staphylococcus streptococcus revision notes microbiology
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staphylococcus streptococcus revision notes microbiology
- 1. Gram positive cocci Staph atrept
- 2. Micrococcus Staphylococcus Streptococcus G+ve in tetrads G+ve in cluster G+ve in chain Strict aerobe Aerobes & facultative anaerobes Aerobes & facultative anaerobes Catalase +ve Catalase +ve Catalase –ve Oxidase +ve Oxidase –ve Oxidase –ve
- 3. Hugh leifson Oxidative fermentative media
- 4. Hugh leifsons OF medium Micrococcus strict aerobic oxidative reaction Streptococcus & staphylococcus (facultative anerobes) fermentative
- 6. staphylococci • G +ve • Grape like cluster arrangement • Division along all three planes • Facultative anaerobes • Microcapsule +/-
- 7. • Catalase +ve • Oxidase –ve • Ferment sugars
- 8. Culture of staphylococcus • Blood agar • Beta hemolysis (complete hemolysis) • Nutrient agar • Golden yellow colur • d/t beta carotene production (staphyloxanthine)
- 9. Increased pigment production in staph • Incubared @22*C • Aerobic conditions • Tween 80/1% glycerol monoacetate / milk
- 11. Selective media • Selective medium • – Salt milk agar (8-10% NaCl) • Most staphylocooi can grow in presence of 10 % NaCl • – Mannitol salt agar-with 7.5% NaCl • – Ludlam’s lithium chloride and tellurite media
- 12. Blood agar Pin head beta haemolytic McConkey Tiny lactose fermenting Transport medium Robertson cooked meat broth media Selective • 10 % salt agar • Ludlam media • Salt milk agar • Baird parker media Selective & differential • Mannitol salt agar (s aureus only species fermenting mannitol)
- 14. Phage typing • MC method for typing of S.aureus Phage typing (pattern method) • MC epidemic phage type in hospital- 80/81 • 23 standard types of phages are used
- 15. Biochemical reaction • All staphylococci • Catalase +ve • Streptococci gram –ve • Staph aureus • Coagulase +ve • Detected by tube coagulation test • Ferments mannitol • Heat stable mannitol • Phenolphthaein phosphatase+ • Lysostaphin sensitive • UREASE + / DNAse+/reduce nitrates
- 17. Tube coagulase test • For tube coagulation CRF is needed
- 19. Slide coagulase is d/t clumping factor
- 21. • Slide coagulase test • Tube coagulase test Both positive Both negative CoNSStaph aureus
- 22. Slide coagulation is d/t bound coagulase/clumping factor
- 25. Coagulase • Most important virulence factor • Coagulase Most common association of virulence with coagulase & lesser extent with mannitol fermentation • 2 forms • Bound form/ clumping factor • Responsible for adherence of organism • Acts directly on fibrinogen • Fibrinogen fibrin • Detected by slide coagulation • Coagulase / free coagulase • Excreted from cell surface as free enzyme • Eight types (A to H)
- 26. Free coagulase Bound coagulase
- 27. Hyalouronidase • Helps in persistent skin infection
- 28. • Hemolysins: • alpha • beta hot cold phenomenon • hemolytic properties are increased by exposure of the RBCs to cold temperature • Hemolysis starts @ 37*c but completed only at 4*C • gamma • Delta
- 29. α hemolysin β haemolysin γ hemolysin • Lethal • Dermonecrotic • Leucocidal • Paradoxical reactivation b/w 80- 100 *C • Sphingomyelinase in nature • Hot cold phenomenon • Does not damage human RBCs • Membrane damaging • Bicomponent synergo hymenotropic
- 30. Protein A • Antiapoptotic • Antiphagocytic • Used co-agglutination test • Binds to Fc portion of IgG
- 31. Panton valentine leucocidin • Dermonecrotic • Leucocidal in nature • Also synergohymenotrpic • Phage mediated lysogenic conversion • In community acquired MRSA
- 32. Synergohymenotropic agents • Secreted as Bicomponent & create pores in membrane • Gamma hemolysin and leucocidin (D-Panton-Valentine toxin) • Membrane-damaging and bicomponent toxins have been grouped as synergohymenotropic toxins.
- 33. Exfoliative / Epidermatolytic Toxin / ET / Exfoliatin • There are two type : ETA and ETB, • toxin possess serine protease activity which triggers exfoliation. • EFA chromosomally mediated • EFB plasmid mediated • Serine proteases Cleave desmoglein 1 in stratum granulosum staphylococcal scalded skin syndrome (SSS).
- 35. Staphylococal toxin mediated infection • Toxic shock syndrome • Scalded skin syndrome • Food poisoning
- 36. • Infection /colonisation of staph aureus EF A& EFB hematogenous dissemination staphylococcal scalded skin syndrome
- 37. • Severe form is called Ritter’s disease in neonate and toxic epidermal necrolysis in elderly. • Milder form are pemphigus neonatorum and bullous impetigo
- 39. Diagnosis of SSSS • ELISA / PCR for EF A or EFB in serum
- 40. Protein A • Used in co agglutination test for salmonella typhi • Protein A binds with Fab fragment of Ab against salmonella typhi
- 41. • Protein A binds with Fab fragment of IgG
- 42. Super Ag produced by staphylococci • TSST • Enterotoxin – B &C Pyrogenic toxinc A,B & C of streptococcus are also super Ag
- 43. TSST • Enterotoxin F • Belongs to pyrogenic super Ag • Super Ag
- 45. TSST • Menustral • It is seen in women using highly absorbent tamponade • TSST-1 = Enterotoxin F = Pyrogenic Exotoxin C is responsible for most cases • Non menustral • Enterotoxin B or C • Streptococcus pyogenes • TSST 1
- 51. Diagnosis of TSST • Detection of toxin in serum by ELISA / latex agglutination by PCR
- 52. Enterotoxin Food poisoning • Enterotoxin : (A, B, C1-3, D, E and H) • Type A toxin is responsible for most cases. • Chromosomally mediated • Preformed, heat stable toxin, • Not destroyed by cooking • meat and fish, milk or milk products. • Stimulates CTZ • No fever • Malaise N V Abdominal pain & diarrhea • 2-6 hrs after consuming (short incubation period) • Similar B cereus causing emetic type of food poisoning (short IP)(5-6 hrs) • Source - usually food handler which is carrier • Mechanism – • Toxin acts directly on autonomic nervous system (Vagal stimulation) and vomiting centre. • Other toxins act on mucosa
- 53. • Rx of staphylococcal food poisoning • Resolves with in 8 to 10 hours • Aymptomatic Rx only • no Abx needed
- 54. Normal carriage of staphylococci • Mc nose & oropharynx • Others • Skin • Hair • Perineum • Vagina
- 56. Staph aureus • 6 biotypes • A • Most human pathogenic strains • B • C • D • E • F
- 57. Skin & sof tissue • Skin and soft tissue infection MC infection by staph • Botryomycosis • Mycetoma like esion with multiple discharging sinuses • Also caused by strep pyogenes Ps aeruginosa E coli proteus • Folliculitis, furuncle, carbuncle, Mastitis and Hidradenitis suppurativa • Bullous impetigo • Ecthyma • Post operative wound infection
- 58. Musculoskeletal • Tropical pyomyositis – S. aureus, (acute bacterial myositis – Group A Streptococcus) (Overall - S.aureus) • Osteomyelitis and septic arthritis (MC- knee)
- 59. Respiratory tract infections • Pneumatocele MC cause • Other causative pneumococcus klebsiella pneumocystis • Pnemonia
- 60. Staphylococcus aureus is the most common cause of • Surgical wound infection • Postoperative parotitis • Bacterial endocarditis (native valve) • 2nd most common cause of BE in prosthetic valve after staph epidermidis • Endocarditis in IV drug abusers • Pneumatocele • Osteomyelitis • Tropical mysositis • Epidural abscess
- 61. • Most common cause of primary bacteremia staph epidermidis • 2nd most common cause staph aureus
- 62. Hospital infections • most common cause of hospital-acquired infections • Post operative wound infection • Phage 80/81 hospital strain • In case of outbreak • search for carriers (30 % of general population are carrier) • Mc in anterior nares • Also in vagina skin axilla perineum • Rx with topical chlorhexidine & mupirocin • Strict hand washing most economical & effective
- 63. • Most common source of infection • Endogenous infection from individuals own colonizing strains • Especially from skin
- 64. • 5th generation CP • Fifth-generation cephalosporins such as ceftobiprole and ceftaroline are effective against MRSA. • Ceftobiprole has additional activity against penicillin-resistant Streptococcus pneumoniae, Pseudomonas aeruginosa, and Vancomycin resistant Enterococci(VRE).
- 65. Mechanism of resistance to antibiotics
- 66. Mode of resistance • Production of beta lactamase MC • Plasmid mediated • Mc spread by transuction • Alteration in penicillin binding proteins • Development of tolerance
- 67. Production of beta lactamase • Plasmid mediated • Penicillinase plasmids are transmitted to the staphylococci by both transduction and conjugation • Mainly transduction • Four types of penicillinases (A, B, C, and D) • Hospital strains produce type A
- 68. Beta lactamase is produced by plasmid
- 69. Beta lactamase / penicillinase
- 70. Alteration of PBP • Chromosomal-mediated resistance • Large mobile genetic element SCC (staphylococcal cassete chromosome ) • It contains mec A gene codes for PBP2a • Major mechanism of methicillin resistance • MRSA • Methicillin resistance is more expressed @ 30 * c
- 72. Detection of MRSA • Dilution method • MIC > 4ug/ml • Disc diffusion method • Cefoxitin disc /oxacillin disc • Zone of inhibition < recommended
- 73. for expression of mec A gene • Best medium • Mueller Hilton agar with 2-4 % salt • Temperature • 30-33*C
- 74. • Gold standard for detetction of MRSA is PCR for mec A gene • Rapid diagnosis latex agglutination against altered PBP2a
- 75. Types of MRSA • Types of MRSA- • – Community acquired- • mediated by mec A subtype IV and Possess PV Toxin, • – Hosp acquired – • mediated by mecA subtype I,II,III, multi drug resistance
- 77. CA MRSA HA MRSA Current or Recent exposure • Hospital / nursing home • Rehabilitation facility • Dialysis centre • Indwelling central venmous catheter More virulent • Less virulent > 95 % a/w produce panton valentine toxin • No such association Isolated from staphylococcal skin infection or necrotising pneumonia • Normal flora / wound infection / blood stream infn Resistant to beta lactams • Often MDR
- 78. Detection of MRSA • Antibiotic susceptibility test on Muller Hilton agar (Disk diffusion test) • By using Methicillin or Oxacillin disc or cefoxitin disc • Cefoxitin disc diffusion more reliable than oxacillin for MRSA • Latex agglutination for PBP2a • Detection of mec A gene by PCR
- 79. • Conditions to be maintained- • – On media containing 4% NaCl, • – Incubation at 30°C, • More growth of MRSA AT 30 *C • – Full 24hr incubation
- 80. Rx • Isolation of patient • Wash hands after each patient • Mc mode of spread is via hands of health care workers • Penicillin sensitive • Penicillin G • MRSA • Vancomycin • VRSA • Sreptogrammins , linezolid
- 81. VRSA • High level of resistance d/t van A gene on a transposon acquired from VRE • Rx • Linezolid • Streptogramins • Daptomycin
- 82. CoNS
- 83. Resistant to novobiocin • S saprophyticus • S xyloses Beta haemolytic • S aureus Bound coagulase • S lugdunsesis Secrete coagulase • S intermedius
- 84. Staph epidermidis • Normal flora • Non pignmented / CoNS /nonhemolytic / absent mannitol fermentation/ novobiocin sensitive • Cause of early prosthetic valve endocarditis up to 12 months • Prediliction for grow on implanted Fb such as artificial vlaves shunts bacteremia • mC bacteremia in IV catheterised • Mc infection in orthopaedic implants • Mc dialysis associated peritonitis • Mc meningits in CSF shunts • Common cause of stitch absecess • Biofilm production • Rx’ • vancomycin
- 85. Staph saprophyticus • Skin & female genital tract infection • Non pignmented / CoNS /nonhemolytic / absent mannitol fermentation/ • Novobiocin resistant • UTI in young woman
- 86. Staph lugdunensis • CoNS (has bound coagulase) • Non pignmented / CoNS /nonhemolytic / absent mannitol fermentation/ novobiocin sensitive • Skin commensal
- 88. • Endocarditis • • MC cause of Native valve endocarditis – S.aureus • • MC cause of Prosthetic valve endocarditis • – Early prosthetic valve endocarditis (<12months) - S.epidermidis • – Late prosthetic valve endocarditis (>12months)- Strept Viridans • – Overall MC cause of prosthetic valve endocarditis- S.epidermidis • • MC cause of Endocarditis in IV drug users: - • – Rt sided – S. aureus, • – Lt sided – Enterococcus > S. aureus • – Over all-S. aureus • • MC cause of Sub acute endocarditis – Strept Viridans
- 89. streptococci
- 90. • Gram positive • Non spore forming / nonmotile • Capsulated grp A B C& D of beta haemolytic streptococci • Catalase negative (oxidase negative)
- 91. • Arranged in chain • Plane of division along one plane only
- 92. • Longest chain is formed by streptococcus salivarius
- 93. • Ferment sugars • Catalase –ve • Oxidase –ve
- 94. Streptococci • Grow only on enriched media
- 95. Based on haemolysis browns classification
- 96. α hemolysis Partial haemolysis • Viridans • Pneumococcus β hemolysis Complete haemolysis • Further classified by lancefield γ hemolyis Incomplete haemolysis • Enterococcus • Nonenterococcal gp D
- 97. Alpha hemolysis Viridans pneumococci Colony Convex colonies Carrom coin / draughtsman Arrangement Chain Pairs Bile solubility (most specific) -ve Bile soluble Optochin sensitivity Resistant Sensitive Inulin fermentation -ve +ve
- 98. Beta haemolytic • Further classified by lancefield • On the basis of C carbohydrate in cell wall • Into 21 seogroup (no I or J) • Most pathogenic is streptococcus group A (streptococcus pyogenese)
- 99. Based on oxygen requirement Based on haemolytic pattern Without I & J Lancefield classification is based on carbohydrate antigen
- 100. Methods of extraction of C antigen (embedded in cell wall )for lancefeld classification Lancefield HCl Rantz & Randall Autoclaving El kholy Nitrous acid FULLER Formamide Maxted Pronase
- 101. Grp A streptococci • Streptococcus pyogenes Streptococcus pyogenes / grp A streptococci Grp B streptococci / streptococcus agalactiae • Bacitracin sensitive • Ribose fermentation –ve • cAMP test +ve • Hydrolyse Hippurate
- 102. γ haemolytic streptococci Enterococcus Non enterococcal Gp D Bile resistance (ability to grow in 40 % bile) +ve +ve Esculin hydrolyisis + + Growth in 6.5 % NaCl + - PYR test + -
- 108. STREP. PYOGENES (GROUP A) • Capsule made of hyalouronic acid (group A & C ) • d/t hyalouronic acid it is non antigenic • Group B & D made up of polysaccharide
- 109. Biochemical reaction for streptococcus pyogenes • Catalase negative • Bacitracin sensitivity (maxsted observation) • Dosenot ferment +ve • Not soluble 10 % bile (pneumococci is soluble) • Hydrolyse PYR
- 111. • Griffith typing based on M protein in cell wall group B streptococci • 80 subtypes • earlier M type RTI • Latter M type skin & superficial tissue infection
- 112. Culture • Transport media- Pike’s media • Blood agar - Pinpoint colony with wide zone of β Haemolysis • Selective media: • – Crystal violet- Blood agar • More resistant to crystal violet therefore incorporated in selective media • – PNF medium (Polymyxin B Neomycin Fusidic acid)
- 113. • Capsulated strains are pathogenic • Capsulated strains produce mucoid colonies matt type of colonies • While non pathogenic (non capsulated )produce glossy colonies
- 114. • More than 90 % streptococcal infection is caused by Group A beta haemolytic streptococci
- 115. Virulence factors of streptococcus pyogenes • M protein most important virulence factor
- 116. Virulence factors in grp A streptococci Cell wall associated Hyalouronic acid Peptidoglycan & teichoic acid Adhesion protein Fibronectin binding protein M protein Antiphagocytic & anticomplementary Most important virulence factor Filamentous dimeric protein Carboxy terminal of protein is embedded in cell membrane Encoded by EMM gene Griffith classification Secreted virulence Streptodornase /DNAase Streptokinase Streptolysin O & S Dick/scarlatiniform/erythrogenic NADase Serum opacity factor
- 117. Virulence factors secreted Streptolysin O Streptolysin S Oxygen labile Oxygen stable Active only in reduced state (as it is oxygen labile) Active in reduced & oxidised state Hemolysis d/t SLO is seen only in pourplate culture Hemolysis around surface colonies Antigenically similar to 4 hemolysins Antigenic Nonantigenic Also found in grp C & G
- 118. Secreted virulence factors streptokinase • Antigenic ab are produced against it • Used in thrombolysis in MI (from group c streptococcus equisimilis) • Activates plasminogen lysis of fibrin • Spread of infection along with hyalouronidase
- 119. Streptodornase • Depolymerises DNA • Responsible for serous character of exudate • Used to liquefy thick pus collection • 4 serotypes A B C & D • B DNAaseB highly antigenic • Used in retrospective diagnosis of glomerulonephritis
- 120. Secreted pyrogenic toxins • Erythrogenic /dick / scarlatiniform toxins • 3 antigenic types A B & C • A & C phage mediated • B chromosome mediated • Pyrogenic toxin super Ag( like staphylococcal TSST ) streptococcal TSS • Also causes scarlet fever
- 121. • Antigens in cell wall • Group specific cell wall Ag • M protein if present + virulent • Capsule made of hyalouronic acid inhibit phagocytosis • Enzymes & toxins • Pyrogenic • Streptokinase • Hyalouronidase • DNAases • 4 types Type A B C D • Type B is most important • Streptolysin O & S
- 122. Pyrogenic toxin (erythrogenic toxin ) • aka streptococcal pyrogenic exotoxin (SPE) • Cause rash in scarlet fever • Streptococcal toxic shock ds • Dick test • Used to detect susceptibility to scarlet fever • By intradermal injn
- 123. Streptococcal TSST • Caused by pyrogenic exotoxin A
- 124. Staph TSST • By TSST 1 • No bacteremia • Rash + Strept TSST • By pyrogenic exotoxin A • Bacteremia + • No rash
- 125. SPE • 3 Types (SPE A, B and C)- All are e.g. of Superantigens • Type A&C bacteriophage coded, • B toxin chromosomal • Streptococcal SPE type A TSS • Dick test -ID injection produces erythema in susceptible individuals • Schultz Charlton reaction • (blanching of rash after injection of antibodies)- diagnostic for scarlet fever
- 126. Scarlet fever
- 128. Strawberry tongue in scarlet fever
- 129. Capsule made up of hyalouronic acid • Inhibit phagocytosis • Large amount of hyalouronic acid mucoid appearance • Hair like pili (fimbriae ) project through the capsule • Consisting partly of M protein
- 130. M protein • Mediates adherence to epithelial cells, inhibits phagocytosis • Major virulence factor • Inhibiting phagocytosis • Ab to M protein is protective
- 131. Streptodornase • Hydrolyse DNA • Liquifies thick pus • DNAase B used in diagnosis of AGN
- 133. Hemolysis
- 134. Streptolysin O is similar in action to • Clostridum perfringens (perfringolysin O) • Clostridium teatni (tetanolysin) • S pneumoniae (pneumolysin) • Listeriolycin • bacillus cereolsysin • B thuringiensis
- 136. Non suppurative sequelae • Due to antigenic cross reactivity, antibody produced against previous Streptococcal infection antigens, cross reacts with human tissue to produces lesions. This accounts for non-suppurative complications (like acute rheumatic fever and glomerulonephritis).
- 137. (M protein)
- 140. Rx • Penicillin DOC • Erythromycin is the DOC in penicillin allergic patients
- 141. Group B streptococci (streptococcus agalactiae)
- 142. • Most common grp B streptococcus agalactiae • Infection is d/t polysaccharide capsule (in grp B & D) • Hydrolyses Hippurate • Positive CAMP test • Postive CAMP is also seen in listeria monocytogenes • Reverse CAMP is seen in clostridium perfringens
- 144. CAMP test (Christine atkin munch Peterson test )/ arrow head hemolysis CAMP factor is a diffusible substance that completes haemolysis of shep RBC exposed to sphingomyelinase C (staphylococcal toxin)
- 145. • Strptococcus agalactae single most common cause of neonatal meningitis • In adults peurperial sepsis
- 146. Group B streptococci • Orange pigment granadas media / islams media • Selective media kanamycin BA
- 148. Neonatal meningitis • E coli • Klebsiella • Staph aureus • H influenza • Listeria monocytogenes • Streptococcus monocytogenes
- 149. Group C streptococci
- 150. Strep equismilis • Part of normal flora of throat • Ferment trehalose & ribose • Streptococcus pyogenes ferment only trehalose not ribose • Used as source of streptokinase used in thrombolysis • Cause URTI pneumonia endocarditis brain abscess • Rx • Penicillin G
- 151. Non enterococcal streptococci • Streptococcus bovis • Streptococcus equinus • They cannot grow in presence of bile & 6.5 % NaCl • Causes • UTI • Endocarditis in patient with neoplasm of GIT (polyp or carcinoma) • Rx • Penicillin
- 152. Enterococci (previous lancefield group D) • Normal inhabitants of large intestine • Enterococci faecalis • Most commonly isolated • Enterococci faecium • Catalase negative • Non haemolytic • Grow in presence of • 40 % bile • 0.1 % methylene milk • 6.5 % sodium chloride at 45 * c (relative heat resistant) • Relatively resistant to heat surving 60 * c for 30 minutes
- 153. • Arranged in pairs at angle to each other
- 154. Hydrolyses bile esculin agar
- 156. Clinical infection • UTI • Nosocomial UTI MC infection caused by enterococci • Nosocomal bacteremia in patients with intravascular catheters • Bacteremia with out endocarditis is the mc presentation • Abdominal surgical wound infection
- 157. Rx • Synergestic action of aminoglycosides + penicillin • Vancomycin resistance +
- 158. Alpha haemolytic
- 159. Alpha haemolytic Viridans Pneumococcus • S mitis • S mutants • S sanguis • S salivarius forms longest chain /non-pathogenic
- 160. Commonest cause of dental caries S mutans Mc cause of late onset prosthetic valve endocarditis S sanguis Mc cause of infection following human bite S anginosus Other causes (eikenella corrodens & peptostreptococci)
- 161. Virudence group • α haemolysing + greenish discolouration • Normal flora of mouth & URT • Transient bacteremia following tooth extraction or other dental procedure • It can cause endocarditis • Str sanguis • Prophylactic Abx before tooth extraction • Str mutans • Dental caries
- 163. Streptococcus mutans dental caries
- 164. Pneumococci
- 165. • G+ve • Capsulated • Lanceolate appearance
- 166. • Catalase negative • Ferments inulin • Pneumococci are bile soluble
- 167. Bile solubility test • d/t presence of autolytic amidase that cleaves the bond b/w alanine & muramic acid in the peptidoglycan
- 168. Culture • Alpha hemolysis • On further incubation colonies become flat with raised edges & central umbonation • concentric rings • Draughtsman colonies • Carrom coin appearance
- 171. optochin sensitivity • Growth is inhibited around optochin disk
- 173. Quellung reaction • In the presence of homologous anti serum the capsule becomes apparently swollen sharply delineated & refractile
- 175. Virulence factors • Capsule • Most important virulence factor capsular polysaccharide • Non capsulated strains are avirulent • Type 3 pneumococci has abundant capsular material, so more virulent • Ab to capsular polysaccharide protects immunity against infection • Penumolysin : • Membrane damaging toxin which has cytotoxic • Destroys respiratory ciliary cells & PMNs • complement activating properties (classical pathway by C1q ) • Autolysin
- 176. Increased risk • Increased risk after splenectomy • Old age is an independent risk factor for pneumococcal infection
- 177. Infections • Commonest infection by pneumococcal infns otitis & sinusitis • Most common cause of pneumonia • Meningitis is the most serious infection • Empyema is the most common complication • Source of infection • Carriers in URT • Less often from patients
- 178. Pneumococcal vaccine • Polyvalent 23 polysaccharide vaccine • Made of capsular polysachharide of 23 serotypes • Not recommended for children under 2 years • indications • With absent or dysfunctional spleen • c/c liver or renal or lung or cardiac ds • CSF leak • Immunodeficiency • Conjugate vaccine (coupled to a protein • 7 most common capsular polyscharides • Can be given to children under 2 years of age
- 179. Rx • Beta lactam antibiotics DOC • DOC for ASOM amoxicillin • DOC for meningitis s vancomycin + ceftriaxone