Lab 24: Oxidative and Fermentation Tests

What is the purpose of exoenzymes?

Enzymes that are exported outside of the cell to break down large molecules into smaller subunits that can be transported into the cell

What are the two processes of generating energy used by the cell?

Oxidative respiration or fermentation

What is the final electron acceptor for oxidative respiration?

Oxygen

Which yields more energy, oxidative respiration or fermentation?

Oxidative respiration

What is the final electron receptor for fermentation?

Organic molecules

Why can fermentation occur under more conditions than oxidative respiration?

Does not require oxygen

What type of organism can only use oxidative respiration?

Obligate aerobes

What type of organism are incapable of surviving in an environment containing oxygen?

Facultative anaerobes

What does the O/F test determine?

Whether oxidative metabolism or fermentative pathway was used

What medium is needed for the O/F test?

Hugh and Leifson’s

What is the pH indicator for the O/F test?

Bromothymol blue dye

How does fermentation affect pH?

Decreases it because of acid production

What is done to create anaerobic conditions and encourage fermentation in O/F test?

Mineral oil dispensed on top to decrease oxygen

O/F test with and without oil shows a green/blue color. What is the result?

Negative, no carbohydrate metabolism

O/F test with and without oil shows a yellow color. What is the result?

Oxidative and fermentative metabolism (O/F)

O/F test with oil shows a green color and without oil shows a yellow color. What is the result?

Oxidative metabolism (O)

O/F test with oil shows a yellow color and without oil shows a green color. What is the result?

Fermentation (F) 

What is the purpose of fermentation tests?

If a particular sugar can be used as an energy source and how

In a fermentation test, methyl red shows a red color. What is the pH?

<4.4

In a fermentation test, methyl red shows a orange color. What is the pH?

5.2

In a fermentation test, methyl red shows a yellow color. What is the pH?

>6.0

In a fermentation test, phenol red shows a yellow color. What is the pH?

<6.8

In a fermentation test, phenol red shows a orange color. What is the pH?

7.9

In a fermentation test, phenol red shows a red color. What is the pH?

>8.4

In a fermentation test, bromothymol blue shows a yellow color. What is the pH?

<6.0

n a fermentation test, bromothymol blue shows a green color. What is the pH?

7.0

In a fermentation test, bromothymol blue shows a blue color. What is the pH?

>7.6

For a fermentation test, what is used in the medium to capture gas?

Durham tube

Which broths use phenol red as a pH indicator?

Glucose, lactose, mannitol

What is mannitol?

Sugar alcohol that is a stereoisomer of sorbitol used by diabetics as a sugar substitute

What two tests can the MR-VP broth do at once?

Methyl red test and Voges-Proskauer test

What is the purpose of the MR-VP test?

Distinguish between different enteric microorganisms

What does the Methyl Red (MR) test observe?

Mixed acid fermentation by measurement of pH

What is the pH indicator for Methyl Red test?

Methyl red

What does the Voges-Proskauer (VP) test observe?

Detect organisms that can ferment glucose and convert acid produced to acetoin and 2,3-butanediol

What is added to detect presence of acetoin for Voges-Proskauer test?

Barritt’s reagent

What can be used instead of Barritt’s reagent for Voges-Proskauer test?

15 drops of Voges-Proskauer reagent A and 5 drops of Voges-Proskauer reagent B

What color is a positive VP test?

Pink to red

What medium is used for the Citrate test?

Simmon’s citrate agar

What substances are in Simmon’s citrate agar?

Sodium citrate, ammonium dihydrogen phosphate, bromomythol blue pH indicator

What is being detected in the Citrate test?

Use of fermentation pathway to break down citrate and produce amino acids

What indicates a positive Citrate test?

Blue

What indicates a negative Citrate test?

Green

How does fermentation of citrate and use of ammonium dihydrogen phosphate affect pH?

Increases

What medium is used for the catalase test?

TSA agar

What medium is used for the Nitrate Reduction test?

Nitrate broth

What is the catalase test used for?

Detect if catalase was/was not produced

For the catalase test, what is added to produce results?

3% H2O2

What are the positive indications for catalase test?

Bubbles

For the nitrate reduction test, what is added to produce results? 

Nitrate reagent A/Sulfanilic acid and Nitrate reagent B/dimethyl-alpha-naphthylamine

For the nitrate reduction test, what is indicated for the presence of NO2?

Dark red

What type of organism uses nitrate (NO3) as a final electron acceptor?

Gram-negative rods in anaerobic respiration

What are 3 possibilities for why nitrate reduction test yields negative results?

Organism does not reduce nitrate. Organism did reduce nitrate to nitrite but it can make nitrite reductase which further reduces nitrite to ammonia. Nitrate has been denitrified; nitrite reduced to nitrogen gas.

For the nitrite reduction test, what is added to clarify negative results?

Nitrate reagent C (Zinc)

For nitrite reduction test, what is indicated to confirm the negative result after adding Nitrate reagent C?

Red color