remain intact, but the hemoglobin is converted to biliverdin. This causes dark greening of the blood agar plate around the colonies.

  • Pneumococci
    S. pneumoniae: Causes bacterial pneumonia, otitis media and meningitis. S. pneumoniae sensitivity to optochin (ethyl hydrocupreine hydrochloride) is the basis of a diagnostic test (Figure 1, Table 2).

    Figure 1:  Differentiation and identification flow chart of Gram-positive cocci

    Table 2.  Diagnostic tests for identification and differentiation of streptococci
    Cat. No. Brand Description
    88597 Sigma-Aldrich Catalase Test
    08382 Sigma-Aldrich Bacitracin Disks
    74042 Sigma-Aldrich Optochin Disks
    67886 Sigma-Aldrich PYRase Strips

  • Viridans and Others
    S. mutans: Associated with dental caries.
    S. viridans: Causative agent for endocarditis and dental abscesses.
    S. salivarius: Considered to be an opportunistic pathogen.
    S. salivarius subsp. thermophilus: Used in the production of cheese and yogurt.

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Beta-Hemolytic Streptococci

Beta hemolysis is a complete hemolysis of erythrocytes by the enzyme hemolysin. Clear zones will appear around the colonies on the blood agar plate. Beta-hemolytic Streptococci are further divided into serological groups using specific antibodies that recognize surface carbohydrate antigens (e.g. A, B, etc.).

  • Group A
    S. pyogenes: Causes infections like strep throat, acute rheumatic fever, scarlet fever, acute glomerulonephritis and necrotizing fasciitis. It is sensitive to bacitracin (Fig. 1), a fact that is employed in a Streptococcus diagnostic test (Table 2). Other Streptococci species may also possess the Group A antigen but are not human pathogens. These include S. dysgalactiae subsp. equisimilis and S. anginosus strains.
  • Group B
    S. agalactiae: Causes meningitis in neonates and the elderly. Occasionally colonizing the female reproductive tract, they increase the risk for premature rupture of membranes and transmission of the infection to the child.
  • Group C
    S. equi: Causes strangles infection in horses.
    S. zooepidemicus: Causes infections in cattle, horses and other mammals.
  • Group D (Enterococci)
    Many former Group D Streptococci have been reclassified to the genus Enterococcus (e.g. S. faecalis, S. faciem, S. durans, S avium).
    S. bovis and S. suis: Still in the Group D streptococci.

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Non-Hemolytic or Gamma-Hemolytic Streptococci

Gamma hemolysis is a misnomer as there is actually no hemolysis. Non-hemolytic or gamma-hemolytic Streptococci rarely cause disease.


Table 3.  Gram staining kit and component solutions
Cat. No. Brand Description
77730 Sigma-Aldrich Gram Staining Kit
94448 Sigma-Aldrich Gram‘s Crystal violet Solution
75482 Sigma-Aldrich Gram‘s Decolorizer Solution
87794 Sigma-Aldrich Gram’s Fuchsin Solution
90107 Sigma-Aldrich Gram‘s Iodine Solution
94635 Sigma-Aldrich Gram‘s Safranin Solution

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  1. Beilharz K, et al. (2012). Control of cell division in Streptococcus pneumonia by the conserved Ser/Thr protein kinase StkP.  Proceedings of the National Academy of Sciences, 109(15), E905-E913.


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