Copyright © 1989 by Nan Fielding
former NCS Microbiology Consultant

  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