Proteins, amino acids and enzymes IMViC tests

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IMViC test

Methyl red
Voges proskauer


Methyl red test

Identifies bacterial ability to produce stable acid end products by mixed acid fermentation of glucose


Methyl red test and voges proskauer test medium




Soln with onky peptone, glucose and phosphate buffer


What do peptone and glucose provide

Protien and fermentatble carb


What is the phosphate buffer

Potassium phosphate resists pH changes in medium


MR test

Methyl red test
Detects organisms capable of performing mised acid ferm which overcomes phosphate buffer and lowers pH, acids produced are stable


How is mixed acid fermentation verified

Addition of methyl red indicator dye
Red at pH 4.4
Yellow at pH 6.2
Between two pH is orange


Red color

Only true positive result
Orange is negative or inconclusive
Yellow is negative


Mixed acid fermenter

E. coli p 82


Voges proskauer test

Identifies organisms able to produce acetoin from degradation of glucose during 2,3 butanediol fermentation


What was voges proskauer test designed for

Organisms that are able to ferment glucose but quickly convert acid end products to acetoin and 2,3 butanediol


Reagent in vp test

Barritts reagent
40% KOH


What happens when reagents added

Oxidizes acetoin (if present) to diacetyl which reacts with guanidine nuclei from peptone in medium and produces red color (positive result)


Copper color

Negative vp resulting from interactions between reagents
P 98


Citrate test

Used to determine organisms ability to use citrate as sole carbon source


Where is citrate used

Citrate (citric acid) is produced as acetyl coa (from oxidation of pyruvate or b oxidation of fatty acids) reacts with oxaloactetate at entry to Krebs cycle
Citrate then converted back to oxaloactetate beginning cycle again


What happens in medium containing citrate as only carbon source

Bacteria that have citrate permease can transport the molecules into the cell and enzymatic ally convert it to pyruvate
Pyruvate then converted to a variety of products depending on pH of enviornment


Citrate chemistry

P 64 draw image


If acidic pH what is formed

Pyruvate decarboxylated to form acetoin, acetate and lactate


If alkaline pH what is formed

Pyruvate converted to acetate and formate


Simmons citrate agar

Defined medium
Contains sodium citrate as only carbon source
Ammonium phosphate only nitrogen source


Bromthymol blue dye

Citrate test reagent
Green at pH 6.9
Blue at pH 7.6


What happens to bacteria that survive in bromthymol bkue

Utilize the citrate and a,so convert the ammonium phosphate to ammonia and ammonium hydroxide...both alkalinize the agar
As pH rises medium changes from green to blue


Positive citrate test

Conversion of medium to blue


No color change

Sometimes citrate positive organism will grow in citrate slant without producing color change bc too short incubation
No color change but growth on slant is positive citrate test


Image of citrate results



What is component in SIM deeps that makes medium suitable to detect production of indole by bacteria



What organic molecule is necessary to detect mixed acid fermentation by bacteria



Why do you shake the MR-VP CULTURE

to oxygenate the culture


Can a bacteria that ferments using the 2,3 butanediol path also use the mixed acid route?

Yes the use of butanediol path decreases acid formation and causes acetoin production


Why is chemically defined medium necessary for detection of citrate utilization by bacteria

Because citrate is onky carbon source

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