STREPTOCOCCI - HUMSC
Transcript of STREPTOCOCCI - HUMSC
STREPTOCOCCI
Microbiology Lecture 2 RS Module
Ashraf KhasawnehFaculty of Medicine
The Hashemite University
Streptococci
• Characters of Streptococci
– Gram positive round or lancet-shaped cocci
– 1µm in diameter
– Chains or pairs
– Usually capsulated
– Non motile
– Non spore forming
– Facultative anaerobes
– Catalase negative (Staphylococci are catalase positive)
Classification of Streptococci
• Streptococci can be classified according to:
– Oxygen requirements
• Anaerobic (Peptostreptococcus)
• Aerobic or facultative anaerobic (Streptococcus)
– Serology (Lanciefield Classification)
– Hemolysis on Blood Agar (BA)
Serology: Lanciefield Classification
Streptococci
Group A
S. pyogenes
Group B
S. agalactiae
Group C
S. equisimitis
Group D
Enterococcus
Other groups
(E-U)
Lanciefield
classification
Streptococci classified into many groups from A-K & H-V
One or more species per group
Classification based on C- carbohydrate antigen of cell wall
Groupable streptococci
A, B and D (more frequent)
C, G and F (Less frequent)
Non-groupable streptococci
S. pneumoniae (pneumonia)
viridans streptococci
e.g. S. mutans
Causing dental carries
Classification of Streptococci Based
on Hemolysis on Blood Agar
Streptococci
-hemolysis -hemolysis -hemolysis
Hemolysis on BA
– -hemolysis
Partial hemolysis
Green discoloration around the colonies
e.g. non-groupable streptococci (S. pneumoniae & S. viridans)
– -hemolysis
Complete hemolysis
Clear zone of hemolysis around the colonies
e.g. Group A & B (S. pyogenes & S. agalactiae)
– -hemolysis
No lysis
e.g. Group D (Enterococcus spp)
Hemolysis on Blood agar
-hemolysis
-hemolysis
-hemolysis
Group A streptococci
• Include only S. pyogenes
• Group A streptococcal infections affect all ages peak
incidence at 5-15 years of age
• 90% of cases of pharyngitis
Pathogenesis and Virulence Factors
• Structural components– M protein M, which interferes with opsonization and lysis of the
bacteria
– Lipoteichoic acid & F protein adhesion
– Hyaluronic acid capsule, which acts to camouflage the bacteria
• Enzymes– Streptokinases
– Deoxynucleases
– C5a peptidase
• Pyrogenic toxins that stimulate macrophages and helper T cells to release cytokines
• Streptolysins (pore forming cytotoxin)– Streptolysin O lyse red blood cells, white blood cells, and platelets
– Streptolysin S Oxygen stable
facilitate the spread of streptococci through tissues
Disease caused by S. pyogenes
• Suppurative
– Non-Invasive• Pharyngitis (“strep throat”)-inflammation of the pharynx
• Skin infection, Impetigo
– Invasive• Scarlet fever-rash that begins on the chest and spreads across the body
• Pyoderma-confined, pus-producing lesion that usually occurs on the face, arms, or legs
• Necrotizing fasciitis-toxin production destroys tissues and eventually muscle and fat tissue
• Non Suppurative
– Rheumatic fever: Life threatening inflammatory disease that leads to damage of heart valves muscle
– Glomerulonephritits• Immune complex disease of kidney
• inflammation of the glomeruli and nephrons which obstruct blood flow through the kidneys
Differentiation between -hemolytic
streptococci
• The following tests can be used to differentiate
between -hemolytic streptococci
– Lanciefield Classification
– Bacitracin susceptibility Test
• Specific for S. pyogenes (Group A)
– CAMP test
• Specific for S. agalactiae (Group B)
Bacitracin sensitivity
• Principle:– Bacitracin test is used for presumptive
identification of group A
– To distinguish between S. pyogenes(susceptible to B) & non group A such as S. agalactiae (Resistant to B)
– Bacitracin will inhibit the growth of gp A Strep. pyogenes giving zone of inhibition around the disk
• Procedure:– Inoculate BAP with heavy suspension of
tested organism
– Bacitracin disk (0.04 U) is applied to inoculated BAP
– After incubation, any zone of inhibition around the disk is considered as susceptible
CAMP test
• Principle:
– Group B streptococci produce extracellular protein (CAMP factor)
– CAMP act synergistically with staph. -lysin to cause lysis of RBCs
• Procedure:
– Single streak of Streptococcus to be tested and a Staph. aureus are made
perpendicular to each other
– 3-5 mm distance was left between two streaks
– After incubation, a positive result appear as an arrowhead shaped zone
of complete hemolysis
– S. agalactiae is CAMP test positive while non gp B streptococci are
negative
CAMP Factor TestS. aureus
(Spingomyelinase C)Group B
Streptococcus
(CAMP Factor)
Group A
Streptococcus
Enhanced Zone of
Hemolysis
CAMP is an acronym for the authors of this test (Christie, Atkinson, Munch, Peterson). The CAMP
test takes advantage of the capacity of GBS-group B strep to produce this CAMP factor; most other
hemolytic streptococci do not produce CAMP factor. Enhances the ability of S. aureus to produce
Beta hemolysis
Differentiation between -hemolytic
streptococci
• The following definitive tests used to differentiate between S. pneumoniae & viridans streptococci
– Optochin Test
– Bile Solubility Test
Optochin Susceptibility Test
• Principle:
– Optochin (OP) test is presumptive test that is used to identify S. pneumoniae
– S. pneumoniae is inhibited by Optochin reagent (<5 µg/ml) giving a inhibition zone ≥14 mm in diameter.
• Procedure:
– BAP inoculated with organism to be tested
– OP disk is placed on the center of inoculated BAP
– After incubation at 37oC for 18 hrs, accurately measure the diameter of the inhibition zone by the ruler
– ≥14 mm zone of inhibition around the disk is considered as positive and ≤13 mm is considered negative
• S. pneumoniae is positive (S) while S. viridans is negative (R)
Optochin Susceptibility Test
Optochin susceptible
S. pneumoniae
Optochin resistant
S. viridans
Bile Solubility test
• Principle:
– S. pneumoniae produce a self-lysing enzyme to inhibit the growth
– The presence of bile salt accelerate this process
• Procedure:– Add ten parts (10 ml) of the broth culture of the organism to be
tested to one part (1 ml) of 2% Na deoxycholate (bile) into the test tube
– Negative control is made by adding saline instead of bile to the culture
– Incubate at 37oC for 15 min
– Record the result after 15 min
Bile Solubility test
• Results:
– Positive test appears as clearing
in the presence of bile while
negative test appears as turbid
– S. pneumoniae soluble in bile
whereas S. viridans insoluble
Differentiation between -hemolytic streptococci
CAMP testBacitracin
sensitivity
Hemolysis
NegativeSusceptibleS. pyogenes
PositiveResistantS. agalactiae
Inulin
Fermentation
Bile
solubility
Optochin
sensitivity
Hemolysis
Not fermentSolubleSensitive (≥
14 mm)
S. pneumoniae
FermentInsolubleResistant
(≤13 mm)
Viridans strep
Differentiation between -hemolytic streptococci
Outline of differentiation between Gram-
Positive cocci
e.g. S. epidermidis
Clinical Infections
- Diverse group of acute suppurative (pus-forming) & nonsuppurative diseases
Skin and Soft tissue infections
A- Cellulitis
Streptococcal cellulitis is a spreading infection →bacteraemia
B- Erysipelas
Acute spreading skin lesion with lymphatic involvement
Marked oedema & erythema of skin; face and lower extremities, skin and subcutaneous tissues
C- Impetigo:
Contagious pyoderma with superficial yellow weeping lesions) - children
D- Necrotizing fasciitis: (a.k.a., “flesh-eating bacteria”):
Infection deep in subcutaneous tissues that spreads along fascial
planes, destroying muscle and fat; Initially cellulitis followed
by bullae (fluid filled blisters; bulla is singular), gangrene,
systemic toxicity, multiorgan failure and mortality in more than
50% of patients
E- Wound Infections
Erysipelas
NOTE:
erythema
bullae
Erysipelas is a type of
superficial cellulitis with
dermal lymphatic
involvement. Diagnosis
is clinical. Treatment is
with oral or IV
antibiotics.
Erysipelas is characterized
clinically by shiny, raised,
indurated, and tender
plaques with distinct
margins. High fever, chills,
and malaise frequently
accompany erysipelas.
There is also a bullous
form of erysipelas.
Flesh Eating Bacterium
Pictures courtesy of e-medicine
A patient out of the surgery room
after a flesh-eating bacteria
disease infected his leg.
An arm infection.
Streptococcus Pyogenes Clinical Infections
Upper Respiratory tract Infections
A- Tonsillitis / Pharyngitis
The commonest bacterial cause of tonsillitis
B- Scarlet fever
Streptococcal sore throat (Children < 10 years)
Incubation period is 2-3 days
Generalized erythema & Strawberry tongue
Scarlet fever: Complication of streptococcal pharyngitis when infecting strain is lysogenized; Frequently develop scarletina rash on upper chest spreading to extremities
C- Acute Otitis media & Sinusitis
tonsillitisOtitis media
Scarlet fever
Other Suppurative Diseases
•Puerperal & neonatal sepsis (child bed fever)
•Lymphangitis: Inflammation of lymphatic vessel(s)
•Pneumonia
Systemic Disease
•Streptococcal Toxic Shock Syndrome (TSS):Multisystem toxicity following soft tissue infection
progressing to shock and organ failure (not to be confused
with Staphylococcal Toxic Shock Syndrome where
hyperabsorbent tampons have been identified as an
important risk factor)
•Bacteremia
Group A Streptococcal Diseases
Nonsuppurative Sequelae
Post-infection complications of Group A streptococcal disease; Serious
complications in pre-antibiotic era; still important in developing countries
Acute rheumatic fever (ARF):
Inflammation of heart, joints, blood vessels, sub-cutaneous tissues
Rheumatic heart disease (RHD):
Chronic, progressive heart valve damage
Acute glomerulonephritis (AG):
Acute inflammation of renal (kidney) glomeruli
Nonsuppurative Sequelae of Acute Group A
Streptococcal Infection
Acute Rheumatic Fever (ARF)
ARF follows respiratory, not skin, infection
Inflammatory reaction characterized by arthritis, carditis, chorea (disorder of
CNS with involuntary spastic movements), erythema marginatum (skin redness
with defined margin), or subcutaneous nodules
Within 1-5 weeks following pharyngitis (Children 6-15 years)
• Epidemic pharyngitis: ARF in as many as 3%
• Sporadic pharyngitis: ARF in 1 per 1000
Morbidity & mortality linked to subsequent disease of heart valve (Rheumatic
Heart Disease)
Poorly understood pathogenesis with several proposed theories including cross-
reactivity of heart tissues & strep AGNs (M protein)
Nonsuppurative Sequelae of Acute Group A
Streptococcal Infection
Acute Glomerulonephritis
Follows either respiratory (pharyngitis) or cutaneous (pyoderma)
streptococcal infection
Associated with well-defined group of M-types
Incidence varies from <1% to 10-15%
Most often seen in children manifesting as dark, smokey urine with RBC's,
white blood cells, depressed serum complement, decreased glomerular
filtration rate (Oedema, Hypertension, Haematuria & Proteinuria)
Latent period: 1-2 weeks after skin infection and 2-3 weeks after pharyngitis
Granular accumulations of immunoglobulin due to deposition of immune
complexes within the kidney
Lab Identification of
S. pyogenes (Group A)
• Primary culture
• Domed grayish/opalescent
colonies
• Encapsulated cells produce
mucoid colonies
Beta-hemolytic• Zone several times greater than diameter of colony
Lab Identification of
S. pyogenes (Group A)
Catalase Negative: Differentiates from Staphylococcus
Bacitracin test: presumptively distinguishing between Group
A beta-hemolytic streptococci (bacitracin POS) and other beta-
hemolytic streptococci that are isolated from pharyngeal swabs
(95% sensitivity for Grp A strep)
When grown on blood agar, Group A streptococci are sensitive to
(killed by) the antibiotic bacitracin
Rapid Identification Tests:Based on extraction of Group A carbohydrate directly from
throat swabs
• ELISA, Coagglutination, Fluorescent Antibody
Streptococcus Pyogenes
Treatment All GAS are sensitive to penicillin G (no
resistance)
Pts allergic to penicillin, treat with Erythromycin or Azithromycin
Adequate treatment of strep pharyngitis within 10 days of onset prevent rheumatic fever but not glomerulonephritis.
Prevention of GAS
Penicillin prophylaxis with long acting preperations after primary attack of ARF (5-15 yrs)
No prophylaxis in case of AGN
Group B Streptococcus
Streptococcus agalactiae
Gram positive cocci in chains
Beta hemolytic
Catalase –ve
Bacitracin Resistant
CAMP test positive
Group B Streptococci:
Streptococcus agalactiae
•Major cause of neonatal sepsis
• 1 in 4 pregnant women are vaginal or
rectal carriers of GBS
• 50% of women transmit the disease to
infant (42-72%)
•Infect neonate during delivery
•1-2% colonized infants develop Group
B strep infection
•Induces pneumonia in the first 7 days
of life
• Later may induce septicemia ,
osteomyelitis and meningitis
Epidemiology of Neonatal Group B
Streptococcal Disease
Prevention /Detection
- 2002 WHO universal screening for GBS colonization by vaginal and rectal swab culture for pregnant women at 35-37 weeks gestation.
Lab confirmation
- Blood and CSF Culture , gram stain
- CAMP test
- Hippurate test (positive)
Treat GBS positive mothers with Penicillin G, Ampicillin or Eryrthromycin
Group D Streptococci (S.bovis)
– Gram-positive cocci,
– α, β or non-haemolytic on blood agar.
– Lancefield group D streptococci will grow on media containing bile and may be differentiated from other
streptococci by rapid hydrolysis of esculin in the presence of 40% bile. Bile esculine postive
• Endocarditis
Non-Lancefield Group Streptococci
Viridans StreptococciDental Caries: Streptococcus mutans
Streptococcus sanguis; Streptococcus
salivarius; Streptococcus mitis
Streptococus pneumoniae – # 1 cause of
community acquired pneumonia
Resistant to optochin
Streptococcus pneumoniae
• Commonly referred to as pneumococcus
• Formerly Diplococcus pneumoniae
• Diplococci lancet shaped
• Alpha hemolytic
• Optochin sensitive
S. pneumoniae
• Diplococcus
Virulence factors
• Colonization and migration factors
– IgA protease: help in colonization
– Pneumolysin- Pore forming toxin
• Tissue destruction
– Teichoic acid: colonization and dissemination processes
– Hydrogen peroxide: DNA damage and apoptosis in lung cells
– Pneumolysin- epithelial cell destruction
• Phagocytic survival
– Capsule – antigenic variation avoid phagocytosis -90 serotypes
• Autolysin: assist in pathogenesis due to its ability to break down the wall or lyse a portion of the invading pneumococci and release potentially lethal toxins into the cell
• S. pneumoniae is a leading cause of community pneumonia in all ages (particularly the very young and old), often after "damage" to the upper respiratory tract (e.g. following viral infection).
RUST COLOUR SPUTUM• Fever and chills. Difficulty breathing, cough
productive of purulent sputum and sometimes blood tinged, pleuritic chest pain
• It also causes middle ear infections (otitismedia).
• The organism often spreads causing bacteremiaand meningitis.
Lack structural damage to the lung
Complete resolution on recovery
Immunity
• Provided by antibody directed against the specific pneumococcal capsular type
• After ab binding, C3b is deposited and phagosytosis can take place.
• Diagnosis
• Gram stain: +ve diplococci
• Blood agar: alpha hemolysis, optochin sensitive
• Blood culture
• Detection of capsular antigen
Streptococcus pneumoniae
Diagnostic Laboratory Tests
• Optochin sensitivity (Taxo P disc)
Treatment
• Penicillin 1st choice
• First-generation cephalosporins are equally effective against infections caused by penicillin-susceptible S pneumoniae. All first-generation cephalosporins are equally effective against pneumococci;
• For empiric treatment of meningitis, third generation
cephalosporins may be administered in conjunction with vancomycin or rifampin.
• Azithromycin has activity against penicillin-susceptible strains of S pneumoniae.
Prevention
VACCINE
• Given the 90 different capsular types of pneumococci, a comprehensive vaccine based on polysaccharide alone is not feasible.
• Thus, vaccines based on a subgroup of highly prevalent types have been formulated. (PPV)
• The number of serotypes in the vaccine has increased from four in 1945, to 14 in the 1970s, and finally to the current 23-valent formulation (25 mg of each of serotypes 1, 2, 3, 4, 5, 6B, 7F, 8, 9N, 9V, 10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, and 33F).
• These serotypes represent 85-90% of those that
cause invasive disease and the vaccine efficacy is estimated at 60% .
• T-cell independent; used in pts >2yrs• In 2000: PCV, 7-valent vaccine stimulte TH2
response and can be given at 2months. At 2 yrs give PPV
• Underutilization of the vaccine is so extensive that the pneumococcus remains the most common infectious agent leading to hospitalization in all age groups due to cost
• Further complicated by the fact that polysaccharides are not immunogenic in children under the age of 2 years where a significant amount of disease occurs
Immunization protocol
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3324398/
• Polysaccharide antigens are postulated to stimulate splenic marginal zone B (MZB) cells, which do not mature until the second year of life [9]; therefore, the purified polysaccharide contained in 23vP, a T-independent antigen, is poorly immunogenic in young children. Chemical conjugation of pneumococcal polysaccharide to a carrier protein creates a T-dependent vaccine (pneumococcal conjugate vaccine [PCV]) that generates higher affinity antibodies, immunological memory, and induces responsiveness to booster doses of vaccine, resulting in a vaccine that is both immunogenic and highly effective from early infancy [10]. Because the splenic marginal zone is immature in early life, MZB cell responses are not present, and it is postulated that the conjugated polysaccharides in PCV are processed by the follicular origin (FO) B cells at that age [11]. Despite the immunological advantages of PCV in early childhood, both PCV7 (a 7-valent PCV) and 23vP induce similar antibody concentrations in adults [12], and it is therefore unclear whether the conjugate vaccine has any immunological advantage over 23vP or whether the same B-cell subsets are involved in the response.