MIP 302 Final

Catalase test

determines if the bacterium produces the enzyme catalase, aerobic

catalase

enzyme that breaks down hydrogen peroxide in cells into water and oxygen gasses

hydrogen peroxide

catalase test; reagants added to sample

positive

catalase test; bubbles observed

negative

catalase test; no bubbles observed

Oxidase test

determines if a bacterium produces cytochrome oxidase, aerobic

cytochrome oxidase

enzyme that transfers electrons between cytochrome c and oxygen, the final electron acceptor of the ETC

redox indicator disks

oxidase tests; indicator for test

positive (oxidized)

oxidase test; dark blue to maroon color

negative (reduced)

oxidase test; colorless

PR carbohydrate broth

fermentation; differential media; broth that contains a sugar as the sole carbohydrate source. Purpose is to see if the organism ferments that sugar to produce acid. Also contains peptone and beef extract that allows bacteria to grow even if they don't ferment the sugar (not selective)

Also has durham tube to catch gas bacteria produces

phenol red

pH indicator that is yellow in acidic conditions and red in basic conditions

positive

PR carbohydrate; yellow

negative

PR carbohydrate; red/orange

Nitrate broth

anaerobic; differential; distinguishes bacteria's ability to produce nitrate and nitrite reductase

Nitrate/nitrite reductase

enzyme that reduces nitrate/nitrite respectively during anaerobic respiration

48 hours

Nitrate broth; time of incubation

Nitrate A and B, zinc

Nitrate broth; reagants added after incubation

+/-

Nitrate broth; red upon addition of Nitrate A and B (write answer it charge/charge format)

inconclusive, add zinc

Nitrate broth; colorless upon addition of Nitrate A and B

-/-

Nitrate broth; red upon addition of nitrate A, B, and zinc

+/+

Nitrate broth; colorless upon addition of nitrate A, B, and zinc

MacConkey agar

selective for growth of gram negative bacilli, bile salts and crystal violet; differential for ability to ferment lactose, lactose sugar, peptones and beef extract.

Neutral red

pH indicator, red in acidic environment and yellow in basic environment

positive for lactose fermentation

MacConkey; red colonies on pink plate with precipitated bile salts

negative for lactose fermentation

MacConkey; transparent colonies on orange-ish plate

Gram positive organism

MacConkey; no growth

PEA agar

selective medium that inhibits growth of gram negative organisms, phenylethyl alcohol

Blood agar

used for cultivation of medically significant bacteria; differential for lysis of blood cells

alpha hemolysis

incomplete lysis of blood cells

beta hemolysis

complete lysis of blood cells

gamma hemolysis

inability to lyse blood cells

Pseudosel Agar

selective for Pseudomonas aeruginosa. Differential for pyocyanin, MgCl and KSO4 stimulate growth

pyocyanin

blue-green pigment that is water soluble, can be seen under normal room lighting, associated with Pseudomonas aeruginosa

Pseudomonas Agar F

differential for ability to produce flourescin, magnesium and potassium salts enhance production. Need UV light to visualize. Differentiates members of the genus Pseudomonas

fluorescin

lime-green pigment visualized under UV light, associated with Pseudomonas

Indole test

differential for bacteria that produce tryptophanase, can utilize tryptophan as a carbon source, tryptone broth

Tryptophanase

enzyme that breaks down tryptophan into indole, pyruvic acid and ammonia

48 hours

Indole test; incubation time

Kovac's reagant

Indole test; reagant added after incubation

positive

Indole; red ring at surface of broth

negative

Indole; colorless

MRVP test

differential, tests for glucose fermentation by two pathways, production of mixed acids or acetoin. Contains glucose and peptone

48 hours

MRVP; incubation time

methyl red

pH indicator; red in acidic conditions and yellow in basic conditions

Methyl red

Methyl red test; reagant added after incubation

Barritt's A then Barritt's B

Voges-Proskaeur test; reagant added after incubation and transfer to new tube

positive

methyl red test; red; VP opposite result in glucose fermenters

negative

methyl red test; yellow; VP opposite result in glucose fermenters

positive

VP test; red layer in upper portion of broth; MR opposite result in glucose fermenters

negative

VP test; no color change or green/copper color; MR opposite result in glucose fermenters

Citrate test

selective for use of citrate as a carbon source and ammonia and nitrogen souce and differential for production of citrate permeases; simmon's citrate agar

bromthymol blue

pH indicator; yellow in acidic conditions, green in neutral conditions, blue in basic conditions

48 hours

citrate test; incubation time

positive

citrate test; growth on blue slant

negative

citrate test; no growth, green slant

Urease test

differential for production of urease, urea agar, contains urea and glucose, contains phenol red

urease

enzyme that hydrolyzes urea to form ammonia and CO2.

positive

urease test; hot pink slant

negative

urease test; no color change or yellow

LDC test

differential for production of lysine decarboxylase, contains lysine, glucose, amino acid, and pH indicator

ODC test

differential for production of orthinine decarboxylase, contains orthinine, glucose, amino acid, pH indicator

lysine/orthinine decarboxylase

enzyme that under acidic conditions (fermentation of glucose) functions to decarboxylate amino acids and produce alkaline conditions that revert the broth

48 hours

LDC/ODC test; incubation time

positive

LDC/ODC test; turbid purple broth

negative

LDC/ODC test; yellow or clear purple broth

KIA slant

differential for glucose and lactose fermentation as well as production of H2S and CO2. Contains peptone, glucose, lactose, ferric ammonium citrate, sodium thiosulfate, phenol red. Must stab butt and then do fishtail streak

positive for CO2 gas

KIA slant; bubbles in agar

negative for glucose fermentation

KIA slant; red butt

positive for glucose fermentation

KIA slant; yellow butt

positive for H2S and glucose fermentation

KIA slant; black butt

positive for lactose fermentation

KIA slant; yellow slant

negative for lactose fermentation

KIA slant; red slant

Gram stain

differential stain; tests for gram staining quality of the cell wall (peptidoglycan thickness)

positive

Gram stain; purple

negative

Gram stain; pink

crystal violet

Gram stain; primary stain

gram's iodine

Gram stain; mordant

alcohol acetone mixture

Gram stain; decolorizer

safranin

Gram stain; secondary stain

acid fast stain

differential stain; tests for mycolic acid in cell walls, somewhat hydrophobic and does not stain well with gram stain

positive

acid fast; pink

negative

acid fast; blue

carbol fuschin

acid fast; primary stain + heat

acid alcohol mix

acid fast; decolorizer

methylene blue

acid fast; secondary stain

endospore stain

structural stain for production of endospores

malachite green

endospore; primary stain + heat

water

endospore; decolorizer

safranin

endospore; secondary stain

add enough inoculum to fill tube, add oil

underlined wells of API20E

fill tube and cupule

boxed wells of API20E

fill tube

all other wells of API20E

direct elisa

detect presence of particular antigen in a sample, antibody carrying reporter enzyme is bound directly to the target antigen

Indirect elisa

detect presence of specific antibody directed against a particular antigen, or presence of a particular antigen, enzyme will only be present to react with substrate if the secondary antibody that it is attached to which is also attached to the primary antibody

antigen capture elisa

antibody directed against a target antigen is coated directly onto a solid matrix. Binds with any antigen present in the sample. Used to detect presence of a particular antigen but also used to detect presence of antibody directed against a particular antigen.

antibody capture elisa

directed against a particular antibody type, is coated directly onto a solid matrix. Second antibody becomes sandwiched between the capture antibody and the antigen. Captured by the antibody bound to the solid matrix.

phenol red

pH indicator used for PR carbohydrate test

neutral red

pH indicator used for MacConkey Agar