Bacterial fermentation

Bacteria mainly produce ATP by :

1. Aerobic respiration : is an oxidative process which uses oxygen as a final electron acceptor.# ATP/glucose : 36-38

2. Fermentation : uses an organic molecule as a final electron acceptor.# ATP/glucose : 2organic molecule are mainly carbohydrates (glucose, lactose, maltose, sucrose …etc)

3. Anaerobic respiration : is similar to aerobic respiration, but it uses an inorganic molecule other than oxygen as the final electron acceptor.# ATP/glucose : The ATP yield per glucose molecule varies, depending on the final electron acceptor used.inorganic molecule may be : Fe, Mn, Co, NO3, sulfate or others

all fermentative bacteria can ferment the simplest sugar {dextrose (D-glucose)}

Fermentation end products:

1. Acid end product2. Acid and gas end product

• Detection of acid production is carried out by pH indicator like phenol red which turns to yellow below pH 6.8 (pH decreases by acids production)

6.8 8.4• Detection of gas production carried

out by Durham tube

MR-VPtest

objective

Used to differentiate between enteric bacilli (Coliform) as well as indole and citrate test {IMViC tests}

This test composes from to portions1. The MR portion (methyl red) is used to determine if

glucose can be converted to acidic products like lactate and acetate.

2. The VP portion (Voges-Proskauer) is used to determine if glucose can be converted to acetoin.

Principle

A)Methyl red

• Some Coliform will ferment the dextrose to acid products that will cause the pH to drop below pH 5. This is called a mixed acid fermentation.

• After incubation the addition of methyl red, a dye which turns red below pH 4.4, will indicate whether such fermentation has occurred.

4.4 6.2

B) Voges-Proskauer test

• Other coliforms will convert dextrose to less acidic products such as ethanol or butanediol. These bacteria are negative in the methyl red test.

• Butanediol fermentation is demonstrated by the Voges-Proskauer test which measures the presence of acetoin (acetyl methyl carbinol), a precursor to butanediol.

• This test uses the same medium as the methyl red test and

both tests are usually performed in parallel.

• Barritt‘s reagents, 5% alpha-naphthol (vp1) and 40% potassium hydroxide(vp2), are added to a 48 hour culture and the tube is shaken to aerate the solution. The development of a pink or red color after agitation is a positive reaction for the production of acetoin

Voges-Proskauer equation

Limitations

• Most members of the family Enterobacteriaceae give either a positive MR test or a positive VP test. However, certain organisms such as Hafnia alvei and Proteus mirabilis may give a positive result for both tests.

• Non-fermentative bacteria will give –ve results for both tests (pseudomonas).

• Read the VP test at 48 hours. Increased incubation may produce acid conditions in the broth that will interfere with the readings of the results.

• VP reagents must be added in the order and the amounts specified or a weak-positive or false-negative reaction may occur. A weak-positive reaction may be masked by a copper-like color which may form due to the reaction of KOH and a-naphthol.