LABORATORY CULTIVATION AND ISOLATION OF BACTERIA

Diagnostic bacteriology is concerned with the isolation and identification of bacteria in a specimen from a patient. These specimens, unless from a normally sterile site of the body, rarely contain a single bacterial type, but are mixtures of the disease-producing bacteria and the host's normal or indigenous flora. Since accurate studies of the biochemical and the antigenic properties of a bacterial species are possible only through the use of pure cultures, it is necessary to have a reliable and rapid method that will permit the isolation of possible pathogenic organisms.

An inoculum from the specimen is streaked on solid agar in a manner, which physically separates most of the bacterial types, permitting them to form discrete colonies. This procedure is facilitated whenever possible by the use of either a selective medium that inhibits the growth of species not sought or by the use of a differential medium, which imparts a recognizable appearance to the colonies of the type sought. It is possible that colonies of the bacterial type selected for by the selective medium will be contaminated with bacteria of a different type that are inhibited from growing but not killed by the selective medium. Upon transfer of this mixed colony to a medium without the inhibitors, both types of bacteria may grow, and a pure culture will not be obtained. Consequently, it is often necessary to streak a second plate of the same selective medium with a colony from the first selective plate in order to obtain a pure culture of the bacterial species that you are attempting to isolate.

The method used most often for colony isolation from a clinical specimen or mixed culture utilizes the four-phase streaking pattern described on the following page.