L2 Streptococci Exam Notes 2025 PDF

460 MBIO Medical Bacteriology Lecture 2 Micrococcus (Streptococcus and Enterococcus ) 0 Learning Outcomes (LLO): By the end of this lecture, you should be able to: 1. Identify the general characteristics and morphology of Micrococcu...

460 MBIO Medical Bacteriology Lecture 2 Micrococcus (Streptococcus and Enterococcus ) 0 Learning Outcomes (LLO): By the end of this lecture, you should be able to: 1. Identify the general characteristics and morphology of Micrococcus and Streptococcus species. (CLO 1.1 and CLO 1.2) 2. Explain the classification of Streptococcus using Brown's, Lancefield's, and Sherman’s classification systems. (CLO 1.1 and 2.1) 3. Describe the diagnostic methods used to identify Streptococcus species, including biochemical tests like the PYR test, and serological methods like the Lancefield grouping.(CLO2.1) 4. Discuss the pathogenicity and virulence factors of Streptococcus pyogenes and Streptococcus agalactiae, relating them to the diseases they cause.(CLO 1.2 and 1.3) 5. Interpret the clinical relevance of Streptococcus infections, especially in immune-compromised individuals, neonates, and pregnant women.(CLO 1.2 ) 1  Micrococcus General characteristics Morphology: Gram +ve cocci Arrangement : Tetrads (Fours) Non motile, non capsulated, non sporulated. Habitat: Domain Bacteria A normal presence in the upper respiratory tract and skin Phylum Actinomycetota (Commensal/sometimes opportunistic in Class Actinomycetia immunocompromised) Order Micrococcales Family Micrococcaceae Notable Species : Genus Micrococcus 1- M. varians** 2- M. luteus (Milk Micrococci) Scientific Classification of Micrococcus sp 3- M. roseus** (Dust of the air) Culture: Strictly aerobic at 37°C incubation (24 hr) Grow on ordinary media (Nutrient agar) Grow on Blood agar (Gamma) Grow on mannitol salt agar given rise to rose or pink colonies except M. luteus.(yellow) ** They were classified into the genus Micrococcus but have now been removed from Micrococcus based on phylogenetic and chemotaxonomic analysis into 2 Kocuria spp.  On Blood Agar K.varians (Yellow-ish) M.luteus (Yellow) K.roseus (Pink) Previously (M.varians ) Previosly (M.roseus ) No hemolysis (Gamma) 3 Diseases: (In Immune compromised) Acute endocarditis Urinary tract infection Mastitis Chest infection Brain abscesses Diagnostic Tests Biochemical tests: Catalase (+ve) Coagulase (-ve) Oxidative-fermentation of glucose : (oxidative positive) Antibiotic testing: To differential between Micrococcus and Staphylococcus by Furazolidone and Bacitracin tests. Furazolidone test result:  Micrococcus → resistance.  Staphylococcus → sensitive Bacitracin test result:  Micrococcus → sensitive  Staphylococcus → resistance Resistance: Most Micrococcus isolates are susceptible to a wide range of antibiotics (Penicillin, methicillin, gentamicin, and erythromycin); however, resistance to a variety of antibiotic classes has been reported, and vancomycin is the preferred agent for empiric therapy. 4  Streptococcus spp General characteristics Morphology: Gram (+ve) cocci Arrangement : chain Non motile Non sporulated Capsulated or not. Domain Bacteria Notable Species Phylum Bacillota S.pyogenes, S. agalactiae, S pneumoniae, Class Bacilli and E. faecalis Order Lactobacillales Family Streptococcaceae Culture Genus Streptococcus Facultative anaerobic Scientific Classification of Streptococcus sp On broth media: Sediment Grow very poorly on ordinary media (dew drops like colonies), but growth is enhanced with blood or serum and glucose. Biochemical Catalase (-ve) 5  Classification of Streptococci and Enterococci based on Due to the heterogeneity of the Streptococcus group, multiple classification systems have been introduced: 1. Brown`s classification is based on the degree of blood haemolysis. (Alpha, Beta, Gamma). 2. Lancefield classification relies on the presence of carbohydrate antigens in the cell wall. 3. Sherman`s classification relies on haemolytic reaction, Group carbohydrate antigen, and specific physiological characteristics, including fermentation and tolerance tests. (such Temp (10°C and 45°C), pH, and salt) 6 1. Brown`s classification: Depend on degree of haemolysis reactions on blood agar  Beta-hemolytic ( β) Complete lysis of red cells surrounding the colony, it appears as a clear zone. most pathogenic group , e.g., Strep. Pyogenes  Alpha-hemolytic (α) Partial hemolysis (greenish discoloration) associated with reduction of red cell hemoglobin. Mild pathogenic group , e.g., Strep. pneumoniae and Strep. viridans  Gamma-hemolysis (γ ) Non hemolytic group (Mainly saprophytic) e.g. , S. lactis (now is called Lactococcus lactis) and Enterococcus faecalis 7 2. Lancefield classification Streptococci are identified based on the serologic reactivity of polysaccharide antigens present in their cell walls, as outlined by Rebecca Lancefield. The serological detection of these cell wall antigens allows for alphabetical classification of streptococci, with Lancefield groups A–H, K–V, and an untippable group defined by the possession of specific cell wall antigens. For example : Group A : S. pyogens - pharyngitis, skin infections. Group B: S. agalactia -- neonatal infections. Group D: Enterococcus faecalis (previously identified as Streptococcus faecalis)- - nosocomial infections, endocarditis. Untippable group : S. pneumonia → pneumonia in humans.. 8  Lancefield classification (Latex Agglutination test ) The presence of agglutination (as The image shows the latex seen B)on the left image indicates agglutination test principle. Latex a positive reaction, identifying the beads coated with specific specific Lancefield group of the antibodies clump together when tested Streptococcus species. In mixed with a bacterial sample this case, image B shows a positive containing the corresponding agglutination reaction, suggesting antigen, indicating a positive the bacterial strain belongs to the result. Lancefield Group B. 9 3. Sherman`s classification (Lancefield’s/hemolysis/Biochemical /Physiological properties) Sherman’s Groups’ Classification Pyogenic Viridians Lactic Enterococcus Representative spp S. pyogens S. viridans S. lactis E. faecalis S.pneumoniae Lancefield's Groups’ Classification A,B,C,E,F, and G - N D Hemolysis Beta Alpha “S.pneumoniae”/No heam No heam No heam/sometime Alpha (Brown’s classification) Growth at 10 C - - + + Growth at 45 C + + - + Growth on pH 9.6 - - - + Growth on high salt (6.5% NACL) - - - +  (Group A streptococcus) Streptococcus pyogenes General characteristics Morphology: As general characters, (have a capsule composed of hyaluronic acid Culture As general character + β haemolysis Biochemical As a general character Disease Strep. pyogenes typically act as an exogenous secondary invader, often occurring after viral diseases or disruptions in the normal flora. S. pyogenes cause a variety of diseases, such as: a) Pharyngitis (Strep Throat), Scarlet fever (Complication of strep throat), skin infections (impetigo, necrotizing fasciitis, cellulitis) b) Post-streptococcal diseases, such as Rheumatic fever, are autoimmune conditions triggered by similarities between heart muscle antigens and Streptococcus antigens. Antibodies formed against Streptococcus may attack the heart, leading to carditis. Other post-streptococcal conditions include Rheumatic kidney. 11 Pathogenicity and Toxins S. pyogenes possesses a variety of virulence factors that contribute to its pathogenicity. a) Surface protein: M Protein, a major virulence factor that helps the bacteria evade phagocytosis by the host's immune cells. It also plays a role in adherence to host tissues. Capsules: The bacterium has a hyaluronic acid capsule that helps it resist phagocytosis by immune cells. Other: Protein F (Adherence on host cells), and lipoteichoic acid (Gram - positive cell wall) b) Enzymes: Hyaluronidase: An enzyme that breaks down hyaluronic acid in connective tissue, facilitating the spread of the bacteria through tissues. Streptokinase: An enzyme that dissolves blood clots, allowing the bacteria to spread more easily through the bloodstream and tissues. Other: Dnase, C5a peptidase (Inactivates the C5a component of the complement system for the purpose of immune evasion). c) Toxins: H emolysins (Streptolysin O and S): These toxins cause lysis of red and white blood cells, contributing to tissue damage and the spread of infection. Exotoxins (e.g., SpeA, SpeB): S. pyogenes produces streptococcal pyrogenic exotoxins (Spe), which can cause toxic shock syndrome and scarlet fever. 12 Illustration depicting the various virulence factors utilized by S. pyogenes to enhance pathogenicity (the image was taken from (Makthal, 2019)) 13  Diagnostic Tests a) Samples: From blood, throat swabs, or other infected sites. b) Initial testing: Smear (Gram stain*), Blood Culture (Beta ), Antibiotic sensitivity (Bacitracin sensitive. Used to differentiate from S. agalactiae, which is resistant.) c) Biochemical testing : PYR test positive (Distinguish from other beta hemolysis species). d) Rapid antigen detection: A rapid latex agglutination tests from throat swabs. e) Serological tests: These are used to detect antibodies against the bacteria, such as Lancefield Grouping. Others usually used, for the identification of post-streptococcal diseases, such tests include: Anti-streptolysin O titer (ASO) is a blood test to measure antibodies against streptolysin O. (detected after 7-10 days of infection) Other: C-reactive protein (CRP) Anti-streptolysin O titer test result. The PYR test is a rapid method used to distinguish S. pyogenes from other similar streptococci. For instructions, visit: https://youtu.be/sCT2Qrsgqpw 14 * Cultures older than 24h old may lose their ability to retain the crystal violet-iodine complex  (Group B streptococcus) Streptococcus agalactiae General characteristics As general characters ,Beta hemolytic, and Bacitracin resistant Resides gener ally in the genital and urinary tracts of about 10-30% of the population (Verani et al., 2010). It can cause significant infections, especially in newborns, pregnant Disease women, and immunocompromised individuals. Such as : Neonatal sepsis and pneumonia, UTI in pregnant women might lead to preterm labor. Diagnostic tests a) Samples: Vaginal/rectal/wounds Swaps, urine, blood, Amniotic Fluid, or Placental Tissues b) Serological tests: Lancefield Grouping will be used to identify the Lancefield Group B antigen, confirming the presence of S. agalactiae. c) CAMP Test: S. agalactiae produces an arrowhead-shaped zone of enhanced hemolysis when grown near Staphylococcus aureus on blood agar. 15 CAMP test: The group B streptococci are streaked perpendicular to a streak of S. aureus on blood agar. Results: A positive reaction appears as an arrowhead zone of hemolysis adjacent to the place where the two streak lines come Lancefield Grouping (agglutination into proximity. test): A sample is mixed with latex beads coated with antibodies. Corresponding to S. agalactiae specific carbohydrate antigen. Results: If the antigen is present, it will react with the antibodies , causing the latex beads to clump (1). 16  (Group D streptococcus) Enterococcus faecalis General characteristics Arranged typically appears in pairs or short chains. Commensal bacterium(Normal colonists of human large intestine) high salt concentrations (6.5% NaCl) and bile presence (See Strepto Classification). Usually, non-hemolytic or α hemolytic Naturally high levels of antibiotic resistance Diseases Is a major cause of nosocomial (hospital-acquired) infections, including urinary tract infections (UTIs). It is particularly problematic in patients with indwelling medical devices, such as catheters and central lines. Diagnostic tests a) Samples: Blood, Urine, CSF, wound swabs, and Catheter Tips b) Biochemical tests: PYR Test E. faecalis is PYR-positive c) Special: Growth in 6.5% NaCl, E. faecalis can grow in high salt concentrations, differentiating it from non-enterococcal streptococci. Why is it unreliable to use Lancefield Grouping for E. faecalis (Group D)as an identification method? 17  Hint For group A streptococci, this is rhamnose-N-acetylglucosamine. For group B, it is rhamnose-glucosamine polysaccharide. For group C, it is rhamnose-N-acetylgalactosamine. For group D,it is glycerol teichoic acid containing d-alanine and glucose. ”This slide is Not required in any course test." 18  Lancefield group ungroupable Viridans group streptococci (VGS) Streptococcus pneumoniae (Diplococcus pneumoniae) They possess no Lancefield antigens Streptococcus pneumoniae Vs Viridans group streptococci (VGS) 19  Viridians group streptococci (VGS) General characteristics A heterogeneous group of organisms (Human commensals or Pathogens). Alpha hemolytic or nonhemolytic. Optochin resistant. Not soluble in bile salts (biochemical tests are often used for their identification). Notable Species Streptococcus mutans (dental caries), Streptococcus salivarius Diseases Causes endocarditis and bacteremia. Diagnostic tests To differentiate between VGS and S. pneumoniae, two tests are commonly used: the Optochin Susceptibility test and the Bile solubility test 20 bile solubility test  Streptococcus pneumoniae (Diplococcus pneumoniae) General characteristics Usually, pairs of cocci (diplococci) , and lancet-shaped. Fastidious and Grows best in 5% CO2. (Fermentative aerotolerant anaerobe) Alpha hemolytic Young colonies resemble dew drops due to capsule-spontaneous autolysis of bacteria. Later, a central plateau with an elevated rim is formed. Habitat: A normal inhabitant of the human upper respiratory tract. Diseases A leading cause of bacterial pneumonia, meningitis, and other serious infections (such as osteomyelitis, endocarditis, and brain abscesses), particularly in young children, the elderly, and immunocompromised individuals. It also causes. Pathogenicity a) Polysaccharide Capsule: The capsule is the most important virulence factor, protecting the bacterium from phagocytosis by host immune cells. There are more than 90 different serotypes of S. pneumoniae based on the composition of the capsule. 21 Diagnostic Tests a) Samples: Sputum, Blood, CSF, Nasopharyngeal swabs b) Special: Sensitivity to optochin and bile solubility tests help confirm the identification. c) Serological tests: Quellung Reaction a serological test in which antibodies bind to the bacterial capsule, causing it to swell and become more visible under a microscope. This helps in serotype identification. 22 23