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BACTERIOLOGICAL MEDIA is the substrate on which bacterial culture is done and it provides nutrition
and pH to the organism
Liquid and Solid Bacteriological Media
Both are extensively used in bacteriological laboratories. To obtain a pure growth one has to use solid
medium because of the following disadvantages with the liquid media:
1. Growth does not show character on the basis of which a tentative diagnosis can be established.
2. If it is a mixed culture of more than one organisms, their separation cannot be performed.
However, once a pure growth has been obtained on solid media, liquid media are used to perform further
tests because of the homogenous growth in these media.
Agar Agar
Agar agar (commonly called as agar) is used as the most effective solidifying agent.
Properties of Agar are:
• Derived from certain seaweeds
• Melts at 95°C and solidifies at 42°C
• Remains firm at temperatures of incubation
• Heat labile material can be added to it even at temperature of 45°C thus retaining their potency
• Bacteriologically inert
• Resistant to action of all medically important bacteria
• Easily available
• Economical
Sugar Media
The term 'sugar' denotes fermentable substance. The usual sugar consists of 1 % of the chosen type of
sugar (lactose, sucrose, mannose, dextrose inulin, arabinose, etc) in peptone water alongwith an
appropriate indicator. A small tube (Durham's tube) is kept inverted in the sugar tube to detect gas
production.
Anaerobic Media
Medium such as Robertson's cooked meat medium is used for growing anaerobic organisms.
Preparation of Media
The components of any medium are combined together in recommended proportion without sterile
precautions. Sterilization process is then followed. All media are distributed as liquids. Blood for use in
bacteriology laboratory must be collected under sterile precautions from natural source and should be
preserved in sterile container. It should be rendered noncoagulable by defibrination or by the addition of
anti-coagulants such as citrates or oxalates. Serum can be sterilized by filtration.
The prepared medium is distributed in test tubes (prior to sterilization) and in petridishes (usually called
as plates).
Storage of Media
If the sterilized medium is in screw capped bottles, it can be stored at room temperature for weeks.
Medium in petri dishes deteriorates rather quickly at room temperature. Blood, serum and media can be
preserved for longer time if these are kept at low temperatures. Plates of media can be kept in a
refrigerator for 7 to 10 days. Screw capped bottles having media can be stored for months in refrigerator.
Blood and serum can be stored for still longer periods at -10°C to -40°C. Media should never be frozen
since it is detrimental to quality of nutrition.
CULTIVATION OF BACTERIA
The indications for culture of bacteria include:
· To obtain pure culture
· To characterize and confirm the isolate
· To carry out antibiotic sensitivity tests
· To carry out other epidemiological marker studies
· To estimate viable counts
· To store the bacterial isolates for future use.
ANAEROBIC CULTURES
For obligatory anaerobic bacteria, oxygen acts as a lethal poison and hence for the growth of these an
environment which is free of oxygen has to be created. The MacIntosh and Fildes technique of 1916 is still
widely practised with some modifications. In this the inoculated medium to be incubated anaerobically is
placed in the air tight jar and with the help of vacuum pump it is evacuated to -660 mm Hg. Hydrogen
(90%) and carbon dioxide (10%) are added to this through built in valves. The oxygen is removed by its
combination with hydrogen in the presence of palladium catalyst which is present in the jar. More
hydrogen or gas mixture is then introduced to equilibrate the pressure and the jar is incubated at 37°C.
Glove box or anaerobic chamber is used in modem laboratories with high quantum of work.