Bacterial Testing
Copyright © 1989 by Nan Fielding
former
NCS Microbiology Consultant
Copyright Notice & Disclaimer
Statement
Escherichia Coli (E Coli) and Salmonella are bacteria which belong to
the same family and tribe of microorganisms and infect in a similar
manner. Both cause diseases which can have varying symptoms in birds from
intestinal or renal involvement to septicemia (infection in the blood
stream). These organisms can also be present in birds in the carrier state
with no outward signs of disease but can be spread to susceptible
individuals causing disease.
Both organisms are gram negative rods which are not distinguishable by
Gram Stain from each other or from many other organisms of this type.
Therefore further testing must be done in order to make a definite
identification. This testing involves growing the organism in pure
culture, then inoculating from this pure culture onto a number of
different media (substances that the organisms can use as food) in order
to determine the growth characteristics and chemical activity of the
organisms involved.
Inoculation as used by the microbiologist means picking up a small
amount of a specimen or of a bacteria culture with a microbiologist loop
or needle and transferring it to the media which will be used for testing
or growth.
Some media contain chemicals which do not effect the growth of the
organism being sought but discourage the growth of other groups of
bacteria. The are called selective media. Some media contain
dyes or other chemicals that allow the observer to distinguish types of
bacterial colonies which develop after incubation by their appearance.
Such media are called differential media. Both of these
types of media are used in order to give the best chance of isolating a
pure culture of the organisms under study.
After a pure culture is obtained, other media containing specific
chemicals and designed to give specific reactions to the growth of
different organisms must be inoculated and incubated. After collecting the
results of all of these tests, the microbiologist refers to a table
containing the known reactions of various organisms to these tests and
makes a decision as to what organism is present. In order to run the
various tests the microbiologist must have available many different media.
For the small lab these media are available in a ready to use form. There
are also rapid tests for some of these reactions which are available in
kit form. These rapid tests usually require very small quantities of media
and inocolum and their accuracy is somewhat less than their comparable
full scale and full length tests but they can be valuable as a tool for
giving presumptive evidence in order to begin a treatment
program.
In keeping with my last article of introducing various pieces of
equipment which will be used in an avian microbiology lab, I will proceed
now to discuss two such pieces which are essential in performing these
bacterial tests.
An incubator must be used in growing your cultures since
these organisms do not grow easily at room temperature. The incubator you
use must have a thermostatically controlled heat source to maintain the
proper temperature and a laboratory thermometer to check the accuracy of
that temperature. I must caution each of you that an accurate thermometer
is essential and therefore, the "discount store" types should not be
considered. Provision must be made for maintaining proper humidity and air
circulation.
INCUBATORS
Such incubators can be purchased for approximately $275 for a small
bench top incubator up to many thousands of dollars for large ones of the
type used in commercial labs. Some people have effectively used incubators
like those intended for incubating eggs and other have built their own.
After your cultures have been grown and your identification made, you
must sterilize all media and glassware used in your work. For this purpose
you will need a sterilizer which is often called an autoclave.
AUTOMATIC AUTOCLAVES/DRYERS
This is a fancy pressure cooker! It must be capable of maintains 15
pounds of pressure at 121 C over varying periods of time depending on what
organisms you may have present. The normal times involved vary between 15
minutes and 2 hours. This autoclave must also have a steam escape valve
for safety reasons. The least expensive one I have found is available at a
cost of approximately $350.
In the case of both Salmonella and E Coli, once identification is made,
sensitivity testing is necessary to determine what antibiotics are
effective against the particular organism you have. The required equipment
comes in a kit with instructions for its use. This usually involves
flooding a plate of media with a pure culture and depositing discs soaked
in various antibiotics on the surface of the media. After incubation you
can read the growth reaction of the organisms to the antibiotic.
To summarize the various pieces of equipment necessary for this form of
bacterial testing you will need:
- pure culture
- inoculating needle and loop
- selective media
- differential media
- bacterial table (found in many lab manuals and Microbiology books)
- incubator
- autoclave
- antibiotic sensitivity testing kit
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