Download Anaerobic Jar

General Microbiology Laboratory
Anaerobic Bacteria
Categories Based Upon Oxygen Requirements
• Aerobic bacteria
Require oxygen as electron acceptor
• Microaerophilic bacteria
Require oxygen in reduced quantity (2-10%)
• Facultative bacteria
Grow either with or without oxygen
• Anaerobic bacteria
Both obligate and aerotolerant (Aerotolerant do
not use aerobic metabolism but have some
enzymes that detoxify toxic forms of oxygen)
Methods for excluding oxygen
1. Fluid Thioglycollate Media (FTM)
• supports aerobic & anaerobic
bacteria.
• Sodium thioglycollate: removes O2
from the medium
• Resazurin as an oxidation
reduction indicator. dye that turns
pink in presence of O2
2- Anaerobic
Jar
Inoculate FTM tube :
B. subtilis or P. aeruginosa (strict aerobes)
S. pyogenes (microaerophile)
E. coli (facultative anaerobe)
C. sporogenes (strict anaerobe)
• Place in anaerobic jar.
• Incubate overnight at 37C.
Anaerobic Jar
Anaerobic Jar
 For this exercise, we use the GasPak anaerobic system. This
system consists of a polycarbonate jar, a lid with a gasket to
prevent air flow, a strip infused with methylene blue, and a
pouch containing sodium borohydride, sodium bicarbonate,
citric acid, and a palladium catalyst.
 When water is added to the pouch, the sodium borohydride,
sodium bicarbonate, and citric acid react to form hydrogen and
carbon dioxide. The palladium catalyzes a reaction between
the hydrogen and the oxygen within the jar; this reaction
creates water, which forms as condensation on the inside of the
jar.
 Methylene blue is blue in the presence of oxygen but is
colorless in an anaerobic environment. When the oxygen is
converted to water and condensation forms on the side of the
jar, the indicator strip will turn from blue to white.
Candle jars
 The candle flame will
consume most of the
oxygen in the jar.
 Produce an elevated level
of carbon dioxide.
 These conditions are ideal
for the growth of
microaerophilic
organisms.
Growing Microaerophilic Microbes
CO2 Generating Packet
Anaerobic Chambers
The Vinyl Anaerobic Chambers provide a
strict anaerobic atmosphere of 0-5 parts per
million (ppm) using a palladium catalyst and
hydrogen gas mix of 5%.
End of lecture