Experiment 15: Metabolic Activities of Bacteria

what is carbohydrate fermentation?

some bact ferment simple carbs making acidic, alcoholic or gaseous end products in the process

many specieis can be distinguished based on?

1. carbs they use

2. carbs they dont use

3. nature of end products made in the fermentation rxn

what do media that test for carb fermentation have?

prepared as broth tubes with inverted fermentation/Durham tube for trapping gas

each broth has nutrients: one specificed carb + colour indicator bromocresol purple to indiviate change in pH if acid is produced

organisms that grow in the broth tube but dont ferment carbs?

NO colour change and NO gas production

some might make acid prodycts in fermenting sugar but NO gas and some might make acid and gas

organisms that dont ferment carbs might make alkaline/basic products by using?

protein nutrients in broth

detetc w colour change

what is indole?

by product of metabolic breakdown of AA tryptophan

how to test for indole production?

add Kovac’s reagent

YES indole = mixe w kovac, RED

NO indole = no colour other than colour of reagent

some bacteria split urea molecules to make?

CO₂ + NH_{4 —>} alkaline conditions = DARK PINK

mediated by urease enzyme can be seen if urea is added as substrate to culture

pH indicator is phenol red

which species rapidly produce urease?

Proteus species and some enterics

when is hydrogen sulfide made?

when amino acids containing sulfur are metabolized by microorganisms

if medium has metallic ions like Pb, Bi, Fe + appropriate AA, what happens?

the hydrogen sulfide combines w metallic ions to make metal sulfide = BLACK

what medium do we use to do hydrogen sulfide detetction?

SIM media - semisolid agar with Fe and blackens in the form of ferrous sulfide

also detects acid and visualizes bacterial motility after stabbing agar w needle and flagella will move away from the stab line if bact is motile

most that make hydrogen sulfide are also motile

exp 15 learning objectives

characterizie bact based on metabolism

analyze results of colour, gas, by-products

exp 15 materials (in fours)

1. E. coli

2. Staphylococcus aureus lvl 2

3. Bacillus subtilis

4. Proteus mirabis

5. purple glucose broth

6. pruple lactose broth

7. pruple surcros broth

8. urea broth w phenol red inside alr

9. SIM agar

10. sterile Pasteur pipette

exp 15 method

1. labe and put one drop of each bact into each tube

2. FOR SIM AGAR: use incinerator to sterilize needle and inoculate and stab straight down and bring it back up in the same path dont wiggle

3. incubate at 37deg for 24hr

4. put 3 drops of Kovacs in SIM agar

Results - Carbohydrate Fermentation

Results - Indole

Results - Urease Activity

Results - Hydrogen Sulfide Production