8.2) Methyl Red/Voges Proskauer Test:(MR-VP)
Microorganisms when grown in MR-VP or Clark and Lub's medium containing a limited amount of carbohydrate ferment it to produce acids which lowers the pH of the medium. Certain genera go on producing more acid in a mixed acid fermentation making the pH too low. Basic principle of this test is to detect the ability of an organism to produce & maintain stable acid end products. If an organism produces large amount of organic acids like formic acid, acetic acid ,lactic acid, succinic acids as end products from glucose, the pH of the medium lowers down.
To test if sufficient amount of acid has been produced in the culture, a pH indicator methyl red is added to it. This indicator turns red at Ph 4.4 & below & becomes yellow at pH 6.2 & above. In a positive MR test, the reagent remains red indicating that large amounts of acids were produced by bacteria growing in the medium. The negative reaction is orange or yellow indicating less acid production.
All enteric bacteria catabolize glucose for their energy
needs; however, the end products vary depending on
the enzyme pathways present in the bacteria. The pH
indicator methyl red detects a pH
change to the acid range as a result of acidic end products
such as lactic, acetic, and formic acids. This test is
of value in distinguishing between E. coli (a mixed
acid fermenter) and E. aerogenes (a butanediol fermenter).
Mixed acid fermenters such as E. coli produce
a mixture of fermentation acids and thus acidify
the medium. Butanediol fermenters such as E. aerogenes
form butanediol, acetoin, and fewer organic
acids. The pH of the medium does not fall as low as
during mixed acid fermentation. At a pH of 6, the indicator
turns yellow—a negative methyl red test.
The Voges-Proskauer test (named after Daniel Voges,
German physician, and Bernhard Proskauer, German hygienist, in the early twentieth century) identifies bacteria
that ferment glucose, leading to 2,3-butanediol
accumulation in the medium. The addition of 40%
KOH and a 5% solution of alpha-naphthol in absolute
ethanol (Barritt’s reagent) will detect the presence of
acetoin—a precursor in the synthesis of 2,3-butanediol.
In the presence of the reagents and acetoin, a cherry-red
color develops. Development of a red color in the culture
medium 15 minutes following the addition of Barritt’s
reagent represents a positive VP test; absence of a red color is a negative VP test.
The following medically important bacteria are MR (+) : Escherichia
coli (opportunistic urinary tract infections), Salmonella
typhi (typhoid fever), Shigella dysenteriae (bacterial
dysentery), and Yersinia pestis (plague). The following
is MR(negative) : Enterobacter aerogenes (urinary tract infections).
Bordetella pertussis (whooping cough) is citrate negative
whereas all other Bordetella species are citrate positive.
The enteric bacteria such as Klebsiella pneumoniae
(pneumonia) and Enterobacter are citrate positive and can
be distinguished in the clinical laboratory from the opportunistic
pathogen Escherichia coli (urinary tract infections)
which is citrate negative.
Please refer to these links for more information;
Cited By Kamal Singh Khadka
Assistant Professor in Pokhara University, Regional College Of Science & Technology, Prithivi Narayan Campus
Nobel Academy, LA