Streptococcus.pdf

GRAM POSITIVE COCCI STREPTOCOCCI  STREPTOCOCCI Gram (+) spherical cells, arranged in chains or pairs Grayish pinpoint translucent to slightly opaque colonies Catalase (-), oxidase (-) Growth is enhanced by blood, serum or glucose incorporated in agar plate ...

GRAM POSITIVE COCCI STREPTOCOCCI  STREPTOCOCCI Gram (+) spherical cells, arranged in chains or pairs Grayish pinpoint translucent to slightly opaque colonies Catalase (-), oxidase (-) Growth is enhanced by blood, serum or glucose incorporated in agar plate  STREPTOCOCCI Behave-like facultative anaerobe, some species require increased CO2 for growth Aerotolerant anaerobe (peptostreptococci – obligate anaerobe  STREPTOCOCCI ALL streptococci EXCEPT the viridans group have a layer of C carbohydrate – serological classification Young cultures are characterized by the presence of capsule Are commonly found as part of normal human flora.  STREPTOCOCCI However, when these organism gain access to normally sterile sites they can cause life threatening infection. S. pyogenes and S. pneumoniae - notorious pathogens Aerococcus, Lactobacillus, Leuconostoc and Pediococcus – resemble streptococci  STREPTOCOCCI Colony variation a. Mucoid b. Matte or rough – large amount of CHON c. Smooth/glossy – little amount of CHON A. ACADEMIC/BERGEY’S CLASSIFICATION BASED ON TEMPERATURE CLASSIFICATION OF STREPTOCOCCI  1. PYOGENIC GROUP Will not grow neither on 10°C or 45°C Produce pus; mostly β-hemolytic Species: Streptococcus pyogenes  2. VIRIDANS GROUP Will grow at 45°C NOT part of the lancefield group Alpha hemolytic OR Non hemolytic Species: S. salivarius, S. mutans, S. mitis, S. sanguis  3. ENTEROCOCCUS Will grow both at 10°C and 45°C Normal flora of human intestine Species: E. faecalis  4. LACTIC GROUP Will grow on 10°C Non hemolytic organism Often found in dairy products Species: S. lactis B. SMITH AND BROWN’S CLASSIFICATION BASED ON HEMOLYTIC PATTERN CLASSIFICATION OF STREPTOCOCCI 1. ALPHA-HEMOLYTIC STREPTOCOCCI Partial/incomplete hemolysis of RBC around colony Green discoloration around colony Species: S. pneumoniae (green streptococci) 2. BETA-HEMOLYTIC STREPTOCOCCI Complete hemolysis of RBCs around the colony Clear area/zone around colony Yellow discoloration Species: S. pyogenes, S. agalactiae  3. GAMMA-HEMOLYTIC STREPTOCOCCI NO lysis of RBCs around colony/red cells immediately surrounding the colony are unaffected (no change) Species: E. faecalis (GROUP D)  C. LANCEFIELD BASED ON THE ANTIGENIC NATURE OF CELL WALL CARBOHYDRATES – C POLYSACCHARIDE Rebecca Lancefield – found out that C carbohydrate can be extracted from the streptococcal cell wall by placing the organism in dilute acid and heating for 10 min. CLASSIFICATION OF STREPTOCOCCI HEMOLYTIC REACTION LANCEFIELD GROUP SPECIES α none S. pneumoniae β A S. pyogenes B S. agalactiae C S. Dysagalactiae subsp. equisimilus γ D S. bovis a, β or γ D Enterococcus spp. a and γ none Viridans streptococci Small colony variants Anginosus group of A, C, F, G or ungroupable  GROUP A STREPTOCOCCI Pathogenic to man Not considered part of the normal flora Acquired thru contaminated droplets by cough or sneeze Species: S. pyogenes – “fever producing bacteria,” flesh eating bacteria (involves deeper tissues and organs)  GROUP A STREPTOCOCCI Principal virulence factor: M-protein (type specific; anti-phagocytic) Other virulence factor: a. Protein F – mediates epithelial cell attachment b. Lipoteichoic acid – bacterial adherence to the respiratory epithelium ENZYMES AND TOXINS PRODUCED BY S. pyogenes 1. HEMOLYSINS A. STREPTOLYSIN O - “Oxygen labile”; highly antigenic - responsible for subsurface hemolysis on BAP - cause lysis of WBC, platelets, tissue cells - ASO test – detects recent infection w/ S. pyogenes ENZYMES AND TOXINS PRODUCED BY S. pyogenes B. STREPTOLYSIN S - “Oxygen stable”; non antigenic - responsible for surface hemolysis on BAP - it causes lysis of WBCs 2. DEOXYRIBONUCLEASES - degrading DNA via hydrolysis of its phosphodiester backbone ENZYMES AND TOXINS PRODUCED BY S. pyogenes 3. STREPTOKINASE - a protein that binds to plasminogen and activates the production of plasmin - it allows the bacteria to move from clotted area - it activates a host blood-factor that dissolves fibrin clots  ENZYMES AND TOXINS PRODUCED BY S. pyogenes 4. HYALURONIDASE - to separate the tissue and spread the organism 5. PYROGENIC (ERYTHROGENIC) TOXINS - serotypes A, B, C - exotoxin B (cysteine protease) – degrades proteins INFECTIONS AND DISEASES 1. PHARYNGITIS OR TONSILITTIS – “STREP THROAT” 2. SCARLETT FEVER (SCARLATINA) Results from a throat infection with a strain of S. pyogenes that carries a lysogenic bacteriophage. Acquired through respiratory droplet Cardinal signs: diffused red rash on the upper chest and spreads to the trunk and extremities and strawberry colored-tongue Caused by the release of erythrogenic toxin/strep pyrogenic exotoxin  2. SCARLETT FEVER (SCARLATINA) Red cheeks and rash Strawberry tongue 2. SCARLETT FEVER (SCARLATINA) DICK’S TEST (ERYTHROGENIC TOXIN) - susceptibility test for scarlet fever - (+) erythema – redness SCHULTZ CHARLTON REACTION - diagnostic test for scarlet fever - used to diagnostic present scarlet fever - (+) “blanching phenomenon” – rash fade  3. SKIN INFECTIONS CELLULITIS - is a diffuse, spreading infection of subcutaneous skin tissue characterized by a defined area of redness (erythema) and accumulation of fluid (edema)  3. SKIN INFECTIONS ERYSIPELAS - is an acute infection and inflammation of the dermal layer of the skin characterized by painful reddish patches that enlarge and thicken with sharply defined edges - may also lead to necrotizing fasciitis and myositis – “galloping gangrene”  3. SKIN INFECTIONS RHEUMATIC FEVER ACUTE GLOMERULONEPHRITIS OR BRIGHT’S DISEASE - inflammatory disease of the renal glomeruli; results from the deposition of antigen-antibody complexes. DIAGNOSTIC TESTS  BACITRACIN DISK TEST Differentiates S. pyogenes from other β- hemolytic group Result: (+) any zone of inhibition Group C and G are also susceptible  SULFAMETHOXAZOLE- TRIMETHOPRIM TEST Positive: Any zone of inhibition SXT Negative: No Zone of inhibition (Group A and Group B) SXT  L-pyrrolidonyl-β-napththylamide (PYR) TEST Specific than Bacitracin test (+) bright/cherry red color (β-napththylamide) upon adding 0.01% cinnamaldehyde reagent (-) no color change Other PYR test positive organisms: Enterococcus, Aerococcus, Gemella  GROUP B STREPTOCOCCI Part of the normal flora of female genital tract and lower GIT Nosocomially transmitted by unwashed hands of mother or healthcare personnel. Infection of fetuses and infants – infection during passage through the colonized birth canal, and premature rupture of mother’s membranes  GROUP B STREPTOCOCCI On culture, it is grayish white mucoid w/ small zone of beta hemolysis Species: S. agalactiae Virulence factor: capsule (sialic acid – signifiant component of the capsule) Enzymes: hemolysin, CAMP factor, neuraminidase, deoxyribonuclease, hyaluronidase and protease  DISEASES AND INFECTIONS Pneumonia Neonatal Meningitis Neonatal sepsis Postpartum infection Osteomyelitis UTI endocarditis DIAGNOSTIC TESTS  CAMP TEST To differentiate S. agalactiae from other β- hemolytic streptococci It uses β-lysin – producing strain of S. aureus or disk impregnated w/ β lysin Result (+) arrow-head β-hemolysis near S. aureus growth / (+) bowtie appearance  HIPPURATE HYDROLYSIS TEST S. agalactiae possesses the enzyme hippuricase Reagent: sodium + ninhydrin Results (+) Purple color after adding Ninhydrin rgt (indicates hippurate hydrolysis) (-) no color change  GROUP C STREPTOCOCCI Main source of streptokinase; animal pathogen Species: S. equi, S. equisimilis, S. dysagalactiae, S. zoopedimicus  GROUP D STREPTOCOCCI Enterococci: E. faecalis, E. faecium, E. avium, E. gallinarium Non enterococci: S. bovis, S. equinus Enterococci are not highly pathogenic; cocci in pairs and long chains DIAGNOSTIC TEST GROWTH IN BILE ESCULIN MEDIUM Reagent: Esculin + 1-4% bile salt Result: (+) Black color (BOTH enterococci and non enterococci)  6.5% NaCl (nutrient broth base) test Result: (+) turbididty – enterococci (also PYR +) (-) non enterococci  LAP Test Detects for the ability of the bacteria to hydrolyze Leucine-β-naphthylamide by enzyme leucine aminopeptidase to β-naphthylamide. Positive: Development of a red color within 1 minute after adding cinnamaldehyde reagent (S. Pneumoniae and S. pyogenes, Enterococcus, Pediococcus ) Negative: No color change or development of slight yellow color (Aerococcus and Leuconostoc)  NOTES Enterococci – (+) bile esculin, growth in 6.5% NaCl. PYR test and LAP test; vancomycin resistant Leuconostoc, Pediococcus, Globicatella, S. urinalis, Lactococcus – (+) growth in bile esculin and 6.5% NaCl All enterococci isolated from human hydrolyze PYR  NOTES Isolation of S. bovis from a blood culture may be the first indication that the patient has an occult tumor in the GI tract (GI cancer) 85% of Group B streptococci is (+) w/ growth in 6.5% NaCl but (-) in bile esculin test E. faecalis requires cyanocobalamin (B12) as growth factor.  GROUP F and G Minute streptococci w/ wide zone of beta hemolysis Species: S. anginosus  VIRIDANS STREPTOCOCCI Alpha hemolytic streptococci that lack the lancefield group antigens and do not fall on the criteria for S. pneumoniae, can be non- hemolytic. Oropharyngeal commensals Opporunistic pathogens of low virulence Glucans and dextrans – enhance attachment to host cell surfaces  VIRIDANS STREPTOCOCCI Laboratory tests: bile insoluble, optochin resistant, no growth in 6.5% NaCl, (-) inulin fermentation and PYR (-) Species: S. mutans, S. salivarius, S. mitis Infections: subacute endocarditis, dental carries (S. mutans)  ABIOTROPHIA Pyridoxal-dependent or Vit B6 dependent and symbiotic streptococci Formerly known as nutritionally variant streptococci (NVS) Thiol compounds – cystein, Vit B6 and pyridoxal This group of organism were found as satellites around an organism that produces pyridoxal – E. coli, Klebsiella spp.,Enterobacter spp., yeasts  ABIOTROPHIA They produce satellatism w/ Staphylococcus Opportunistic pathogens of low virulence Variable morphology – gram (+) to gram (-) to gram variable; pleomorphic forms It causes 5-6% endocarditis STREPTOCOCCUS SPP. LABORATORY DIAGNOSIS 1. Gram Stain 2. Culture – BAP, PEA, CNA, CAP For isolation of Group A streptococci from throat swab, BAP w/ SXT is used to suppress the growth of normal flora. To detect genital carriage of Group B streptococci during pregnancy, Todd Hewitt broth w/ antibiotics (gentamicin, nalidixic acid or colistin and nalidixic acis) is used to suppress vaginal flora.  Visualization of beta hemolysis is enhanced by anaerobic condition (BAP should be inoculated by stabbing the inoculating loop into the agar several times) CAP is incubated w/ 5-10% CO2 Abiotrophia will not grow on BAP or CAP unless pyridoxal (B6) is supplied. Streptococci has tendency to produce increase amount of lactic acid and other organic acid from fermentation of sugar resulting to acidic medium and eventually their growth is inhibited. 3. Susceptibility Test – bacitracin disk test SXT test 4. CAMP and Hippurate test 5. Bile Esculin and Salt Tolerance Test 6. Serological Test a. Latex agglutination b. ASO titer test c. ELISA d. Co-agglutination  Streptococcus pneumoniae Diplococcus/pneumococcus Gram (+) cocci in pairs, oval or lancet shape; facultatively anaerobe Causative agent of lobar pneumonia Considered part of the normal flora (25-50%) of the URT of preschool children Most common bacterial pneumonia in elderly as well as in patients w/ underlying disease.  Streptococcus pneumoniae Most common cause of bacterial meningitis in adults Common isolate both as a pathogen and as a member of the normal respiratory tract. Principal virulence factor: antiphagocytic capsular polysaccharide – type 3 strains Opsonization of the capsule renders the organism non virulent  Streptococcus pneumoniae The capsule is antigenic and can be identified w/ appropriate antiserum; it is composed of hyaluronic acid. Alpha hemolytic colonies, mucoid, flattened with depressed center. “dome-shaped,” glistening, wet, mucoid – young colonies “coin w/ a raise rim” – old colonies (due to autolytic changes)  Streptococcus pneumoniae Dimple-shaped/donut-shaped colonies Enzymes: hemolysin, immunoglobulin A protease, neuraminidase and hyaluronidase INFECTIONS/DISEASES PRODUCED 1. Pneumonia - pneumonia due to S. pneumoniae is not usually a primary infection but rather a result of disturbance of the normal defense barriers. 2. Meningitis 3. Otitis media 4. Bacteremia 5. Endocarditis 6. Peritonitis  Differential Test Result 1. Optochin Susceptibility/P disk Test Presumptive test > Or = 14mm Ethylhydroxycuprein hydrochloride BAP (sensitivity media) 2. Bile Solubility Confirmatory test Differentiates pneumococcus from viridans streptococci Sodium desoxycholate When a heavy suspension of (+) soluble pneumococcus is added to bile salt, the cloudiness of the broth clears after 3 hrs incubation.  Differential Test Result 3. Neufeld Quellung Reaction The antipnemococcal serum is mixed w/ sputum, CSF and SWELLING CAPSULE other sources along w/ methylene blue Examined under OIO 4. Mouse Virulence Death of the animal 5. Inulin Fermentation Test (+) Fermentation