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103 Cards in this Set
- Front
- Back
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What is the catalase test used for?
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to differentiate between aerobic and anaerobic bacteria
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What reagent does the catalase test involve?
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Peroxidase is added to a colony of bacteria.
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What is catalase?
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Catalase is an enzyme that breaks down hydrogen peroxide in the cell
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What does a positive catalase result look like? What does a negative result look like?
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positive: vigorous bubbling will be seen when peroxidase is added to a bacterial colony
Negative: either no bubbling or only slight bubbling will be seen when adding peroxidase to bacteria |
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What types of bacteria are catalase positive? Why?
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aerobes, falcultative anaerobes, and microaerophiles.
They all oxidize carbohydrates and produce Hydrogen peroxide |
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Is Staphylococcus catalase positive or negative?
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Positive
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Is Streptococcus catalase positive or negative?
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Negative
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Is Bacillus catalase positive or negative?
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positive
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Is Clostridium catalase positive or negative?
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negative
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Is Listeria catalase positive or negative?
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positive
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Why should you be careful when performing a catalase test on a colony from a blood agar plate?
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RBCs contain catalase and can give a false positive test.
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What is the purpose of the A disk tests?
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it differentiates between group A Streptoccoci and Nongroup A Streptococci.
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What is th basis of the A disk test?
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Sensitivity of streptococci to the antibiotic, Bacitracin
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Why is the A disk test only used on beta - hemolytic Streptococci?
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alpha hemolytic Strep. are sensitive to Bacitracin
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How does the A disk test work
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Bacteria is streaked on an agar plate and a paper disk impregnated with Bacitracin is placed on top.
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What does a positive A test look like? What does this mean?
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any inhibition of growth of the bacteria
bacteria is sensitive to Bacitracin |
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What does a negative A test look like? What does this mean?
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Growth of bacteria up to the disk
Bacteria is not sensitive to Bacitracin |
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What physically distinguishes alpha hemolytic bacteria from beta hemolytic bacteria (how would it look on the agar plate)?
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Alpha hemolytic bacteria - appears to have a greensh tint on agar plate
Beta hemolytic bacteria - appears clear on the agar plate |
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What does the citrate test determine?
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Bacteria's ability to utilize citrate as a sole energy source
citrate --> acetate --> OAA (oxaloacetate) --> CO2 and Pyruvate |
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What are two types of bacteria that are citrate positive?
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Enterobacteria aerogenes and Citrobacter sp.
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How does the citrate test work?
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the surface of a green Simmons citrate slant is streaked with a loop of bacteria and incubated for 24 - 48 hrs.
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What does a positive citrate test involve?
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Bacteria grows on the slant and an alkaline reaction occurs to turn the originally green indicator dark blue.
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What does a negative citrate test look like?
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No growth will appear on the slant, and no color change will occur
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What is the coagulase test used for?
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differentiates between Staph aureus (+) and other strains of staphylococci (-)
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What does the coagulase test test for?
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production of extracellular coagulase enzyme in bacteria
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How does coagulase test work?
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A loopful of Bacteria is added to a drop of plasma to look for clotting.
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What happens in a positive coagulase test?
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Bacteria will clump on the slide and will not be able to be broken up
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What does a negative coagulase test look like?
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Bacteria will form a milky homogeneous emulsification on the slide
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Is Staph aureus coagulase positive or negative?
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positive
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Is Staph epidermidis coagulase positive or negative?
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negative
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What is the medical importance of the coagulase test?
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~95% correlation between coagulase production and pathogenicity, therefore the method is considered a test for S. aureus pathogenicity.
[Note: not definitive b/c some coagulase neg. strains are pathogenic] |
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What does MIO stand for?
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Motility Indole Ornithine Decarboxylase
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What family of bacteria does the MIO test characterize?
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Enterobacteriaeae
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How is the MIO test performed? (basic procedure; ie loop/needle or streak/stab)
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A needle laced with bacteria is stabbed straight down into the medium
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What does the Motility test do?
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Tests whether the bacteria are able to move freely in the medium or if they are relatively immobile
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What does a positive motility test look like?
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Bacteria migrate away from the stabline and into the medium; tube appears cloudy
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What does a negative mobility test look like?
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Bacteria stay on the stab line
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What is indole?
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breakdown product of tryptophane in Enterobacteiaceae
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What reagent is used to test for Indole in the MIO tube?
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Kovac's reagent
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What does a positive Indole test look like?
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when Kovac's reagent is dropped on the surface of the MIO tube, a red color should immediately appear
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What does a negative Indole test look like?
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No reaction with the MIO medium will occur; drop of Kovac's reagent will remain its original color (yellow)
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What does the ODC test test for?
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Bacteria's ability to decarboxylate orthinine.
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What does the yellow color in the MIO tube indicate?
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glucose fermentation
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What does a positive ODC test look like?
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The tube reverts from yellow to purple at the bottom of the tube
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What does it mean when the purple color advances from the top of the tube?
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The purple comes from the oxidative metabolism of peptone and should be ignored.
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The oxidase test identifies bacteria that produce what enzyme?
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cytochrome oxidase
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What family of bacteria are known as oxidase positive? (2 organisms)
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Psuedomonas and Neisseria
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Most gram positive bacteria are oxidase positive or negative?
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negative, but the test is not generally useful in their idenification
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How is the oxidase test performed?
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A piece of filter paper is wet with the oxidase enzyme and a colony of bacteria is rubbed onto the paper to look for a color change
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What does a positive oxidase test look like?
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the bacteria rubbed onto the filter paper will turn purple after 15sec
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What does a negative oxidase test look like?
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There will be no color change on the filter paper
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Is a color change that occurs after one minute in the oxidase test consider positive or negative?
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it is a false positive and is considered to be negative
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What is the purpose of the P test?
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to differentiate S. pneumoniae from other alpha-hemolytic streptococci based on sensitivity to the drug optochin
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Sensitivity to what agent is the basis of the P test?
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optochin
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What group of streptococci are most sensitive to the agent in the P test?
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S. pneumoniae
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How are the P test results interpreted differently from other identification tests?
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results are interpreted as the bacteria either being sensitive or resistant to the agent rather than having positive and negative results.
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If the zone of inhibition of the bacteria in the P test is greater than 15mm, the test is consider to be what? What organism is it?
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sensitive;
S. pneumoniae |
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If there is no zone of inhibition in the P test, the bacteria are considered to be what?
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resistant
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The P test should not be performed on an organism that is catalase positive or negative?
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positive
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What does TSI stand for?
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Triple Sugar Iron
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What organism is TSI used to identify?
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gram negative bacilli, especially members of th Enterobacteriaceae
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The TSI test is based on an organism's ability to do what?
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ferment glucose, lactose, or sucrose; produce H2S
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How is the TSI test performed?
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the medium is inoculated by stabbing a wire needle, laced with bacteria, to the bottom of the medium and streaking it across the slanted surface
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The TSI medium has both acidic and basic indicators on the ends (bottom and slant) which is which?
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slant: basic (alkaline) indicator
bottom: acidic indicator |
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What does it mean if the bottom of a TSI tube turns yellow and the slant turns red? Is the organism aerobicc, anaerobic, or falcultative
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glucose fermentation only, no lactose or sucrose is fermented;
the organism is falcultative |
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What does it mean when both the bottom and top of the TSI tube turn yellow?
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the bacteria is a lactose and/or sucrose fermenter
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What does it mean if the both the bottom and the slant of the tube turn red?
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no carbohydrate fermentation
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Observance of a blackened bottom (butt) of a TSI tube indicates the production of what?
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H2S
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What does the urease test test for?
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an organism's ability to hydrolyze urea into ammonia, CO2 and water through the urease enzyme
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How is the urea test performed?
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The surface of the medium is streaked with a wire loop, laced with bacteria
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What does a positive urease test look like?
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color change from pink to red that may be on the slant or extend into the entire but.
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What does a negative urea test look like?
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no color change in the agar
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What compound is contained in MacConkey agar?
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lactose
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How do lac+ bacteria look on the MacConkey agar plate?
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the colonies will appear red
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How do lac- bacteria look on the MacConkey agar plate?
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the colonies will appear white or colorless
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Why does a surgical scrub show more bac than no handwashing at all
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surgical scrubbing mechanically removes microorganisms that may have been picked up from the environment, but also exposes other organisms that were previously under layers of skin.
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What would happen if a gram stain is overdecolorized?
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gram pos will look pink and the test will be hard to interpret
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What type of bac does the MacConkey agar plate grow?
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gram neg
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What color is the MIO tube before color change?
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purple
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Wht color is the citrate tube before color change?
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green
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What color is the TSI tube before color change?
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orange
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What color is the urea tube before color change?
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peach
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A/A
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yellow slant/butt;
ferments glucose and lactose/sucrose |
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K/A
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red slant/yellow butt;
ferments glucose and doesn't ferment sucrose/lactose |
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K/A plus H2S production
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red slant/dark butt;
ferments glucose and doesn't ferment sucrose/lactose |
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K/K
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red slant/butt;
doesn't ferment glucose or lactose/sucrose |
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Why do you incubate agar plates upside down?
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to keep the agar from drying out and to keep moisture from the condensation on the cap from dropping onto the bottom of the plate
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What is the purpose of the ELISA assay?
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to determine if a person was exposed to a particular antigen by testing the levels of antibody to that antigen.
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What are the steps involved in an ELISA assay?
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-a microtiter plate is covered in antigen and rinsed
-a primary Ab is diluted four-fold and each dilution is added to a well in the plate -the plate is rinsed to remove any unbound Ab -a secondary Ab is added to the plate and binds to the primary Ab -a substrate that acts with the secondary Ab is added to th plate -the wells are observed for a color change in order to determine the exposure of the patient to the antigen |
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What is colony morphology?
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what the cells look like on the plate (large, small, smooth, rough, mucoid, etc)
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What is cell morphology?
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What the cells look like under the microscope (bacillus, coccus, etc)
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What is the purpose of the phage plates?
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to quantify the amount of phage in a sample.
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What are the steps of making phage plates?
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-grow bac as a law on an agar plate
-introduce phage -repeat until plaque is formed |
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Does alcohol sanitizer work on all bacteria?
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No; alcohol sanitizer only acts against bacteria with lipid envelopes
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What is the purpose of phase contrast microscopy?
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to observe the morphology and motility of live unstained bacteria.
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What are the steps of gram staining?
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1 - prepare a smear from broth, isolated colonies, or a clinical specimen
2 - heat fix the slide and dip in crystal violet dye for 1min. Rinse 3 - add a iodine to the slide to make it more difficult to decolorize. Rinse 4 - decolorize smear with 95% alcohol 5 - add counterstain to the smear. 6 - observe under microscope |
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What is the chemical name of the reagent that is used to react with the stain and make it more difficult to decolorize? What was the specific chemical used in the experiment?
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mordant;
iodine |
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What color do gram negative bacteria stain? Why (presumably)?
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Reddish pink; because the thin lipid containing cell wall is dissolved by the dye causing leakage of macromolecules and subsequent leakage of the crystal violet complex formed in the cell after staining
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What color do gram positive bacteria stain? Why (presumably)?
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Dark purple; the thick peptidoglycan cell wall contains little to no lipid and therefore the crystal violet complex doesn't leak from the cell
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What is the name of the red dye used in gram staining?
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Safranin
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What is the purpose of the bile esculin test?
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characterizes group O streptococci.
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What does a positive bile esculin test look like?
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medium turns brown
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What does a negative bile esculin test look like?
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medium turns red
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