1. Exercise 39: Oxidation and Fermentation Tests
Lab 9 Goals and Objectives:
Exercise 39: Oxidation and Fermentation Tests
Read results: some tubes will require additional reagents
*Share and record all class results*
Exercise 38: Cultural Characteristics
Each person inoculate one nutrient gelatin deep
Exercise 40: Hydrolytic and Degradative Reactions
Each person inoculate one urea broth
Exercise 41: Multiple Test Media
Each person inoculate one SIM medium deep
Exercise 69: Gram Negative Intestinal Pathogens
Each person inoculate one MacConkey agar plate
Make new streak for isolation on BHIA
Each pair will need:
2 Nutrient Gelatin Deeps Methyl Red
2 Urea Broths Alpha Naphthol
2 SIM Medium Deeps KOH
2 MacConkey Agar Plates
2 BHIA plates

2. Durham Sugar Tube Fermentation (Lactose or Mannitol)
Inoculation method: loop transfer
Contains: single carbohydrate (lactose or mannitol) peptone
broth with durham tube for gas collection, Phenol red pH indicator: alkaline pH = red, acidic pH = yellow
Discriminates the ability to ferment a single carbohydrate (lactose or mannitol) into acid products (e.g. pyruvic acid) or acid plus gas
Results: Red = inert, negative for fermentation of specified
carbohydrate
Yellow = positive for fermentation of carbohydrate to acid products
Yellow with bubble = positive for fermentation of carbohydrate to acid + gas
Figure 39.7

3. Acid plus gas
Acid
Negative

4. _ + MR-VP Medium: Methyl Red Test Inoculation method: loop transfer
Contains: peptone, glucose, and buffer (buffer will neutralize weak acids so only strong stable acids will be detected by
methyl red)
Additional reagents added: methyl red pH indicator: acid pH = red, neutral or alkaline pH = yellow
Distinguishes ability to catabolize glucose into stable mixed acids (lactic, acetic, and formic acids) in the mixed acid pathway
Results: Red = positive for mixed acid
formation
Yellow = negative for mixed
acid formation _ +
Figure 39.8

5. _ _ + + MR-VP Medium: Voges-Proskauer Test
Inoculation method: loop transfer
Contains: peptone and glucose
Additional reagents added: Barritt’s A (alpha napthol) and Barritt’s B (KOH) (will react with acetoin to produce a red product, alone will produce a copper colored product)
Distinguishes the ability to catabolize glucose into the neutral end product butanediol (the oxidized product is acetoin) in the butylene glycol pathway
Results: Red = positive for acetoin and thus for 2, butanediol production
Yellow/Orange = no acetoin, negative for 2, butanediol production _ _ + +
Figure 39.8

6. _ + Simmon’s Citrate Agar
Inoculation method: streak and stab slant with needle
Contains: citrate as sole carbon source, ammonium salts as sole
nitrogen source, bromthymol blue pH indicator: neutral pH = green, alkaline = prussian blue
Discriminates organisms that can produce citrase to metabolize citrate into oxaloacetate and pyruvate. These organisms are forced to utilize ammonium salts as the nitrogen source producing alkaline ammonia waste.
Results: Prussian blue slant and or butt =
positive for citrase production
Green = negative for citrase
production _ +
Figure 39.9

7. New Inoculations from single colonies on BHIA:
Nutrient Gelatin: stab with needle
(Exercise 38)
Urea Broth: loop
(Exercise 40)
SIM Medium: stab with needle
(Exercise 41)
MacConkey Agar: streak for isolation with loop
(Exercise 69)
BHIA: streak for isolation with loop
Discard previous plates and tubes
Share all results with class

8. Exercise 39: Oxidation and Fermentation Tests
Lab 9 Goals and Objectives:
Exercise 39: Oxidation and Fermentation Tests
Read results: some tubes will require additional reagents
*Share and record all class results*
Exercise 38: Cultural Characteristics
Each person inoculate one nutrient gelatin deep
Exercise 40: Hydrolytic and Degradative Reactions
Each person inoculate one urea broth
Exercise 41: Multiple Test Media
Each person inoculate one SIM medium deep
Exercise 69: Gram Negative Intestinal Pathogens
Each person inoculate one MacConkey agar plate
Make new streak for isolation on BHIA
Each pair will need:
2 Nutrient Gelatin Deeps Methyl Red
2 Urea Broths Alpha Naphthol
2 SIM Medium Deeps KOH
2 MacConkey Agar Plates
2 BHIA plates