Growth of Microbes in Pure Culture

Unlike Pasteur, who was a university professor, Koch took up a medical practice in a small Polish-German town. To make space in his home for a laboratory to study anthrax and other deadly diseases, his wife curtained off part of his patients' examining room. Anthrax interested Koch because its epidemics in sheep and cattle caused economic hardship among local farmers. Today, anthrax is no longer a major problem for agriculture, as its transmission is prevented by effective environmental controls and vaccination. It has, however, gained notoriety as a bioterror agent because anthrax bacteria can survive for long periods in the dormant, desiccated form of an endospore. In 2001, anthrax spores sent through the mall contaminated post offices throughout the northeastern United States, as well as an office building of the United States Senate, causing several deaths. To investigate whether anthrax was a transmissible disease, Koch used blood from an anthrax-infected carcass to inoculate a rabbit. When the rabbit died, he used the rabbit's blood to inoculate a second rabbit, which then died in turn. The blood of the unfortunate animal had turned black with long, rod-shaped bacilli. Upon introduction of these bacilli into healthy animals, the animals became ill with anthrax. Thus, Koch demonstrated an important principle of epidemiology: the chain of infection, or transmission of a disease. In retrospect, his choke of anthrax was fortunate, for the microbes generate disease very quickly, multiply in the blood to an extraordinary concentration, and remain infective outside the body for long periods.
        Koch and his colleagues then applied their experimental logic and culture methods to a more challenging disease: tuberculosis. In Koch's day, tuberculosis caused one-seventh of reported deaths in Europe; today, tuberculosis bacteria continue to infect millions of people worldwide. Koch's approach to anthrax, however, was less applicable to tuberculosis, a disease that develops slowly after many years of dormancy. Furthermore, the causative bacteria, Mycobacterium tuberculosis, are small and difficult to distinguish from human tissue or from different bacteria of similar appearance associated with the human body. How could Koch prove that a particular bacterium caused a particular disease? What was needed was to isolate a pure culture of microorganisms, a culture grown from a single "parental" cell. This had been done by previous researchers using the laborious process of serial dilution of suspended bacteria until a culture tube contained only a single cell. Alternatively, inoculation of a solid surface such as a sliced potato could produce isolated colonies, distinct populations of bacteria, each grown from a single cell. For M. tuberculosis, Koch inoculated serum, which then formed a solid gel after heating. Later he refined the solid-substrate technique by adding gelatin to a defined liquid medium, which could then be chilled to form a solid medium in a glass dish. A covered version called the petri dish (also called a petri plate) was invented by a colleague, Julius Richard Petri (1852-1921). The petri dish consists of a round dish with vertical walls covered by an inverted dish of slightly larger diameter. Today, the petri dish, generally made of disposable plastic, remains an indispensable part of the microbiological laboratory. Another improvement in solid-substrate culture was the replacement of gelatin with materials that remain solid at higher temperatures, such as the gelling agent agar (a polymer of the sugar galactose). The use of agar was recommended by Angelina Hesse (1850-1934), a microscopic and illustrator, to her husband, Walther Hesse (1846-1911), a young medical colleague of Koch (Fig.1.16). Agar comes from red algae (seaweed), which is used by East Indian birds to build nests; it is the main ingredient in the delicacy "bird's nest soup." Dutch colonists used agar to make jellies and preserves, and a Dutch colonist from Java introduced it to Angelina Hesse. The Hesses used agar to develop the first effective growth medium for tuberculosis bacteria. (Pure culture is discussed further in Chapter 4.) Note that some kinds of microbes cannot be grown in pure culture without other organisms. For example, viruses can be cultured only in the presence of their host cells. The discovery of viruses is explored at the end of this section