Microbiology

Microbiology Study Guide

Pure Cultures In order to work withmicroorganisms in the laboratory, it is desirable to obtain them in pure cultures. Pure cultures of bacteria can be obtained by spreading out bacteria and permitting the individual cells to form masses of growth called colonies. One can then pick a sample from the colony and be assured that it contains only one kind of bacteria. Cultivating these bacteria on a separate medium will yield a pure culture. To obtain separated colonies from a mixed culture, various isolation methods can be used. One is the streak plate method, in which a sample of mixed bacteria is streaked several times along one edge of a Petri dish containing a medium, such as nutrient agar. A loop is flamed and then touched to the first area to retrieve a sample of bacteria. This sample is then streaked several times in the second area of the medium. The loop is then re-flamed, touched to the second area, and streaked once again in the third area. The process can be repeated in a fourth and fifth area if desired. During incubation, the bacteria will multiply rapidly and form colonies. A second isolation method is the pour plate method . In this method, a sample of bacteria is diluted in several tubes of melted medium, such as nutrient agar. After dilution, the melted agar is poured into separate Petri dishes and allowed to harden. Since the bacteria have been diluted in the various tubes, the plates will contain various dilutions of bacteria, and where the bacteria are most diluted, they will form isolated colonies. Dynamics of Populations Reproduction Patterns Bacteria reproduce by the asexual process of binary fission. In this process, the chromosomal DNA duplicates, after which the bacterial membrane and cell wall grow inward to meet one another and divide the cell in two. The two cells separate and the process is complete. A few bacteria reproduce by budding; they form a small initial outgrowth (bud) that grows until adult size and then separates. One of the remarkable attributes of bacteria is the relatively short generation time, the time required for a microbial population to double in numbers. The generation time varies among bacteria and often ranges between 30 minutes and three hours. Certain bacteria have very brief generation times. Escherichia coli , for example, has a generation time of about 20 minutes when it is dividing under optimal conditions. The Growth Curve The growth of a bacterial population can be expressed in various phases of a growth curve . The logarithms of the actual numbers in the population are plotted in the growth curve along the side axis, and the time is plotted at the base. Four phases of growth are recognized in the growth curve. In the first phase, called the lag phase, the population remains at the same number as the bacteria become accustomed to their new environment. Metabolic activity is taking place and new cells are being produced to offset those that are dying. In the second phase, known as exponential growth phase, or logarithmic phase or log phase, bacterial growth occurs at its optimal level and the population doubles

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