February ATMOSPHERIC O2 REQUIREMENTS MICOBIOLOGY-LAB SUBJECT OF

10, 2010 STUDY 6

Atmosphere O2 Requirements

Materials:
Nutrient Broth Culture of
• Bacillus cereus
• Escherichia coli Only aerobic cells High
• Micrococcus luteus

Medium:
• Broth Thiogloycollate Free exchange
Clostridium sporogenes of O2
The amino acids and glucose in the
medium can be respired, and glucose
is a fermentable source for organism.
as well as Clostridium which Decreased exchange Facultatively
can ferment amino acids. of O2 anaerobic cells

Equipment Used:
Bunsen burner
Inoculating loop Low
Gas pack anaerobic system Complete absents Only anaerobic cells
Test tube rack of O2

The observed growth patterns of organisms in this medium determine their O2 relationship
designation, which brings the abilities of respiration, fermentation, and the catalase reaction as
well as whether there is an inhibitory effect on the organism in the presence of air.

Top

Middle More
Area Even

Less Bottom

Obligate Aerobes Microaerophillic Facultative Aerotolerant Obligate Anaerobe

Anaerobes Anaerobes
Requires O2 for Needs a Greater Growth Anaerobic growth
growth limited amt. growth in occurs only. Ceases in
Occurs where high of O2 for the presents evenly O2 the presents of
Concentrate of O2 growth. of O2, but has no O2.
has occurs effect
throughout
Anaerobic Culture Method Reducing Media
• Contain chemicals thioglycolate that
Peroxide anion: combine O2
(2H2O2 catalase 2H2O + O2) (H2O2+2O+ peroxidase 2H2O)
• Heated to drive off O2
February ATMOSPHERIC O2 REQUIREMENTS MICOBIOLOGY-LAB SUBJECT OF
10, 2010 STUDY 6

Anaerobic Jar
This system is a temporary method for
the exclusion of O2 from a sealed jar
used for incubation of anaerobic cultures
in a non-reducing medium.
CO2 H2
• Duplicate sets of cultures are made
Anaerobic indicator
(methylene blue) • A single-line steak inoculation, of
Hydrogen Petri plate each test organism
Gas Pack (Inoculated)
generator • With corner of generator, insert inside
gas pack jar (one set of cultures in a
inverted position
When water is mixed with chemical pack, hydrogen
and carbon are generated • 10ml of H2O is added (with pipette) to
Hydrogen and atmospheric oxygen in the jar combine gas generator, quickly seal cap
to form water. Removing Oxygen
Indicator shows no color when O2 is removed • Jar is then incubated @ 37°F for 24-
48 hours