Presentation about intrduction and types of anaerobic bacteria and about various methods used for cultivation of anaerobic bacteria

1. CULTIVATION OF
ANAEROBIC BACTERIA
Mr. Krishnakant B. Bhelkar
Asst. Professor,
Gurunanak College of Pharmacy, Nagpur
Unit I
Pharmaceutical Microbiology
B. Pharm III Semester

2. ANAEROBIC BACTERIA
An anaerobic bacteria or anaerobe is any organism that does not
require oxygen for growth.
It may react negatively or even die if free oxygen is present.
Depending on amount of oxygen tolerated, they are divided in
three types
Obligate Anaerobe
Aero-Tolerant Anaerobes
Facultative Anaerobes

3. ANAEROBIC BACTERIA
Depending on amount of oxygen tolerated, they are divided in
three types
Obligate Anaerobe
These are microorganisms killed by normal atmospheric
concentrations of oxygen (20.95% O2). Oxygen tolerance varies
between species,
Some capable of surviving in up to 8% oxygen, others losing
viability unless the oxygen concentration is less than 0.5%
Aero-Tolerant Anaerobes
Survive in presence of oxygen –
Do not use oxygen for energy requirements but
use fermentation to produce ATP. They do not utilize oxygen,
They can protect themselves from reactive oxygen molecules.
Facultative Anaerobes
A facultative anaerobe is an organism that makes ATP by aerobic
respiration if oxygen is present, but is capable of switching
to fermentation if oxygen is absent.

4. METHODS FOR CULTIVATION OF
ANAEROBIC BACTERIA
Candle Jar Method
Anaerobic Jar Method
Anaerobic Chamber
Vacuum and Gas Displacement Method
Thioglycollate Broth Method
Alkalline – Pyrogallol Method
Brewer Anaerobic Culture Plate Method

5. CANDLE JAR METHOD
In this method, media plates with bacterial inoculum is
kept inside the jar.
Seal the jar with the lit candle inside.
The candle flame will consume most of the oxygen in
the jar and will produce an elevated level of carbon
dioxide.
These conditions are ideal for the growth of anaerobic
and microaerophilic organisms.

6. CANDLE JAR METHOD

7. ANAEROBIC JAR METHOD
In this method, media plates with bacterial inoculum is
kept inside the jar.
The jar contains palladium pellets, that produces
hydrogen inside the jar.
The hydrogen reacts with oxygen present inside the jar
and form water molecules.
This cause reduction in oxygen concentration inside the
jar.
The anaerobic condition can be confirmed by indicator,
methylene blue. Indicator becomes colorless in absence
of oxygen
This condition is ideal for the growth of anaerobic
organisms.

8. ANAEROBIC JAR METHOD

9. ANAEROBIC CHAMBER
Anaerobic chambers, also known as anaerobic glove
boxes, are atmosphere control units designed to be used
when working with oxygen sensitive materials.
The chamber contains the sealed environment of H2,
CO2 or N2.
The media plates are kept in the chamber and then N2 is
purged in =side the chamber

10. ANAEROBIC CHAMBER

11. VACUUM AND GAS DISPLACEMENT
METHOD
Anaerobic In this method, the air in the chamber is removed
with the help of applying vacuum and is replaced with the
mixture of N2 and CO2,
This provides the good condition for cultivation of strict
anaerobes.

12. THIOGLYCOLLATE BROTH METHOD
Thioglycollate Broth is a
multipurpose, enriched, differential
medium used primarily to determine
the oxyen requirements of
microorganisms.
Sodium thioglycolate in the medium
consumes oxygen and permits the
growth of obligate anaerobes

13. ALKALLINE – PYROGALLOL METHOD
Alkaline solutions of pyrogallol absorb oxygen
efficiently and are used in determining the oxygen
content of gas mixtures

14. BREWER ANAEROBIC CULTURE
PLATE METHOD
Brewer Anaerobic Agar is used for
cultivating anaerobic and
microaerophilic bacteria
Brewer1 described a special Petri
dish cover that allowed surface
growth of anaerobes and
microaerophiles without anaerobic
equipment.
A small amount of air is caught over
the surface of the medium, and the
oxygen in this space reacts with the
reducing agents to form an
anaerobic environment.

15. BREWER ANAEROBIC CULTURE
PLATE METHOD
Brewer Anaerobic Agar Approximate Formula* Per Liter
Pancreatic Digest of Casein .......................... 5.0 g
Proteose Peptone No. 3............................. 10.0 g
Yeast Extract .............................................. 5.0 g
Dextrose .......................................... 10.0 g
Sodium Chloride .................................. 5.0 g
Agar ............................................... 20.0 g
Sodium Thioglycollate ................................ 2.0 g
Sodium Formaldehyde Sulfoxylate................ 1.0 g
Resazurin .............................................. 2.0 mg