IMViC-Test---identification-of-enteric-bacteria.ppt

Click to edit Master title style,Click to edit Master text styles,Second level,Third level,Fourth level,Fifth level,*,IMViC Test,IMViC is a series of tests that are useful in the identification of enteric bacteria,Tests include:,1.I=Indole test,2.M=Methyl red test(MR),3.Vi=Voges-Proskauer test(VP),4.C=Citrate test,Indole Medium,Tryptone broth,contains extra tryptophan,Trp.(tryptophan)can be utilized as a sole carbon and energy source by some bacteria that produce an enzyme tryptophanase,(tryptophanase),Tryptophan,Indole,+pyruvic acid +NH3,(C-source)(N-source),We test the presence of the bi-product Indole with a chemical called Korvacs reagent,Korvacs reagent reacts with Indole and turns solution red,The alcoholic layer concentrate the red color as a ring at the top.,Indole Procedure,Procedure,:,1,.Inoculate tryptone broth with young bacterial culture.,2,.Incubate at 37 degrees for 48 hours,Indole Results,Results,:,Red ring at the top:+ve;,E.coli,Coloreless ring at the top:-ve;e.g.,Staphylococcus aureuos,Klebsiella,Methyl red,Strains of,E.coli,are mixed acid fermenters;they degrade carbohydrates into acidic end products such as:lactic acid,acetic acid,succinic acid,and formic acid,MR(continued),The Methyl-Red tests for acidic products resulting from the ability of an organism to produce and maintain stable acid end products from glucose fermentation.,These acidic products will drop the pH of the medium to pH 4.5 or below,Methyl red is a pH indicator will turn red if mixed acid fermentation has occurred and remain yellow in pH over 6.2.,The principle:,Glucose Fermentation (acidic end products)MR+,Hydrogenlyase,Glucose 2pyrovate (Lactic acid,CO,2,+H,2,(MR+),formic acid,acetic acid),The Procedure:,Incubate 5 ml of the broth with young bacterial culture for 24-48 hrs at 37C.,Add methyl red reagent into the broth.,The results:,Red color solution:+ve;e.g.,E.coli,faint yellow=-ve;,Klebsiella,The principle:,This test is used to detect microorganism that ferment glucose via the butanediol pathway produce acetoin as an intermediate which can be further reduced to 2,3-butanediol.,Material VP test:,Media(MR-VP media):,Peptone Glucose K,2,HPO,4,broth,Reagents:,VP reagent:,naphthol+KOH which is called(Barritts reagent).,The procedure:,Incubate 5 ml of the broth with young bacterial culture for 24-48 hrs at 37C.Add naphthol to the broth then add KOH.,The results:,Red color broth=+ve,e.g.,Klebsiella,pale yellow=-ve,e.g.,E.coli,Citrate Test(continued),The pH change is induced by CO2,which is given off as a bi-product of citrate utilization.When it reacts with Na and H2O in the agar it raises the pH above 7.6,The organism must contain the enzyme citrase to degrade citrate,EX:,(citrase),Citrate CO2+,Na,HCO +H2O,(blue color change),The materials:,Media:,Simmon Citrate slant agar,.,The Procedure:,The slant is streaked only with the required bacteria,Incubate at 37 degrees for 48 hrs.,Citrate Results,1.Green color is(-)for Citrase:eg.,E.coli,2.Blue color is(+)for Citrase:e.g.,Klebsiella,Urease test,This test is used to detect if the m.o.possess the enzyme,Urease,.,The Principle:,Urea+H,2,O,Ammonia,+CO,2,pH 6.8 pH8.1,Dark Orange Dark pink(,phenol red,is a pH indicator),Ureas,e,The Procedure,Inculate the urea agar with the required m.o.,Incubate at 37C for 24-48 hrs.,The results:,Pink color=+ve;e.g.,Proteus,;orange color=-ve,e.g.,E.coli,Catalase Test,This test used to detect the ability of m.o.(aerobic bacteria)produce the enzyme catalase that degrade H,2,O,2,.,The results:,Vigorous bubbling(due to O2 release)=+ve,Staph.,No bubbling=-ve result(no catalase is produced)e.g.,E.coli,