Agar plate orientation

Different medium

What type of tube is this?

Carbohydrate Fermentation tube

Used to determine the ability to ferment carbohydrates (glucose, lactose, mannitol, sucrose & maltose broths w/ phenol red indicator)

Results:
acid production = yellow (pH< 7)
gas production = bubble formation in Durham tube
negative fermentation = red/pink no gas & not yellow (pH 5)

Glucose (Carb. test)

Left: Fermentation with acid & gas production (A/G) bubble in tube
2nd: Fermentation acid without gas (A/-) no bubble in tube
3rd: control. 4th: No fermentation no reaction (-/-) red no bubble in tube
5th: degradation of peptone w/alkaline end products (K) no bubble in tube

IMViC

Methyl red, Voges-Proskauer, Indole and Citrate

used to distinguish between members of the family Enterobacteriasea and differentiate them from other Gram-negative rods

Methyne red

Mixed acid fermentation

Left MR-positive (+) red / mixed acid fermentation

Right MR-negative (-) no color change/ yellow / no mixed acid fermentation

Voges–Proskauer

left VP- negative (-) / copper color at top means reaction of KOH and alfa-naphthol / no color change / no 2, 3 butanediol fermentation (no acetoin produced)

right VP-positive (+) / red / 2, 3 butanediol fermentation (acetoin produced)

Indole

Organism converts tryptophan to indole

Simmons Citrate slants

Left tube: uses citrate (+) blue
Center tube: does not use citrate (-) no color change
Right tube: control inoculated

Simmons Citrate Agar-used to determine if capable of using citrate as the sole source of carbon w/ production of enzyme citratase.

-Medium contains sodium citrate, ammonium salts & brom thymol blue (pH indicator)
-Procedure: stab/streak method then incubate 24-48 hrs

Nitrate

Organism reduces NO3 --> NO2

Nitrate. Reagents A+B added. Interpret tubes

Tube 1 (left): red, positive result. Nitrate reduced to nitrite
Tube 3, 4: inconclusive, no color change. Must add zinc dust.
Tuble 2: control

Nitrate. +Zinc, interpret tubes 3, 4

Tube 3: red color indicates nitrate is still present. Negative result. Organism does not reduce nitrate.

Tube 4: no color change, organism reduced nitrate to some other nitrogenous compound. Positive result.

Starch Hydrolysis

Top: no clearing, organism does not break down starch.

Bottom: clearing in medium, organism breaks down starch (amylase is present)

Microorganisms excrete hydrolytic extracellular enzymes in order to degrade starch

Glucose

That will be transported into the cell and use for energy production (Glycolysis)

Caesin Hydrolysis test

Casein (proteases)

Organisms producing proteases will show a zone of lipolysis (clear area surrounding the bacterial growth)

Caesin Hydrolysis test

Top: casease-positive (+) present of casease
bottom: casease-negative (-) absent of casease

Lipid Hydrolysis test

Above lipase-positive (+) lipase present / clearing in agar around growth

Below lipase-positive (-) absent of lipase / no clearing in agar around growth

Lipid Hydrolysis test

Lipid ( tributyrin ) (lipases)

Organisms producing lipases will show a zone of lipolysis (clear area surrounding the bacterial growth)

Urea hydrolysis

Left urease-positive (+) rapid urea splitter hydrolysis; strong urease production
Center uninoculated control
Right Urease-negative (-) no urea hydrolysis; urease absent

Gelatin Hydrolysis test

Top: liquid gelatin indicates gelatinase-postive (+) Gelatin is liquid (control is solid) Gelatinase is present in organism

Bottom: solid gelatin indicates gelatinase-negative (-) Gelatin is solid. No gelatinase is present in organism

Organisms producing proteases will show liquefaction of the gelatin, low temperatures of 4°C will not restore the gel characteristic.

SIM MEDIA TEST

SULFUR POSITIVE vs Sulfur NegativeIndole Negative in both(Note: Indole indicator: Kovac)

MOTILITY POSITIVE vs Motility Negative(Note: Motility Indicator: Stabline)

Sulfur reduction in SIM medium
-when a species can reduce sulfide there is a black precipitate that forms turning the agar black.

Left No black in the medium / Sulfur is not reduced / H2 S-negative (-)

Right Black in the medium / Sulfur reduced / H2 S-positive (+)

Indole test

Left indole-negative (-) organism Reagent color is unchanged Tryptopha is not broken down into indole ad pyruvate

Right indole-positive (+) organism / Red in the alcohol layer of Kovac's reagent. Tryptopha is broken down into indole ad pyruvate

Motility in SIM

Left motile organism / Growth radiating outward from stab line

Right nonmotile organism / Non radiating growth

Litmus milk Medium