24 feb STREPTOCOCCUS.ppt
- 1. Introduction ▰ Pharyngitis (or sore throat) - most common upper respiratory tract infections (URTI). ▰ Viral pharyngitis - vast majority of cases – self-limited. ▰ Bacteria - important etiologic agents of pharyngitis, require specific antibiotic treatment - can lead to serious complications and sequelae
- 2. Bacterial Pharyngitis DR MAYURI BHISE
- 3. Etiological agents Streptococcus pyogenes Corynebacterium diphtheriae ▰ Rare causes Other β-hemolytic streptococci (group C and G) Arcanobacterium hemolyticum Fusobacterium necrophorum Mycoplasma pneumoniae Neisseria gonorrhoeae.
- 4. S T R E P T O C O C C U S
- 5. History Billroth, 1874 Ogston, 1881 Rosenbach – 1884. 5
- 7. O2 requirement Aerobes & facultative anaerobes Anaerobes Hemolysis on 5% horse B.A. - hemolytic (viridans) - hemolytic (pathogens) - or non-hemolytic (fecal streptococci) Serogrouping by Rebecca Lancefield (1933) - based on cell wall cabohydrate, A-W (except I & J) Group A – Strepto. pyogenes Serotyping based on M, T, R proteins. >100 Griffith types 7
- 9. Morphology Size – 0.5-1 m. Shape – oval /elliptical. Arrangement – in chains esp. in liquid culture media. (upto 50 cocci in a chain). Divide in one plane. Daughter cells do not seperate. Gram positive Cultures older than log phase may lose gram reaction. 9
- 11. Capsule – • Hyaluronic acid (group A, C). Nonimmunogenic. • Polysaccharide (group B, D ). Nonmotile. Nonsporing. L-forms – cell-wall deficient, require thiol & pyridoxal for growth. Found in blood (due to antibiotics). 11
- 12. Cultural characters Aerobes & facultative anaerobes. 22-42°C; opt.37. pH for growth –opt. 7.4. Capnophilic – 10% CO2. Fastidious; need blood / serum / sugar. Liquid medium (e.g. Todd-Hewitt broth) – granular turbidity + powdery deposits. 12
- 13. Blood agar – • 0.5-1mm, circular, low convex, -hemolytic. • Matt colonies – pathogenic •Glossy colonies – nonpathogenic. Selective medium – •Crystal violet (1:500,000) in B.A. 13
- 14. Biochemical reactions Catalase – ve Sugar fermentation – Glucose, lactose, maltose, trehalose - . (constitutive enzymes). Other sugars & alcohols - ,(inducible enz.). *Ribose sugars – not fermented. *Pyrolidonyl naphthalamide hydrolysis (PYR) - +ve (differentiates gr. A from other groups). 14
- 15. Resistance / viability Delicate organism Survives in dust in dark for many weeks. Susceptible to heat , 54°C x 30 min. Susceptible to common antiseptics. Resistant to – Crystal violet (1mg/litre) – for isolation of gr A. Nalidixic acid (15mg/litre) for isolation of & Colistin (10mg/litre) gr. B Bacitracin sensitive – differentiates gr. A from other hemolytic streptococci. 15
- 16. Antigenic structure Hyaluronic acid capsule- nonantigenic, antiphagocytic, thrombolytic. •Innermost cell wall layer peptidoglycan (NAG-NAM). •Middle layer of cellwall carbohydrate. •Outer layer of proteins, lipoteichoic acid. 16
- 17. Cell-wall CHO – Group sp. Ag. Todd-Hewit growth → Extraction by – HCl (Lancefield) Formamide (Fuller) Enzyme of Streptomyces albus (Maxted) Autoclave (Rantz & Randall). Capillary pptn /ring pptn 17
- 18. M protein – Acid extraction & Serotyping. 100 M-types. T protein – Typing by slide agglutination with trypsinised RBCs. R protein – Present in gr. B, C, G & some serotypes of gr. A (23, 28, 48). 18
- 19. Antigenic cross-reactions Capsular hyaluronic acid synovial fluid. Cellwall CHO cardiac valves. Cellwall peptidoglycan skin. Cyto. membrane vascular intima & cardiac muscle. 19
- 20. Virulence factors A) cellular M protein* - reqd. for invasive infection. Receptor for fibrinogen, factor H, IgG. Capsule - antiphagocytic Group CHO Ag’-invasive properties. Protein F- binds fibronectin on epi. Surface Lipoteichoic acid loosely binds strepto’ to epi surface. F prtn, M prtn secures adhesion. 20
- 21. Virulence factors B) Exotoxins Streptococcal pyrogenic exotoxins (SPE) / erythrogenic toxins / Dick toxins – A, B, C. Act as superantigens. Excessive release of cytokines & interleukins from T-cells → inflammatory shock (TSS). Hemolysins – Streptolysin O & S. 21
- 22. Streptolysin O Oxygen labile Broth culture + sod. hydrogen sulphite Cardiotoxic Freely secreted Antigenic ASO titer imp. in rheumatic fever Streptolysin S Oxygen stable Broth culture + RNA Nephrotoxic Cell bound Nonantigenic Stimulates release of lysosomal enzymes. But exact pathogenic role unknown 22
- 23. Virulence factors C) Enzymes Streptokinase – dissolves fibrin wall around site of inflammation → spread of streptococci. Hyaluronidase – Helps in spread. (spreading factor). Nuclease /DNAse /Streptodornase Dissolve thick pus & help in spread. Neuraminidase – helps in mucosal colonisation. 23
- 24. Host defenses First line defense / nonspecific defense – Barrier of skin Mucociliary defense in resp. tract. Cough & sneeze reflex. Second line defense – Complement activation. Antistrepto. antibodies in serum. Opsonization & phagocytosis. Autophagy of phagocyte if killing impossible. 24
- 25. Epidemiology Reservoirs – widely distributed in nature. + Part of normal flora. Sources – patients & carriers. Transmission – a) droplets / contaminated dust → URTI; b) Milk → bovine mastitis; c) fomites, insects (eye gnats) → skin infn. Age – two peaks; a) 5-8 yrs. b) at 12-14 yrs. 25
- 26. Sex – males > females. Season – winter in temperate climates; all seasons in tropics. Predisposing factors – Crowding Medical problems Immunity – associated with antibody to M-protein. 26
- 27. Diseases A) Suppurative Pharyngitis , tonsillitis, URTI. By earlier serotypes. Swollen, red post. pharyngeal wall. Complications – peritonsillar & retro- pharyngeal abscess, sinusitis, otitis media, mastoiditis etc. 27 Streptococcal sore throat, with enlarged tonsils covered with white fibrinous exudate
- 28. Skin & soft tissue infections By later serotypes. Common in small children. Impetigo – Small papules, vesicles, pustules → thick crusts Ecthyma – Ulcerated lesion (staphylococcal lesions produce larger vesicles & bullous ulcers.) 28
- 29. Erysipelas In older children. Red, swollen, edematous, indurated skin with sharply demarcated lesions. Repeated attacks. Due to SPE- B & C. 29
- 30. Cellulitis – In skin with breach in continuity. Lesion – Red, swollen, painful Not raised or demarcated. Lymphangitis, bacteremia common. 30
- 31. Necrotizing fascitis / streptococcal gangrene – Skin with breach in continuity. 1-2 days – erythema & purple bullae. 7-10 days – Demarkated lesion. Skin breaks open. Underlying necrosed tissue seen. 31
- 32. Streptococcal toxic shock syndrome – Wound infections of skin due to type M1, M3. Due to SPE- A, B, C. Super antigens. → inflammatory shock, renal failure & ARDS. Extensive necrosis of skin & s/c muscles. 32
- 33. Other invasive infections – Puerperal sepsis. Anaerobic infections cause sepsis. Abscess formation in kidney, brain & other organs. 33
- 34. Diseases B) Nonsuppurative Acute rheumatic fever & glomerulonephritis. Both seen 1-3 weeks after ac. strepto. infn. RF seen after pharyngitis but not after cut. Inf. GN seen after dermal infections No virulence factor of streptococcus is solely responsible for both the diseases. 34
- 35. Rheumatic fever Pathogenesis Molecular mimicry & autoimmunity – M protein Heart Membrane antigen Skin & joints Group sp. CHO Brain Genetic predisposition of some B-alloantigens Pathology Thick mitral, aortic & tricuspid valves. Thick chordae tendinae. Thick myocardium. 35
- 36. Thick mitral valve. Thick chordae tendinae. Thick myocardium. Thickened & calcified Aortic valves due to RHD. 36
- 37. Histopathology Aschoff nodule In interstitium around vessels Rosette of giant cells around central necrosis. 37
- 38. Diagnosis – Jone’s criteria Major criteria – • Arthritis • Carditis • Chorea • Erythema marginatum • Subcutaneous. nodules Minor criteria – • Arthralgia • Fever • Raised ESR • Raised CRP • Increased P-R interval in ECG. Antecedent streptococcal infection – positive culture, ag’ detection, rising ab’ titre (ASO). 38
- 39. Glomerulonephritis Pathogenesis – Molecular mimicry & autoimmunity Protein antigens of nephritogenic streptococci endostreptosin under glomerular B.M. Immune complex disease with deposition of IgG + complement under glomerular basement memb. Features – Pallor, edema, hypertension, hypoalbuminemia, rusty/ smoky urine with ↑proteins, RBCs & WBCs. Self-limiting. Resolves without any permanent damage. 39
- 40. Bowman’s capsule
- 41. Differences ACUTE RHEUMATIC FEVER ACUTE GLOMERULONEPHRITIS Site of infection Throat Throat / skin Prior sensitization Essential Not necessary Serotypes Any Pyodermal types Immune response Marked Moderate Complement level Unaffected Lowered Genetic susceptibility Present Not known Repeated attacks Common Absent Penicillin prophylaxis Essential Not indicated Course Progressive / static Spontaneous resolution Prognosis variable good
- 42. Differential diagnosis ▰ Diphtheria and Viral pharyngitis ▰ Scarlet fever: Mediated by streptococcal pyrogenic exotoxins (e.g. SPE-A, B, and C) ▰ Characterized by pharyngitis and rashes (with sandpaper feel), strawberry tongue (enlarged papillae on a coated tongue). Rashes in the skin folds - Pastia’s lines
- 43. Laboratory diagnosis Specimens – Throat swabs, pus, blood, CSF/body fluids, infected tissues from cellulitis/ erysepelas. Collection – Under vision with aseptic precautions in a sterile container. Transport – Immediate / immediate plating on B. A. Transport medium – Pike’s medium (B.A.+ C.V. +Na- azide). 43
- 44. Smears (Gram stain) – Pus & body fluids. No value for throat & genital specimens. Culture – Sheep B. A. without NAD (H. influenzae can’t grow). Incubation in 5-10% CO2. Colony morphology & hemolysis. Selective medium – sheep blood agar + cotrimoxazole or crystal violet (1:500000). Biochemical reactions – Catalase –ve, 10% bile insoluble, sugar fermentation except ribose, PYR +ve. 44
- 45. Grouping & typing Bacitracin sensitivity 6 mm disc, 0.1unit of bacitracin. Zone of inhibition, ≥ 11 mm diameter. Lancefield grouping Griffith typing Phage typing 45
- 46. Antigen detection in samples – Antigen extraction by pronase / nitrous acid. Techniques – Latex agglutination test (LAT). Enzyme immunoassay (EIA). Liposome immunoassay (LIA). Fluorescent antibody staining. Rapid detection of Strepto. pyo. DNA hybridization 46
- 47. Serological diagnosis (retrospective diagnosis in RF & GN) – Antistreptolysin O- inhibit streptolysin O mediated RBC lysis. Titers high in RF; low in GN. Titers >200 Todd units significant. AntiDNAse B – inhibits DNAse B from hydrolyzing DNA-methyl green conjugate. 47
- 48. Other antibodies – Anti NADse Anti hyaluronidase Anti A-carbohydrate Rapid screening tests – Streptozyme test Passive hemagglutination (RBCs coated with streptococcal antigen) 48
- 49. OTHER TESTS FOR STR. PYOGENES 1) Fluorescent antibody technique 2) Typing by precipitation or agglutination 3) Rapid diagnostic test kits available 4) ASO titre estimation 5) AntiDNAseB estimation RAPID DIAGNOSTIC TEST KIT
- 50. Prophylaxis Life long prophylaxis for Rheumatic fever – Prevents reinfection & further damage to heart. Long acting penicillins, erythromycins, sulfadiazine (to prevent recurrent attacks). Not useful in c/o glomerulonephritis as it occurs after single infn and there is no reinfection. 50
- 51. Treatment Cellwall active agents used. Penicillins, macrolides (erythro’, azithro’), clindamycin, vancomycin, cephalexin. Surgical debridement in c/o myositis, necrotizing fascitis etc. For pharyngitis: Benzathine penicillin G, IM single dose or oral penicillin V for 10 days. For recurrent pharyngitis: Clindamycin or amoxicillin- clavulanate. For concomitant pneumonia and empyema: Penicillin G + drainage of empyema. 51
- 52. Other hemolytic streptococci 45% of total strepto. Isolates = Gr. A. 15% ,, ,, ,, ,, = Gr. B. 15% ,, ,, ,, ,, = Gr. C. 25% ,, ,, ,, ,, = Gr. G. 5 % ,, ,, ,, ,, = Gr. F. 52
- 53. Group B streptococci – Identified by hippurate hydrolysis, CAMP test. Hippurate benzene ring + serine Overnight growth in broth culture → Centrifuge → Deposit → Red ppt. on addition of FeCl3. Colony suspension in broth → Incubate for 4 hrs. → Purple colour on addition of ninhydrin. HIPPURICASE 53
- 54. CAMP test Staph. aur Gr. B Gr. A 54
- 55. Infections Bovine mastitis. Colonizes female genital tract → septic abortion & puerperal sepsis. Neonatal septicemia & meningitis. Early onset (within 1wk.) - vertical transmission from vagina. Late onset (2-12 wk.) – horizontal transmission from environment. Adult infection Skin & soft tissue infections, bone infections. Pneumonitis & septicemia. 55
- 56. Group C streptococci Animal pathogens. Strepo. equisimilis – human pathogen. Infections of deep tissues, bones, brain & heart. Important source of streptokinase (used in thrombolytic therapy). Group F streptococci Minute streptococci. Streptococcus MG - -h streptococci in PAP. Used in diagnosis of serological test of mycoplasma pneumonia. 56
- 57. Group D streptococci – Two types – Enterococcal group – Enterococcus fecalis, E. fecium. Nonenterococcal group – Strepto. bovis, Strepto. equinus. Pairs of oval cocci arranged at angles. Dark magenta colored coloured colonies on MacConkey’s agar; black colonies on BPT. Grow at 45°C, at pH 9.6, in 6.5% NaCl, in 40% bile; and hydrolyse esculin. 57
- 58. Bile Esculin Agar Bile Esculin Agar NEG POS 58
- 59. Viridans streptococci Oral streptococci. Grow at acidic pH. Produce dental plaques & caries. Tooth extraction → bacteremia → endocarditis affecting heart valves. 59