Lab Tests for Streptococcaceae (Week 1) – Flashcards
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| Streptococcaceae are catalse |
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| Negative |
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| Micrococcaceae are catalse |
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| Positive |
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| Streptococcaceae are gram what? What shape? |
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| Gram-pos cocci in chains |
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| Micrococcaceae are gram what? Shape? |
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| Gram-pos cocci in clusters. |
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| Bacitracin susceptibility (Taxo A) for Streptococcaceae |
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| Any zone of inhibition is sensitive. Helpful to identify group A Beta-hemolytic Strep. |
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| Latex agglutination test for Streptococcaceae |
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| Used to identify group A Beta-hemolytic Strep. Based on the group-specific carbohydrate antigen from the cell wall. Place colonies on reaction card and rock and roll. Pos=agglutination |
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| PYR test (Pyrrolidonlyarylamidase test) |
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| Presumptive test for both Group A Strep and Enterococcus spp. Reagent A to paper disk and put colonies onto the disk. Incubate and then add reagent B. Pink= pos |
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| CAMP test |
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| Group B strep produce CAMP (Christie, Atkins, Munch-Peterson) factor that acts synergistically with the Beta toxin produced by STA to produce more potent hemolysis. |
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| Method of CAMP test |
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| Innoculate BAP with a center streak of STA. Inoculate the unknown organism and controls perpendicular to the line of STA. Observe for arrowhead hemolysis = pos |
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| Hippurate hydrolysis |
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| Group B Strep are able to hydrolyze hippuric acid to its components, glycine and benzoic acid. |
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| Method of hippurate hydrolysis |
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| Inoculate broth containing sodium hippurate with organism and incubate overnight at 35 degrees Celsius. 2 methods. Method A detects presence of benzoic acid Precipitate of ferric chloride last for more than 10 mins= pos. Method B detects presence of free glycine. Ninhydrin added. Deep blue/purple color = Pos |
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| Bile esculine test |
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| Used for the identification of Enterococcus spp. and Group D Strep. Inoculate bile esculin with organism and incubate. Black color = Pos |
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| Salt tolerance test |
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| Helps differentiate Enterococcus spp. from Group D Inoculate organism into a TSB containing 6.5% NaCl. Incubate and observe for growth. Growth= Pos. |
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| Which spp will be positive for the Salt tolerance test Enterococcus spp. or Group D Strep? |
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| Enterococcus spp. |
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| Susceptibility to optochin (Taxo P disc) |
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| Based on the fact that S. pneumoniae is senstive to optochin (Ethyl hydrocupreine hypochloride) |
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| Method for optochin susceptibility test |
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| Streak BAP for isolation with organism. Place P disc in between the first and second quadrants. Incubate in Co2 and measure zone size. Sensitive >/= 14mm. Resistant is less than 14mm |
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| Strep. pneumoniae prefers what growth environment |
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| Co2 |
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| Bile solubility test |
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| Bile salts dissolve colonies of S. pneumoniae (autolysins) Reagent = 10% sodium. |
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| Bile solubility test method |
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| Place on drop of bile reagent on an isolated colony.Incubate media side down Pos= colony disappears. Neg= colony remains intact. |
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| Which organism is soluble in the bile solubility test? |
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| Strep. pneumoniae, which is sensitive to the P-disk. |
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| Quellung Test |
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| Serologic test for the detection of s. pneumoniae. No longer in use. |
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| Pneumoslide Test |
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| Latex test for the detection of s. pneumoniae. Used on either isolated colonies or organisms grown in broth |
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| Catalse test used to differ between? |
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| Used to differentiate between micrococcaceae and streptococcacea families. Micrococacceae are pos Streptococacceae are neg |
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| Catalase test explanation |
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| In the presence of the enzyme catalse, hydrogen peroxide is broken down into water and oxygen gas. Place h2O2 on a slide and emulsify a colony in it |
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| What can cause false positives in the catalse test? |
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| Wire loops and blood cells from BAP Overreading the results |
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| Glucose fermentation is used to differentiate between what? |
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| Staphylococcus spp. and Micrococcus spp. of the Micrococacceae family |
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| Explanation of glucose fermentation |
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| Organsisms can utilize glucose by three mechanisms: oxidation, fermentation or notat all (nonsaccharolytic) |
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| Media for glucose fermentation |
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| OF media which is a semi-solid media cotaining a low concentration of peptones and a high conc of glucose |
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| Method of glucose fermentation |
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| Inoculate 2 tubes of media with organism. Cover one with oil overlay. Indicator is bomthymol blue. Acid results( organisms that fermented the glucose)= yellow. Alkaline results= blue/green color |
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| Fermenters |
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| Use the glucose in both tubes, producing a yellow color |
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| Oxidizers |
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| Only use the glucose i the tube open to air which will be yellow. Tube with oil will remain blue (due to the absence of oxygen) |
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| Nonsaccharolytic organisms |
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| Both tubes will be blue-green because the glucose was not used |
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| Coagulase Test |
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| Fibrinogen(in serum) in the presence of coagulase will agglutinate to form a fibrin clot. Reagent used is EDTA rabbit plasma. Staph aureus is coag pos. Other staph are coag neg. |
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| Coagulase slide test procedure |
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| Test for bound coagulase. 1 drop of sterile water in two circles drawn on a glass slide. Place on colony into each of the circles. Add a drop of EDTA plasma to one and mix. |
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| Coagulase tube test |
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| Test for free coagulase. Drop pure colony into 0.5ml rabbit plasma and incubate. After 4 hours if still neg, take out and incubate at room temp for 18 hours. |
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| Staphaurex latex test |
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| Latex particles coated with IgG bind to protein A and clumping factor produced by Staph. aureus. Visible clumping with clear backgorund= STA. No clumping with milky background= other staph |
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| Staphaurex latex test procedure |
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| Add one drop reagent to circle on card. Use applicator stick to put a few colonies in reagent. Rotate card for 2 mins. (Nonspecific end binds to STA) |
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| DNase test |
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| STA produces DNase which hydrolyzes nucleic acids. Positive= break down of DNA in media, a pink halo forms around well (STA) Negative= Media remains blue (STN) |
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| DNase Test procedure |
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| Hydrolysis of nucleic acids via STA is detected by placing a well into media and filling the well with a heavy inoculum. Incubate and read results |
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| Novobiocin Susceptibility (Diagnostic Test) |
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| Only 3 Staph spp. are novobiocin resistant. Only Staph. saprophyticus is clinically significant. Reagent used in the test are Novobiocin disk, BAP and 0.5 McFarland standard |
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| Novobiocin Susceptibility method |
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| Make a 0.5 McFarland suspension of organisms (1.0-1.5 x 10^8 organisms/ml suspension) Streak BAP, drop disc in center of plate, incubate, read results |
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| Results of a Novobiocin Susceptibility test |
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| Zone of < or = to 12mm is resistant to novobiocin (S. saprophyticus) Zone of > or = to 16mm is susceptible or sensitive to novobiocin (Other staph) |
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| Mannitol Salt Agar |
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| High concentration of salt selects for halophiles like Staphylococcus spp. STA can ferment the mannitol as well, while other staph cannot |
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| Reagents involved in mannitol salt agar |
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| Peptones, beef extract 7.5% Nacl Mannitol (Fermentable carb) Phenol red indicator |
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| Mannitol salt agar procedure and results |
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| Inoculate mannitol salt agar with organism. If the media turns yellow under the colonies it is acidic and pos (indicative of STA). If the media remains pink/red it is alkaline (indicative of STN) |
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| Differentiation of micrococcus spp and staphylococcus spp. |
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| Glucose utilization Bacitracin susceptibility (Taxo A disc) Furazolidone suscpetibility |
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| Bacitracin susceptibility (Taxo A disc) test procedure |
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| Prepare a 0.5 McFarland suspension of organism. Streak a lawn on BAP Drop disc in center of plate incubate. Read results |
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| Bacitracin susceptibility (Taxo A disc) test |
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| Used to differentiate between micrococcus spp. and staphylococcus spp. Zone of > or = to 10mm is sensitive or susceptible to bacitracin (Micrococcus spp.) Zone < or = to 10mm is bacitracin resistant (Staphylococcus spp.) |
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| Furazolidone susceptibility test procedure |
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| Use same procedure as Bacitracin except use a 100ug disc of furazolidone |
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| Furazolidone susceptibility test |
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| Used to distinguish Micrococcus spp. from Staphylococcus spp. No zone (<6mm) resistant to furazolidone (Micrococcus spp.) Zone > or = 15mm is susceptible or sensitive to furazolidone (Staphylococcus spp.) |
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| Staphylococcus aureus causes |
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| Petigo Food poisoning Scalded skin syndrome Carbuncles |
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| Streptococcus pyogenes causes |
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| Acute pharyngitis (strep throat) Scarlett Fever Skin infections |
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| Staphylococcus saprophyticus |
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| Clinically significant as an etiologic agent of urinary tract infections in women of child bearing age |
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| Streptococcus pyogenes (Group A beta hemolytic strep) |
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| Pharyngitis Yellow/white exudate on tonsils Impetigo |
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| Streptococcus agalactiae (Group B strep) |
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| Normal flora of the genitourinary tract Cause of neonatal sepsis and meningitis Wound and burn infections |
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| Group D non-Enterococcus |
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| Normal flora of the GIT Pathogen in UTI's, abdominal and wound infections |
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| Streptococcus pneumoniae |
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| Pneumonia Meningitis Otitis media |
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| Viridans streptococci |
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| NF or oropharynx, nasopharynx and skin Subacute bacterial endicarditis Seen in patients with underlying heart problems |
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| Enterococcus spp. |
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| NF of the GIT Cause nosocomial UTI's Abdominal wounds Septicemia |
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| Streptococcus milleri group |
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| NF of the oropharynx and nasopharynx, GIT and vagina Cause SBE, pulmonary head and neck infections, oral and abdominal abscesses |
