The PHENOL RED SUGAR or CARBOHYDRATE BROTH or Phenol Red  Fermentation TESTS: 

Atlas page 52

Carbohydrate Fermentation:   

The identification of some bacteria is aided by determining what nutrients the bacteria can utilize and what end products will be produced in the process. These characteristics are controlled by the enzymes which the bacteria produce. Because the type of enzyme(s) bacteria produce is genetically controlled, the pattern of sugars fermented may be unique to a particular species or strain. Fermentation products are usually acid (lactic acid, acetic acid etc.), neutral (ethyl alcohol etc.), or gases (carbon dioxide, hyrogen, etc.).

To determine the products of sugar fermentation, a carbohydrate fermentation broth containing phenol red pH indicator is prepared at pH 7.4 (it is RED a this pH and YELLOW below pH 6.8). This broth contains 3 essential ingredients: 0.5%-1.0% of the carbohyrate to be tested (WE  use: lactose, glucose (dextrose), sucrose, dulcitol, saccharose, sorbitol, and mannitol), nutrient broth, and the pH indicator phenol red. The nutrient broth, which is a light red color, supports the growth of most organisms whether they are able to ferment the sugar or not.

The test organism is inoculated into a broth containing the test sugar and incubated. A bright yellow color indicates the production of enough acid products from fermentation of the sugar to drop the pH to 6.9 or less. Production of gas is determined with a Durham tube, a small inverted vial filled with the carbohydrate fermentation broth. If gas is produced during fermentation of the sugar, it is trapped at the top of the Durham tube and appears as a bubble. Slow fermenters may take a week or more to cause color changes detectable by the human eye. Positive (yellow color or yellow color with gas bubble) and negative results (red color, no gas bubble) are shown in this image.

EVALUATION: incubate these for up to 3 labs or until they turn positive (yellow).  Record them as... FAST (with or without gas), Moderate (with or without gas), SLOW (with or without gas), or NEGATIVE (orange, red, or pink for 3 labs)

There are 8 possible answers for one of these tests if you inoculate on day 1 then:  Day 3 Fast pos with gas or without gas, Day 5 Moderately Fast positive with gas or without gas, Day 7 Slow positive with gas or without gas.  Assuming that you re-incubate each time it is NOT yellow.  If NOT yellow on Day 7 then record it is as negative.  Remember discard anything recorded as a positive (remove marks and tape before discarding).   EXCEPTION: If you have a tube the is whitish-pink with gas it is recorded as ALCOHOLIC.

 

Click on the image to see an enlarged version. 

Carbohydrate fermentation

      (+)without gas             (+)with gas         (-)negative           (-)negative

positive YELLOW tube with a gas bubble trapped in the Durham tube...

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