Purpose:
To identify an unknown microorganism by performing a series of biochemical tests on a pure bacterial culture.
Materials and Methods:
Carbohydrate Utilization: Materials:
- Phenol red lactose Broth
- Phenol Red Sucrose Broth
- Unknown organism # 40
- Inoculating tool
Procedure:
Phenol red lactose and Phenol Sucrose were cultures used to identify fermentation of carbohydrates for the unknown organism #40. Phenol red is a ph. indicator as the inverted Durham tube indicates gas production. The unknown organism was inoculated into each phenol red broth with either sucrose or lactose. The inoculating tool was sterilized by heat between each inoculation. The inoculated broth was then incubated at 37℃ for 24-48 hours. The negative outcomes are: medium broth remains red, no color change. The color of red to magenta is a negative result. The positive outcomes are: medium broth turns from red to yellow, production of acid. The medium turns yellow and a gas bubble in the Durham tube indicate acid and gas production. Indole Production: Materials: - 1% Tryptone broth - Kovac’s reagent - Unknown organism # 40 - Inoculating tool Procedure: 1% Tryptone is a Tryptophan saturated broth that will indicate the breakdown of tryptophan to Indole. …show more content…
Unknown organism #40 was inoculated into 1% Tryptone. Inoculating tool was sterilized with heat before and after inoculation. The inoculated broth was incubated at 37℃ for 24-48 hours. Kovac’s reagent was then added into the inoculated broth after incubation period. Negative outcome of experiment is a remained yellow ring in the reagent layer on top of the broth. A positive outcome is the appearance of a red ring in the reagent layer on top of the broth.
Urea Hydrolysis: Materials:
- 1 Urea broth
- Unknown Organism #40
- Inoculating tool
Procedure:
Urea broth contains phenol red a ph. indicator, urea and yeast extract to detect the hydrolysis of urea. Unknown organism #40 was inoculated into the Urea broth. Inoculating tool was sterilized with heat before and after inoculation. The inoculated broth was incubated at 37℃ for 24-48 hours. Positive outcome of experiment is the color change from orange to magenta. The negative outcomes are no color change or a color change of orange to yellow. Starch Hydrolysis: Materials: - Starch Agar Plate - Unknown organism # 40 - Iodine - Inoculating Loop Procedure: A Starch agar plate made up of nutrient agar and starch is used to detect amylase activity. One loopful of the unknown organism #40 was inoculated in a straight line onto the starch agar plate. Inoculating tool was sterilized with heat before and after inoculation. The inoculated starch plate was then incubated at 37℃ for 24-48 hours. After incubation period the starch agar plate was completely covered with iodine. A positive outcome of this experiment is a clear zone around the bacterial growth with no color formation complex. A negative outcome is the media will turn from straw colored to dark blue or black. Casein Hydrolysis: Materials: - Skin milk Agar - Unknown Organism #40 - Inoculating Loop Procedure: A skin milk agar plate is used to detect the hydrolysis of casein. One loopful of the unknown organism # 40 was inoculated in a straight line onto the skim milk agar plate. Inoculating tool was sterilized with heat before and after inoculation. The inoculated skim milk plate was then incubated at 37℃ for 24-48 hours. A positive outcome of this experiment is a clear zone surrounding the growth. A negative result is no clear zone forming around the organism. Catalase Production: Materials: - Nutrient Agar Plate - Unknown Organism # 40 - 3% Hydrogen Peroxide - Inoculating Loop Procedure: A nutrient agar plate is used to detect enzyme break down of hydrogen peroxide. One loopful of unknown organism # 40 was inoculated in a straight line onto the nutrient agar. . Inoculating tool was sterilized with heat before and after inoculation. The inoculated nutrient agar plate was then incubated at 37℃ for 24-48 hours. A positive outcome is the formation of gas bubbles, a negative result will not produce any gas. Sulfide Production: Materials: - SIM (Sulfide Indole Motility) - NA slant of Unknown Organism # 40 - Inoculating Needle Procedure: To detect the formation of Hydrogen sulfide a
To identify an unknown microorganism by performing a series of biochemical tests on a pure bacterial culture.
Materials and Methods:
Carbohydrate Utilization: Materials:
- Phenol red lactose Broth
- Phenol Red Sucrose Broth
- Unknown organism # 40
- Inoculating tool
Procedure:
Phenol red lactose and Phenol Sucrose were cultures used to identify fermentation of carbohydrates for the unknown organism #40. Phenol red is a ph. indicator as the inverted Durham tube indicates gas production. The unknown organism was inoculated into each phenol red broth with either sucrose or lactose. The inoculating tool was sterilized by heat between each inoculation. The inoculated broth was then incubated at 37℃ for 24-48 hours. The negative outcomes are: medium broth remains red, no color change. The color of red to magenta is a negative result. The positive outcomes are: medium broth turns from red to yellow, production of acid. The medium turns yellow and a gas bubble in the Durham tube indicate acid and gas production. Indole Production: Materials: - 1% Tryptone broth - Kovac’s reagent - Unknown organism # 40 - Inoculating tool Procedure: 1% Tryptone is a Tryptophan saturated broth that will indicate the breakdown of tryptophan to Indole. …show more content…
Unknown organism #40 was inoculated into 1% Tryptone. Inoculating tool was sterilized with heat before and after inoculation. The inoculated broth was incubated at 37℃ for 24-48 hours. Kovac’s reagent was then added into the inoculated broth after incubation period. Negative outcome of experiment is a remained yellow ring in the reagent layer on top of the broth. A positive outcome is the appearance of a red ring in the reagent layer on top of the broth.
Urea Hydrolysis: Materials:
- 1 Urea broth
- Unknown Organism #40
- Inoculating tool
Procedure:
Urea broth contains phenol red a ph. indicator, urea and yeast extract to detect the hydrolysis of urea. Unknown organism #40 was inoculated into the Urea broth. Inoculating tool was sterilized with heat before and after inoculation. The inoculated broth was incubated at 37℃ for 24-48 hours. Positive outcome of experiment is the color change from orange to magenta. The negative outcomes are no color change or a color change of orange to yellow. Starch Hydrolysis: Materials: - Starch Agar Plate - Unknown organism # 40 - Iodine - Inoculating Loop Procedure: A Starch agar plate made up of nutrient agar and starch is used to detect amylase activity. One loopful of the unknown organism #40 was inoculated in a straight line onto the starch agar plate. Inoculating tool was sterilized with heat before and after inoculation. The inoculated starch plate was then incubated at 37℃ for 24-48 hours. After incubation period the starch agar plate was completely covered with iodine. A positive outcome of this experiment is a clear zone around the bacterial growth with no color formation complex. A negative outcome is the media will turn from straw colored to dark blue or black. Casein Hydrolysis: Materials: - Skin milk Agar - Unknown Organism #40 - Inoculating Loop Procedure: A skin milk agar plate is used to detect the hydrolysis of casein. One loopful of the unknown organism # 40 was inoculated in a straight line onto the skim milk agar plate. Inoculating tool was sterilized with heat before and after inoculation. The inoculated skim milk plate was then incubated at 37℃ for 24-48 hours. A positive outcome of this experiment is a clear zone surrounding the growth. A negative result is no clear zone forming around the organism. Catalase Production: Materials: - Nutrient Agar Plate - Unknown Organism # 40 - 3% Hydrogen Peroxide - Inoculating Loop Procedure: A nutrient agar plate is used to detect enzyme break down of hydrogen peroxide. One loopful of unknown organism # 40 was inoculated in a straight line onto the nutrient agar. . Inoculating tool was sterilized with heat before and after inoculation. The inoculated nutrient agar plate was then incubated at 37℃ for 24-48 hours. A positive outcome is the formation of gas bubbles, a negative result will not produce any gas. Sulfide Production: Materials: - SIM (Sulfide Indole Motility) - NA slant of Unknown Organism # 40 - Inoculating Needle Procedure: To detect the formation of Hydrogen sulfide a