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Biology 26 - Microbiology

Biol 26  

Syllabus  

Biol 26 Lectures  

Biol 26 Labs 

 

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Biology 10 - Introductory Biology

Course Syllabus (Green Sheet)  

Lectures

Labs 

 

MR - VP

MR-VP test (Methyl Red, Voges-Proskauer)

 

The Methyl Red test:
  • a differential test for bacterial respiration 
  • used to distinguish/differentiate strains of coliform bacteria (i.e., Escherichia and Enterobacter)
  • methyl red test contains a small amount of carbohydrate fermentable by both organisms
    • typical strains of E. coli ferment CHO’s, producing a variety of acids, producing a pH of 4.5 or lower
    • typical strains of Enterobacter using the same quantity of CHO’s, are not capable of producing so low a pH
      • some end-products of Enterobacter are nonacidic, e.g., acetoin, ethyl alcohol, acetyl methyl carbinol
      • this organism produces only a partial assortment of acid end products
    • the Methyl Red test will be red (+) for E. coli and yellow (-) for Enterobacter

The VP test:

  • confirms the differences between E. coli and Enterobacter
    • E. coli does not form acetoin
    • Enterobacter does
    • this is a simple test to detect acetoin in the medium

The MP-VP combined test

  • the combination of these 2 tests helps confirm the separate identification of these 2 organisms
  • the test demonstrates the practical use of a good

    understanding of microbial respiration

MP Test (fermentation test)

 

 

Escherichia

Enterobacter

Ferments:

Glucose

Glucose

Products:

Abundant assortment of acidic products

Neutral, acetoin, etc

pH results:

< 4.5 or lower

Above 6.0

Resulting in?

Acidic solution, inactivates enzymes, growth stops

Growth never stops, when glucose is exhausted, organism will use peptone which produces alkaline by-products

Add indicator, Methyl Red

Positive (+) = red

Negative (-) = yellow

 

VP Test (production of acetoin)

 

 

Escherichia

Enterobacter

Add potassium Hydroxide + creatine (KOH) & alpha-naphthol,

 

 

Shake vigorously, let stand ~ 5-10 minutes, some references indicate1-2 hours, exposed to air.

Observe at intervals of minutes, 12, 24 hours

 

Negative (-)  = light brown color

Positive (+) = Red/pink

NOTE: the color reaction develops particularly in that part of the medium exposed to air

 

MR-VP Medium

  • buffered peptone
  • dipotassium phosphate
  • dextrose
  • pH 6.9

Day

Procedure

Organisms

1 Monday, 7/24/06

·        Work in groups of 4

·        inoculate MR-VP broth tubes with loop

·        tubes

1.    E. coli

2.    Enterobacter

3.    unknowns for group (2-3 or 4)

·        INCUBATE, 37oC, 48 hr

·        E. coli

·        Enterobacter

·        unknowns

2 Wednesday, 7/26/06

·        add 5 drops of the pH indicator Methyl Red to each tube

·        RECORD results

·        to the same tubes add 40% KOH + creatine and alpha-naphthol solution

·        SHAKE TUBES

·        allow the tubes to stand, see time information above

·        RECORD results

NOTE: formation of red color with acetoin requires large amounts of oxygen, shake vigorously (but CAREFULLY)

See also: http://facultyfiles.deanza.edu/gems/gilleskay/Exercise23VogesProskauertest.doc

 Updated Thursday, October 30, 2008 at 7:52:21 AM by Kay Gilles - gilleskay@fhda.edu
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