Proteins, amino acids, enzymes lab 24, sulfide, indole, motility agar

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1

Purpose of sulfur reduction test

Used to differentiate members of enterobacteriacae
ESP sulfur reducers from non reducers

2

What medium used in indole production, motility, and sulfur reduction test

SIM medium

3

What is SIM medium

Semi solid medium with casein and animal tissues as sources of amino acids
Iron containing compound
Sulfur in form of sodium thiosulfate

4

What is sulfur reduced to

H2S
Anaerobic activity, accomplished 2 ways depending in enzymes present

5

What are the two enzymes that reduce sulfur

1. Cysteine desulfurase
2. Thiosulfate reductase

6

Cysteine desulfurase

Catalyzes putrefaction of amino acid cysteine to pyruvate

7

Putrefaction of cysteine

Cysteine + h2o + h (cysteine desulfurase) pyruvate (for fermentation or respiration)
Byproduct of is h2s and nh3

8

Thiosulfate reductase

Enzyme that catalysts the reduction of sulfur (in form of sulfate) at the end of the anaerobic respiratory etc

9

What do both systems produce

H2s(hydrogen sulfide gas)
When either reaction occurs h2s produced combines with iron (ferrous ammonium sulfate in medium) to form ferric sulfide (feS) a black precipitate

10

What is blackening of media indicating

Positive test
H2S produced combines with ferris ammonium sulfate in media and forms ferric sulfide a back precipitate

11

Putrefaction of cystein

Reaction is mechanism for getting energy ut of amino acid cysteine

12

Indole test

Identifies bacteria capable of producing indole using enzyme tryptophanase
Uses SIM medium

13

How is Nicole production made possible in SIM medium

Presence of tryptophan (in casein and animal protien)

14

What does tryptophanase do?

Hydrolyses tryptophan to pyruvate, ammonia (by deamination) and indole

15

How is tryptophanase detected

Addition if kovacs reagent after incubation

16

Kovacs reagent

Contains DMABA AND HCL dissolved in amyl alcohol

17

What happens when kovacs reagent added to SIM

Forms liquid layer over solid media
DMABA then reacts with any ndole present and produces a quinoa dal compound that turns the reagent layer red

18

Formation of red layer

Positive reaction and presence of tryptophanase p 75

19

Motility test agar

Semisolid medium with reduced agar concentration to .4% just enough to maintain form but also a,low for motility

20

How is motility agar inoculated

Stabbing with needle

21

How is motility detected

As diffuse growth radiating from central stab line p 84


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