These 4 IMViC tests (actually 6 tests if you include motility and H2S) constitute, perhaps, the most critical tests THE INOCULATION PROCEDURES: Indole test . The four reactions are: Indole test, Methyl Red test, Voges Proskauer test and Citrate Procedure: Bacterium to be tested is inoculated in peptone water, which . In IMViC test there are four biochemical tests and this test are carried out IMViC test is carried out to identify members of the Procedure.

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Twst is a series of tests including the following tests: All of the series were designed to differentiate among the Enterics family Enterobacteriaceae though because many microbiology classes do not or cannot perform all of the bacterial tests, the IMViC tests are often used somewhat inappropriately to differentiate among the other families.

Simplified hour IMVic test: an agar plate method.

This test is often discussed as being positive if the tube prlcedure blue anywhere in the tube and negative if it remains green. Yet, this is really an oversimplification because it is only looking at the color change caused by a pH shift. In reality, there are two tests going on at once: Most bacteria can either use both or neither, but there are a ikvic exceptions. If the cell can, it will bring citrate ions into the cell to metabolize them to acetate and pyruvate.

Most bacteria can metabolize pyruvate, and some can metabolize acetate, to different products, depending on the pathway used, but usually to carbon dioxide and water. Most microbes in a microbiology class are aerobes and facultative anaerobes that use the most efficient way to metabolize a carbon source, aerobic respiration.

The metabolism of citrate removes a weak base from the imgic. Yet, if acetate is not metabolized, it will build up and replace the weak base, so the net change in pH is uncertain.

Methyl Red (MR) Test- Principle, Procedure and Result Interpretation

If the cell can reproduce by utilizing citrate, it will likely also utilize ammonium, removing it from the medium to make nitrogenous biomolecules. Leaving the phosphate ions behind, which are weak bases, and removing the tedt the ammonium ions, which proceduree weak acids, will increase the alkalinity of the medium and change the color to blue.

Unfortunately, occasionally other events occur which may interfere with the color change. So, ask your instructor how to score this test before interpreting your results.


Depending on how your unknown bacteria charts are designed, you may either use the interpretation based on color or the interpretation based on utilization.

It is really two tests: It starts as one, but it is incubated and then split into two tests. Provedure will inoculate a single tube for MR-VP test, and then split the contents into the two tubes to perform two tests at a later date.

This test includes the same reagents as the MR test. The addition of the VP reagents to acetoin produces a red color which is scored as a positive for the test. However, a copper color may result a side reaction and is negative for VP. Carefully distinguished copper from a red as the two are similar.

Procedurw color change or a copper color are negative for the VP test. Indole may be lost after prolonged incubation; however, Pickett found no loss after 96 hours. Also, although most media containing normal or higher amounts of protein may be used for the Indole test, the addition of tryptophan or peptides high in this amino acid tryptone stimulates the production of Indole whereas the addition of glucose inhibits it.

Fermentation of sugars also lowers the pH which inhibits tryptophanase activity. Escherichia coli is Indole positive, Enterobacter cloacae is negative.

Most imvix outside of family Enterobacteriaceae are also negative. As a liquid culture may add a source of carbon and nitrogen to the Citrate tubes, cultures should come from solid media. The Citrate test sometimes shows tiny growth in only one spot. This may be from the accidental introduction of a source of carbon to the tube, particularly if prpcedure many cells were added, or small amounts of agar medium were picked up on the needle and added with tezt culture.

Enterobacter cloacae usually has blue growth it grows and turns blueBacillus cereus and Kocuria varians usually have green growth have no color changeand Escherichia coli is usually negative. The few species that exhibit green growth may be doing something else that prevents the pH from changing. The chart above ignores one possibility, a bacterium that does not utilize citrate but does utilize NH 4. This tesst only with autotrophic bacteria which are not generally used, so they will be ignored.

Some of the references discuss the results of this test as if the fermentation of citrate occurred.

Fermentation is unlikely to occur in a slant that is open to the air. In fact, Pos et.


IMViC – Wikipedia

The Malonate and Acetate tests are very similar to the Citrate test except that malonate or acetate respectively are providing the only carbon source in the media. M R requires an extended incubation period. If fermentation of the media occurs, acidic intermediates are initially made pyruvic acidso that all enterics will initially be MR positive after a day of incubation.

Upon further incubation, fermentors that do not perform mixed acid fermentation will metabolize imviv acidic intermediates to mainly neutral products such as acetoin which will fail to overcome the tesr in the media.

Yet, organisms which perform mixed acid fermentation will produce a number of acidic end-products that will overcome the buffering capacity of the media. If a reduced incubation time is required, use a heavier inoculation and incubate for at least two days at degrees C to ensure that most organisms will test ok. Escherichia coli is positive, Enterobacter aerogenes is negative. VP tests for the production of acetoin, but this test is a little more complicated.

First, some organisms do not make acetoin and score as VP negative. Others produce acetoin but quickly metabolize it to something else, also scoring as negative.

Other organisms produce more or less stable acetoin, so they normally score as positves. Yet, acetoin is not the final end product so that an incubation period greater than 5 days may fail to detect acetoin because it has been metabolized further.

How is IMViC Biochemical test performed ?

Exactly when this happens depends on the organism. If more time is required, one may incubate the tubes and refrigerate them to avoid this problem. tes

Another problem is that some organisms are VP positive when grown for 5 days at 30 degrees or less, but are VP variable when grown at 35 degrees particularly Hafnia alvei. Collins and Lyne’s Microbiological Methods, 6th ed.

Difco Manual11th ed. Biochemical Tests for the Identification of Medical Bacteria, 3rd ed. Laboratory Exercises in Microbiology, 5th ed. Methods for identification of flavobacteria. Color Atlas and Textbook of Diagnostic Microbiology, 2nd ed. Citrate is utilized NH 4 is utilized score as “B” blue growth. Citrate is utilized NH 4 may or may not be utilized score as “G” green growth. Citrate not utilized NH 4 is not utilized score as negative.