Micro: Strep Test

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Micro: Strep Test
2010-08-28 21:14:51
Strep Test

MIcro Strep Test
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  1. Gram positive cocci; Catalase negative; Streptococcus, Enterococcus, and strep-like bacteria general characteristics
    • Gram + cocci in pairs or chains
    • Catalase =
    • facultative anaerobes
    • Gray colored colonies
    • Enhanced growth in CO2 or anaerobic conditions
  2. Natural habitats
    • Skin
    • Oral cavity
    • GI tract
    • upper respiratory tract
    • genital urinary tract
    • female genital tract
  3. Strep are Catalase _____?
    Strep are catalase =
  4. Hemolytic reactions: Brown Classification
    Based on an organism's ability to lyse sheep RBCs

    Subsequent testing is based on interpretation of hemolytic reaction
  5. 3 types of hemolysis
    Alpha: Partial lysis of RBCs surrounding a colony causing a greenish gray coloration

    Beta: Complete lysis of RBCs causing a clearing from under and around a colony

    Gamma: no change in RBCs
    Disk diffusion test on sheeps blood and incubated in CO2

    Strep pneumo is sensi to optochin and should have a zone size of up to 14

    all other strep are resistant
  7. Bile solubility
    Certain orgs. lyse in the presence of bile

    Heavy suspension of orgs. is added to sodium deoxycholate solution and incubated at 35C for 1 hour. Clearing of turbidity indicates bile solubility (organism lysis) and positive rxn

    Strep pneumo is bile solubility +

    other alpha-streps are = (cloudy)
  8. Latex agglutination
    white latex particles coated in Abs to select Strep pneumo capsular pollysach. Ags

    Mixing with strep pneumo causes an Ag/Ab rxn resulting in visible clumping of the latex reagent

    Neg. pneumo slide tests should be confirmed by optochin/bile solubility, as not all strep pneumo isolates possess detectable Ag
  9. Lancefield Classification and testing
    Based on detection of group specific carb. Ags attached to the cell wall

    only beta-strep, group D strep, and Enterococcus contain detectable group Ags. All other streps are non-groupable, but may cross-react

    STREPTEX: enz. extraction then latex agglutination. can detect lancefield groups A,B,C,D,F,G, which are the most common human isolates
    • Disk Diffusion
    • any zone of inhibition is considered +
    • beta-strep Group A is sensi. (+)
    • all other beta-strep supposed to be resistant (=)

    10% BetaSC, G and 5% betaSB are +. perform bactitracin in conjunction with SXT, CAMP test to help eliminate false positives.
  11. SXT
    • Disk Diffusion
    • Any zone of inhib. +
    • beta strep group A and B are resistant =
    • other Beta strep are sensi +
  12. CAMP test
    developed by Christie, Atkins, Munch-peterson

    plate streak of Beta-strep and a single streak of hemolytic Staph Aereus perp. to each other about 3-4mm apart

    Beta-strep group B produce CAMP factor which is a protein that interacts with the Beta-hemolysin produced by SA to cause a synergistic hemolytic rxn
  13. Gamma Strep Identification
    • Bile Esculin Agar: 40% bile and esculin
    • no growth: gamma-srtrep
    • growth: gamma-strep also
    • growth w/esculin hydrolysis: agar turns black. Group P-D strep and Enterococcus

    • PYR: presence of pyrrolidonyl arylamidase
    • Org. added to paper disk impregnated w/ L-pyrrolidonyl-beta-naphthylamide and incubated
    • if PYR enz. is present, substrate is hydrolyzed and a color change occurs with the addition of dimethylamino-cinnamaldehyde

    =: no color change. stays white. Group D strep will do this

    +: Pink red color. Enterococcus

    • Biochemical Profiles:
    • Commercial kits using carb fermentation, chromogenic enz. substrate tests, and other standard ID tests
    • can ID most strep species
    • definitive ID reserved for clinically sig. isolates and by Dr.'s request
  14. Alpha Strep: Streptococcus pneumoniae: Clinical sig. and virulence factors
    • 5-10% adults colonized
    • most commonly infect young, old, immunocomprimised
    • cause ear inf., sinus inf., meningitis, endocarditis, sepsis, and pneumonia

    *becoming increasingly resistant to penicillin, cephalosporins, and fluoroquinolones due to the development of penecillin binding proteins (PBP's)

    Virulence factor is pollysacch. capsule which make it resistant to phagocytosis and lysis by WBCs
  15. Strep pneumoniae ID:
    Gram: + cocci in paris or chains

    Catalase: =

    Colony morphology: Alpha hemolytic, can sometimes be mucoid, some times looks loke a checker head colony

    Optochin: sensi (zone clearing)

    bile solubility: +

    Pneumoslide/latex: +
  16. Beta Strep: Streptococcus Pyogenes (BetaSA): Clinical sig.
    • humans are the only host
    • cause pherengitis/strep throat, ear inf., tonsilar abscesses, sinus inf., sepsis, meningitis, skin and soft tissue inf.

    • Acute Rheumatic fever (ARF): Associated with beta strep group A pherengitis and will show up 1-5wks after strep throat
    • multi syst. disease, cardiac abnormalities, arthritis, rash, neurological. Abs body makes to diff strep Ags cross react and attack host tissue, causing disease.

    • Acuute glomerulonephritis: Associated with Beta strep group A pherengitis or soft tissue inf.
    • 10 days following pherengitis, or 3-6wk long skin inf.
    • inflammatory disease of renal glomerulus causing disease of renal glomerulus and renal failure from deposition of immune complexes containing strep AGs/host Abs in glomerular tissue

    • Virulence factors:
    • M-protein: a self surface Ag that helps resist phago and lysis by body's WBCs

    hyaluronidase and streptokinase: aid in break-down and infection of tissue
  17. Beta strep: streptococcus pyrogens (betaSA) toxin production
    • Hemolysins:
    • lethal to cells, especially RBCs and WBCs
    • 2 types:
    • Streptolysin O (SLO): O2 labile. Measurement in serum useful for retrospective diagnosis of recent pharyngeal inf. and may aid in diagnosis of ARF
    • Streptolysin S (SLS): O2 stabile.

    • Streptococcal pyrogenic exotoxins: Cause 2 diseases:
    • Scarlet fever: Strep throat, possible vomiting, bright red rash on face, groin, and underarms that leaves skin dry and pealing.
    • Streptococcal toxic shock-like syndrome: Associated w/ invasive Beta strep group A soft tissue inf., sepsis, and pealing.
  18. BetaSA ID
    • gram: +
    • Catalase: =
    • colony morph: gray colonies w/ large zone of Beta hemolysis
    • Streptex: group A +
    • Bacitracin: sensi +
    • SXT: resistent =
  19. Streptococcus Agalactiae (betaSB): Clinical sig. and virulence factors
    Adults: UTIs, pneumonia, skin and soft tissue inf., endocarditis, and sepsis

    Neonates: sepsis, meningitis, and pneumonia. can infect during birth or in utero. 10-35% of women are colonized. women screened 35-37wks to ident. as carriers. treated w/ antibiotics to reduce chance of infecting the infant.

    • virulence:
    • polysacch. capsule: prevents phago and lysis by WBCs
    • beta-hemolysin: mainly in respiratory inf. lysis pulmonary alveolar cells.
  20. betaSB ID
    • Gram: + cocci in pairs or chains
    • catalase: =
    • Colony morph: gray colonies w/small zone Beta-hemolysis
    • Streptex: group B +
    • bacitracin: resistent =
    • SXT: resistent =
    • CAMP test: + Arrowhead zone
  21. Gamma strep: Group D Strep: Clinical sig.
    causes meningitis, sepsis, endocarditis, UTIs, abscesses,

    • Streptococcus bovis: linked to colon cancer.
    • 56% colon cancer pts. + for it, vs. 10% normal carriage rate.
    • pts. +, particularly in blood, should be screened for accult bowel and liver disease/cancer.
  22. Group D strep ID
    • gram: + cocci in pairs or chains
    • Catalase: =
    • Colony morph: gray colored gamma-hemolytic (non-hemolytic)
    • streptex: group D +
    • bile esculin: hydrolyzer (black)
    • PYR: =
    • 6.5% NaCl/ Salt tolerance: =

    API only way to fully ID species level. reserved for clinically sig. isolates
  23. Enterococcus clinical sig. and virulence factors
    causes UTIs, abdominal and pelvic infections, wound and soft tissue inf., endocarditis, and sepsis

    • 2 most common species:
    • E. faecalis
    • E. Faecium

    Becoming increasingly resistant to antibiotics ampicillin and vancomycin after 1980's. VRE linked to nonsocomial outbreaks/inf. control problems. Certain strains VRE able to pass resistance genes to other Enterococci, gram + orgs., including SA

    • Virulence factors
    • virulence not well understood
    • Extracellular surface protein (ESP): surface protein unique to Enterococcus that allows for resistance to Ab attack
    • Also, intrinsic low-level resistance to aminoglycosides, penicillins, cepalosporins; naturally more resistant to antibiotics.
  24. Enterococcus ID
    • Gram: + cocci pairs and chains
    • Catalase: =
    • Colony Morph.: gray gamma/non-hemolytic colonies
    • Streptex: 90% are Lancefield group D +
    • Bile esculin: hydrolyzer, turn agar black
    • PYR: +
    • 6.5% NaCl/salt tolerance: + growth
    • API: only way to fully ID to species level.
  25. Viridans Strep: Clinical Sig.
    includes several species of alpha and gamma strep

    normally opportunistic, like immunocomp., heart disease, or poor hygiene

    cause endocarditis, sepsis, oral abscesses, deep tissue abscesses

    • Commonly isolated species:
    • S. mutans: Cavities, endocarditis
    • S. anginosus: oral abscesses, deep tissue abscesses, cavities, endocarditis, sepsis. produces diacetyl as a metabolic end product. gives a sweet caramel/ butterscotch odor.
    • S. mitis: sepsis, meningitis
  26. Viridans ID
    • Gram: + cocci in pairs and chains
    • Catalase: =
    • Colony Morph: gray alpha or gamma, sometimes beat hemolytic
    • Streptex: may or may not have Lancefield group Ag
    • Optochin: = resistant
    • Bile solubility: =
    • Bile esculin: variable rxn, growth or no growth
    • PYR: =
    • 6.5% NaCl/salt tolerance: = no growth
    • API only way to fully ID
  27. Streptococcus like bacteria: Clinical sig.
    Catalase =, gram + cocci. genetically unrelated.

    opportunistic pathogens. Attack immunocomp. cause sepsis, endocarditis, eye inf., abscesses

    • Species include:
    • Leuconostoc
    • Pediococcus
    • lactococcus
    • Aerococcus
    • Vagococcus
    • Gemella
    • Abiotrophia: previously known as nutritionally variant strep, or satalighting strep. contains pyridoxal/vit. B6/thiol. think Abiotrophia if blood culture bottles/ thioglycolate broth are positive, but all other media are subculture plates are =. presumptive ID with Staph streak-innoculate sheep's blood w/ blood culture bottle and make 1 streak of SA. leads to a haze of Abiotrophia after 1 night incubation.
  28. Strep-like bacteria ID
    • gram: + cocci in pairs, tetrads, or chains.
    • Catalase: =
    • API: best for ID/ also see flow chart