SUGAR FERMENTATION TEST PRINCIPLE:

Fermentations are energy producing biochemical reactions in which organic molecules severs as both electron acceptor and electron donors. The ability of microorganisms to ferment the carbohydrate the types of formed product are very useful in heir identification. The given carbohydrate may be converted into a number of different end product depending upon the microorganisms involved. The product may be acid gases alcohols are the characteristics of the that particular organism. Test culture suspension was taken from enriched broth Loopful culture was inoculated in peptone water base containing 1% respective sugar Incubate on shaker for 24 hrs at 37 c Observe the result and note down

IMVIC TESTS INDOLE TEST Principle:

The amino acid tryptophan is found in nearly all proteins. Bacteria that contain the tryptophanase can hydrolyze tryptophan to its metabolic products, namely, Indole, pyruvic acid,and ammonia. The bacteria use the pyruvic acid and ammonia to satis fy nutritional needs; Indol is not used and accumulates in the medium. The presence of indol can be detected by the addition of Kovacs reagent. Kovacs reagent reacts with the indol, producing a bright red compound on the surface of medium. Tryptophanindol, pyruvate and ammonia. Test culture suspension prepared in sterile saline Loopful suspension is inoculated in sterile peptone water base

All the tubes incubated at 37c for 24 hrs Results observed and noted CONFIRMATORY TEST 1. BILE TEST Inoculate the loopful inoculum in the bile medium (MRS + bile2% & 3%) Incubate over night on shaker at 37 c for 24 hrs. Check the turbidity in the medium The tubes which shows turbidity will carry cell adhesion

Inoculate the loopful enriched broth in the medium having pH1 & pH 3 Incubate the medium overnight on shaker at 37 c for 24 hrs Check the turbidity The tubes which shows turbidity will carry for cell adhesion

OXIDATIVE FERMENTATIVE TEST

Prepare the medium Inoculate the culture with the help of stab wire In the oxidative test only inoculum is added but in fermentative test, as soon as inoculum added, paraffin oil added to cover the surface. Check the color change and gas formation in the both tubes

ANTIBACTERIAL ACTIVITY

Prepare nutrient agar Spread the sensitive organism on the plate Plates were incubated in refrigerator for 10 min Make the well with the help of borer Inoculate the enriched broth in specific well After incubation, measure the zone of inhibition

NITRATE REDUCTION TEST Chemolithoautotrophic bacteria and many Chemoorganoheterotrophs can use nitrate as a terminal electron acceptor during anaerobic respiration. In this processes, nitrate is reduced to nitrite by nitrate reductase. Test culture suspension prepared in sterile saline Loopful suspension is inoculated in sterile peptone water base All the tubes incubated at 37 c for 24 hrs Results observed and noted

SUGAR FERMENTATION TEST

Fermentations are energy producing biochemical reactions in which organic molecules severs as both electron acceptor and electron donors. The ability of microorganisms to ferment the carbohydrate the types of formed product are very useful in their identification. the given carbohydrate may be converted into a number of different end product depending upon the microorganisms involved. the product may be acid gases , alcohols depending on the characteristics of the that particular organism. Test culture suspension was taken from enriched broth Loopful culture was inoculated in peptone water base containing 1% respective sugar

Incubate on shaker for 24 hrs at 37 c Observe the results and note down

OXIDASE TEST Oxidase enzyme plays important role in electron transport system. The production of the oxidase enzyme can be checked by using reagent N, N,N,N-Tetra methyl -P-Phenylendiamine dihydrochlride

A small quantity of bacteria is taken on filter paper disc soaked in reagent. In positive test, color of dye get changed.

CATALASE TEST Some bacteria contains flavoprotien that reduces O2 resulting production of hydrogen peroxide which is extremely toxic. Survival in the presence of antimetabolite is possible due to enzyme called as catalase. H2O2 2H2O + O2 Inoculate loopful test culture Dipped into catalase reagent

Observe effervesces in positive test

CERTIFICATE
This is to certify that Mrs. Ashwini A pensalwar studying in M.Sc. Microbiology, has worked on a project entitled during the period of in a partial fulfillment of the requirement for Masters degree in microbiology, is record of candidates own work carried out by her under supervision and guidance. The matter embodied this report has not been submitted for award for any other degree.

DELERATION

The project entitled has been submitted to university of pune as a partial fulfillment M.Sc degree under the guidance of Dr. suneeti Gore (Department of microbiology, Fergusson college pune) and literature cited has been written in my own words.

ASHWINI A. PENSALWAR M.SC. MICROBIOLOGY FERGUSSON COLLEGE, PUNE

Contents: number Objective Abstract Introduction Material and Methods Results Discussion References

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Appendix

Nutrient Agar
Peptone - 1gm Meat extract -0.3 Sodium chloride-0.5 Agar -2% D/W -100ml pH -6.8

Peptone water base

Peptone Sodium chloride D/W