Microbio Lec 5 - Streptococcus
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Transcript of Microbio Lec 5 - Streptococcus
MICROBIOLOGY LECTURE 5 – Genus StreptococcusNotes from LectureUSTMED ’07 Sec C – AsM
Generalities
1. Gram positive cocci that grow in pairs or chains2. Widely distributed in nature3. 27 recognized species 4. Some are members of the normal human flora5. Others are associated with important human
disease attributable to infection by streptococci and to sensitization to them
6. The most important of the human pathogens:a. Streptococcus pyogenes (Group A)b. Streptococcus agalactiae (Group B)c. Enterococcus faecalis(Streptococcus faecalis)
(Group D)d. Streptococcus pneumoniaee. some of the oral streptococci
7. Elaborates a variety of extracellular substances and enzymes
8. Streptococci cause a wide variety of infections:a. Streptococcus pyogenes (Group A) streptococci-
leading bacterial cause of pharyngitis and cellulitis; inciting factor of two important immunologic diseases: acute rheumatic fever and acute glomerunephritis
b. Streptococcus agalactiae (Group B streptococci)- leading cause of neonatal sepsis and meningitis
c. Enterococcus faecalis – important cause of hospital acquired urinary tract infections and endocarditis
d. Viridans group streptococci – most common cause of endocarditis
e. Streptococcus bovis – may also cause endocarditis
9. Most species are facultative anaerobes; maybe strictly anaerobic to capnophilic
10. Have complex nutritional requirements necessitating blood or serum enriched medium for isolation
11. Carbohydrates are fermented with the production of lactic acid but not gas
12. Catalase negative13. Can be classified as to:
a. clinical presentation(pyogenic,oral,enteric)b. serological properties (Lancefield grouping A-H,
K-V) based on the antigenic composition of cell wall carbohydrates
serogroups A,B,C,D and G – most commonly found associated with human disease
c. Hemolytic patterns in blood agar plates Beta hemolysis – clear zone of
hemolysis around the colony as a result of complete lysis of the red blood cells
Alpha hemolysis – zone of partial hemolysis with a greenish discoloration of the medium
Gamma hemolysis – no color change or lysis of red blood cells ; nonhemolytic
d. Biochemical (physiological) properties Sugar fermentation reactions Tests for the presence of enzymes Tests for susceptibility or resistance to
certain chemical agentse. antigenic specificity of the capsular
polysaccharide
Beta hemolysis on 5% sheep blood agar
There is complete lysis of RBC surrounding the colony that can be seen macroscopically
There is a clear colorless zone around a streptococcal colony in
which the RBC have undergone complete destruction Alpha hemolysis on 5% sheep blood agar
an indistinct zone of partial lysis of red blood cells causing a green to greenish brown discoloration of the medium immediately surrounding the colony
Alpha hemolysis showing partially hemolyzed RBC immediately surrounding the colonoy
Gamma hemolysis on 5% sheep blood agar
Gamma denotes lack of hemolysis; the RBC surrounding the colonies are intact
Macroscopically there is no apparent hemolytic activity or discoloration produced by the colony
CLASSIFICATION OF STREPTOCOCCI
1. Group A streptococcus – Streptococcus pyogenesa. contains the group A antigenb. beta hemolyticc. main human pathogen associated with
local or systemic invasion and poststreptococcal immunologic disorders
d. PYR positive(hydrolysis of L-pyrrolidonyl-2-napthylamide)
e. usually susceptible to bacitracin
2. Group B streptococcus – Streptococcus agalactiaea. contains the group B antigenb. members of the normal flora of the female
genital tractc. an important cause of neonatal sepsis and
meningitisd. beta hemolytic e. hydrolyzes sodium hippuratef. gives a positive response to CAMP test
3. Groups C and G streptococcia. occur sometimes in the nasopharynxb. may cause sinusitis, bacteremia or
endocarditisc. beta hemolyticd. identified by reactions with specific
antisera for groups C or G
4. Group D streptococcia. enterococcal – Enterococcus
faecium Enterococcus durans
Part of the normal enteric flora Usually nonhemolytic; occasionally
alpha hemolytic PYR positive Grow in the presence of bile
Hydrolyze esculin (bile esculin positive)
Grow in 6.5% NaCl resistant to penicillin G Some strains are vancomycin
resistant
b. Non-enterococcal – Streptococcus bovis Part of the enteric flora Occasionally cause endocarditis Sometimes cause bacteremia in
patients with colon carcinoma Nonhemolytic and PYR negative Grow in the presence of bile Hydrolyze esculin (bile esculin-
positive) Do not grow in 6.5% NaCl Sensitive to penicillin
5. Streptococcus anginosus or Streptococcus milleri, Streptococcus intermedius, Streptococcus constellatus
a. part of the normal florab. may be beta, alpha or nonhemolyticc. includes:
Beta hemolytic streptococci that form minute colonies (<0.5 mm in diameter) and react with groups A,C or G antisera
All beta hemolytic group F streptococci
Voges Proskauer test positive Those that are group A are PYR
negative May be classified as Viridans
streptococci
6. Group N streptococci a. rarely found in human disease statesb. produce normal coagulation (souring )
of milk
7. Groups E,F,G,H and K-U streptococci- occur primarily in animals
8. Streptococcus pneumoniaea. alpha hemolyticb. growth inhibited by optochin
(ethylhydrocupreine hydrochloride)c. colonies are bile soluble
9. Viridans streptococci – Streptococcus mitis, Streptococcus salivarius, Streptococcus sanguis (Group H), Streptococcus mutans
a. typically alpha hemolytic ; may be nonhemolytic
b. growth not inhibited by optochinc. colonies are not soluble
bile(deoxycholate)d. most prevalent members of the normal
flora of the upper respiratory tract e. important for the healthy state of the
mucous membranesf. may reach the blood stream due to
traumag. a principal cause of endocarditis on
abnormal heart valvesh. Streptococcus mutans – synthesizes
large polysaccharides (dextrans and levans) from sucrose which may lead to dental caries
10. Nutritionally variant streptococci or pyridoxal-dependent streptococci – Streptococcus defectives, Streptococcus adjacens
a. require pyridoxal or cysteine for growth on blood agar
b. grow as satellite colonies around colonies of staphylococci and other bacteria
c. usually alpha hemolytic; may be nonhemolytic
d. part of the normal flora
e. occasionally cause bacteremia or endocarditis
f. can be found in brain abscesses and other infections
11. Peptostreptococcusa. grow only under anaerobic or
microaerophilic conditionsb. variably produce hemolysisc. part of the normal flora of the mouth,
upper respiratory tract, bowel and female genital tract
d. often participate in mixed anaerobic infections in the abdomen, pelvis, lung or brain
Antigenic Structure of Group A Streptococcus
1. Capsule - hyaluronic acid; nonimmunogenic;
antiphagocytic
2. Cell walla. group specific carbohydrates of group A – is a
dimer of N-acetylglucosamine and rhamnose
b. type specific protein antigensi. M protein
o a major antigen associated with virulent streptococci
o located at the end of the hairlike fimbriae that are anchored in the cell wall and extending through the capsule
o antiphagocytico anticomplimentary
ii. T or trypsin resistant proteino With M protein – an important
epidemiological marker of group A strains
o no relationship to virulenceiii. R protein
c. Other surface antigensi. F protein or fibronectin binding protein
o Has a receptor for fibronectino Major adhesin for bacterial attachment
to the epithelial cells of the pharynx and skin
ii. Lipoteichoic acido Lipid moiety is implicated to binding to
fibronectin.o Possibly mediates adherence to
epithelial cells
Antigenic structure of Group B Streptococcus
PATHOGENESIS
Group A streptococci (Streptococcus pyogenes) cause disease by three mechanisms:
- pyogenic inflammation – induced locally at the site of the organisms in tissue
- exotoxin production – can cause widespread systemic symptoms in areas of the body where there are no organisms
- immunologic – occurs when antibody against a component of the organism cross-reacts with normal tissue or forms immune complexes that damage normal tissue
Inflammation-related enzymes produced by Group A Streptococcus (Streptococcus pyogenes)1. hyaluronidase(spreading factor)
- degrades hyaluronic acid which is the ground substance of the connective tissue
- Facilitates spreading of the microorganisms- Antigenic – specific antibodies are found in the
serum after infection with hyaluronidase producing organisms
2. streptokinase(fibrinolysin)- Transforms plasminogen of human plasma into
plasmin, an active proteolytic enzyme that digests fibrin and other proteins
- Given intravenously for treatment of pulmonary emboli and of coronary artery and venous thromboses
3. streptodornase (streptococcal deoxyribonuclease)- Depolymerizes DNA in exudates or necrotic
tissue- With streptokinase – used in enzymatic
debridement; Helps liquefy exudates and facilitates removal of pus and necrotic tissue
- Antibody to Dnase develops after streptococcal skin infection (normal limit –100 units)
Toxins and hemolysins produced by Group A streptococci (Streptococcus pyogenes)
1. Streptococcal pyrogenic toxins
- Three antigenically distinct toxins
a. Exotoxin C– classic erythrogenic toxin causes the rash in scarlet
fever Produced only by strains
lysogenized by a bacteriophage carrying the gene for the toxin
Dick test – The injection of a skin test dose of erythrogenic toxin gives a positive result (an erythematous reaction in the skin of nonimmune persons who lack antitoxins)
Schultz Charlton reaction – antitoxin injected into the skin of a patient with scarlet fever causes localized blanching as a result of neutralization of erythrogenic toxin
Dick test – positive – erythema surrounding the injection site
Schultz Charlton reaction. Convalescent serum from a scarlet fever patient blanches the rash of a patient with scarlet fever
b. Exotoxin B a cysteine protease that
rapidly destroys tissue and is produced by strains that cause necrotizing fasciitis
c. Exotoxin A May cause streptococcal toxic
shock syndrome
2. Hemolysins
a. Streptolysin S oxygen stable,
nonimmunogenic cell bound hemolysin capable of lysing eryhtrocytes, leukocytes and platelets
stimulate release of lysosomal contents after engulfment
responsible for the hemolytic zones around streptococcal colonies growing on the surface of blood agar
not antigenic
b. Streptolysin O a protein that is hemolytically
active in the reduced state responsible for the hemolysis
seen when growth is in cuts deep into the medium in blood agar
antigenic – antibodies are formed against streptolysin O following infection with streptococci that produce streptolysin O
ASO serum titer in excess of 160-200 units – suggests:
Recent infection with streptococci
Persistently high antibody levels dueto an exaggerated immune response to an earlier exposure in a hypersensitive person
Pathogenesis of Streptococcus agalactiae (Group B streptococci)
a. is based on the ability of the organism to induce an inflammatory response
b. no cytotoxic exotoxins are producedc. role of enzymes in the pathogenesis of
infection is unknown – deoxyribonucleases, hyaluronidase, neuraminidase, proteases, hiuppurase and hemolysins
d. no evidence for any immunologically induced disease
e. has a polysaccharide capsule – antiphagocytic
f. anticapsular antibody is protective CLINICAL FINDINGS
Types of diseases produced by Streptococcus pyogenes1. Pyogenic diseases
a. pharyngitisb. cellulitis and erysipelas
c. impetigo(pyoderma)2. Toxigenic diseases
a. scarlet feverb. toxic shock syndrome
3. Immunologic diseasesa. rheumatic feverb. acute glomerunephritis
1a. Pharyngitis1. Streptococcus pyogenes is the major cause of
bacterial pharyngitis2. A disease of children 5-15 years3. Spread by person to person by respiratory
droplets
4. Characterized by sore throat, fever, malaise, headache and nausea
5. Posterior pharynx erythematous with an exudate; cervical lymphadenopathy present
6. Can result to complications (tonsillar abscesses, mastoiditis, septicemia, osteomyelitis, rheumatic fever)
1b. Cellulitis1. Cardinal features –
erythema, swelling, heat and pain
2. Erythema may be pink or red but lacks the intense, fiery red or salmon colored appearance of erysipelas.
3. Initiated by infection through a small break in the skin
4. Can invade the subcutaneous tissue and advance rapidly through lymphatics --à septicemia
1b. Erysipelas1. Characteristic appearance – bright red or
salmon red painful confluent erythema in a “butterfly” distribution involving the nasal eminence, cheeks, and nose with abrupt borders along the nasolabial folds
2. Erythema increases over a course of 3-6 days and usually resolves in 7-10 days
3. Erysipelas usually occur on the face, although any skin surface such as the leg, can be affected.
4. Note the sharp line of demarcation and bright red color, features that distinguish it from cellulitis
1c. Impetigo1. A superficial infection
that usually begins as small vesicles progressing to weeping lesions with amber crust and slightly cloudy purulent exudate.
2. Serotypes implicated – M types 2,49, 55 & 57
3. May result to nephritis as a complication
2a. Scarlet fever1. The primary site of the infection is usually the
pharynx, with the distinctive rash resulting from an erythrogenic toxin produced by the streptococcus.
2. The rash appears within 2 days after the onset of the sore throat and disappears in 6-10 days.
Left – scarlet feverRight – measles
2b. Streptococcal toxic shock syndrome (also called toxic shock like syndrome)
1. Characterized by hypotension, diffuse erythroderma, hypoalbuminemia and multiorgan failure(kidney, lungs, liver, heart)
2. serotypes implicated – M1,M3 or M183. due to the production of pyrogenic exotoxins –
exotoxin A
desquamation of skin occurs 10-14
days after infection at sites that were
erythematous during the initial
phase
3a. Rheumatic fever1. most serious sequelae of hemolytic
streptococcal infection because it results in damage to heart muscle and valves
2. occurs 2 weeks after a group A streptococcal infection usually a pharyngitis
3. results in a systemic inflammatory process involving the connective tissue, heart, joints, and CNS
4. Characterized by fever, migratory polyarthritis, and carditis
5. Due to an immunologic reaction between cross-reacting antibodies to certain streptococcal M proteins and antigens of joint and heart tissue
6. Treat promptly with penicillin which is continued prophylactically to prevent recurrence and increased damage
Revised Jones Criteria for the Diagnosis of Rheumatic Fever
The diagnosis of rheumatic fever is highly likely if supported by evidence of a preceding group A streptococcal infection and the presence of two major manifestations or one major and two minor manifestations.
Supporting evidence of antecedent group A streptococcal infection
Positive throat culturePositive streptococcal antigen testElevated or rising streptococcal antibody titer
Major Manifestations
CarditisPolyarthritisChoreaErythema marginatumSubcutaneous nodules
Minor manifestations
Clinical findings: arthalgia, feverLaboratory findings
Elevated acute phase reactants (erythrocyte sedimentation rate, C-reactive protein)
Prolonged PR interval on elevtrocardiography
3b. Acute glomerulonephritis1. Typically occurs 2-3 weeks after streptococcal
skin infections with M types 2,4,12 or 49 (most frequent)
2. More frequent after skin infections than after pharyngitis
3. Characterized by hypertension, edema of the face(especially periorbital edema) and ankles, & “smoky” urine
4. Complete recovery; reinfection with streptococci rarely leads to recurrence.
5. Initiated by deposition of soluble streptococcal antigen-antibody complexes and complement on the glomerular basement membrane – lumpy-bumpy pattern on immunofluorescence
6. Can be prevented by early eradication of nephritogenic streptococci from skin colonization sites
Diseases produced by Streptococcus agalactiae
Neonatal Group B streptococcal disease
LABORATORY DIAGNOSIS
I. Streptococcus pyogenes
1. Microscopy – Gram stain
Gram stain of streptococci in a positive broth culture Gram positive
cocci in chains
2. Cultures
Colonies of group A streptococci on 5% sheep
blood agar small colonies with
a wide zone of beta hemolysis
Group A StreptococcusStreptococcus selective agar – contains sulfamethoxazoleAnd trimethoprim which inhibits the growth of nongroupA betahemolytic streptococci, staphylococci, viridansStreptococci and gram negative bacilli
Left – blood agar plate; right – streptococcus selective agar
PYR TestThe presence of an aminopeptidase enzyme that degrades the substrate is a 10 minute presumptive test for Group A streptococci (beta hemolytic) and Aerococcus, Enterococcusand Gemella(alpha or nonhemolytic)
left(+) colorless; right (+) red color
Bacitracin susceptibility test
An alternative to PYR test for the presumptive identification of Group A beta hemolytic streptococci 0.04 units of bacitracin disk is placed on an inoculum of the microorganism on sheep blood agar
Positive test – zone of inhibition
II. Streptococcus agalactiae - Group B beta hemolytic streptococcus
1. Microscopy- Gram stain of vaginal secretionsGram positive cocci in pairs,suggestive of Streptococcus agalactiae which colonize the genitourinary tract of women2. culture
Colonies of Group B streptococcus on 5% sheep blood agarColonies are larger than other beta hemolytic streptococci. Hemolytic zone surrounding the colony is smaller.Tests
1. Hippurate hydrolysis test
Incubate a suspension of themicroorganism for 2 hours at35 C in a hippurate solution.Add ninhydrin(indicator)
Hydrolysis of sodium hippurateleads to the formation of glycine and sodium benzoate.
Deamination of glycine –purple color
2. CAMP test (Christie,Atkins, Munich-Peterson) - An alternative to hippurate hydrolysis
Demonstrates the arrowhead shaped enhancement of betahemolysis that occurs when the hemolytic beta toxin producedby Staphylococcus aureus acts synergistically with the CAMP factor
III. Group D streptococci
1. enterococcus
Colonies of Enterococcus spp. on 5% sheep blood agarColonies are raised, white to gray white ranging from 0.5 to 1.5 mm. In size and are usually nonhemolytic
Tests
1. Bile esculin and 6.5% NaCl
- Bile esculin slant(left) – indicates that the microorganism can grow in the presence of bile and hydrolyze esculin.
2. 6.5% NaCl broth(right)- growth is indicated by turbidity and change in the indicator from pink to yellow
3. PYR Test
- The presence of an aminopeptidase enzyme that degrades the substrate PYR(L-pyrrolidonyl-B napthylamide) is a 10 minute presumptive test for group A streptococci(beta Hemolytic) and Aerococcus, Enterococcus and Gemella(alpha or nonhemolytic)
IV. Viridans streptococci
- Gram stain of a viridans streptococcus species in Blood culture broth appear in long chains especially when recovered from a blood culture broth
- Optochin susceptibility test. A paper disk containing optochin (ethylhydrocupreine Hydrochloride) is applied to the surface of an inoculated 5% sheep blood agar plate.
No zone of inhibition- Identification of viridans streptococci with
conventional biochemical reactions. Definitive identification requires several substrates including Bile esculin, arginine decarboxylase, 6.5% NaCl, lactose, Mannitol, raffinose, sorbitol, arabinose, inulin, sucrose and Esculin.
TREATMENT1. All group A streptococci are susceptible to
penicillin G.- mild - oral penicillin V
- if allergic: erythromycin or its derivatives (azithromycin)
2. Endocarditis caused by Viridans streptococci is curable by prolonged penicillin treatment
3. Enterococcal endocarditis – eradicated only by a penicillin or vancomycin combined with an aminoglycoside
4. Vancomycin resistant enterococci – linezolid (Zyvox) and quinupristin/dalforpristin(Synercid)
5. Nonterococcal Streptococcus bovis:penicillin G6. Group B streptococcal infections- penicillin G or
ampicillin in combination with an aminoglycoside
7. Peptostreptococci – penicillin G
PREVENTION1. Rheumatic fever can be prevented by prompt
treatment of Group A streptococcal pharyngitis with penicillin.
2. Penicillin prophylaxis for acute rheumatic fever patients to prevent recurrence of the disease; not needed in acute glomerulonephritis
3. In patients with damaged heart valves who undergo invasive dental procedures, endocarditis can be prevented by using amoxicillin perioperatively.
4. In patients with damaged heart valves who undergo gastrointestinal or urinary tract procedures, endocarditis caused by enterococcus can be prevented by using ampicillin and gentamicin perioperatively.
5. Neonatal sepsis caused by group B streptococci can be prevented by administration of parenteral ampicillin perinatally to women who experience prolonged (longer than 18 hours) rupture of membranes, whose labor begins before 37 weeks gestation or who have a fever at the time of labor.
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