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Streptomyces are gram positive, spore-forming bacteria found in soil. They are characterized by their tough, leathery, frequently pigmented colonies and their filamentous growth. When first discovered, these organisms were thought to be fungi, but closer examination revealed a lack of a nuclear membrane and the presence of peptidoglycan, demonstrating their prokaryotic origin. Streptomyces are chemoheteroorganotrophs, growing best at 25°C and pH 8-9. They are capable of using complex organic materials as carbon and energy sources and are involved in the breakdown of these products in the soil. This degradative ability makes these bacteria pivotal in the production of fertile soil for agriculture. They also give soil its characteristic smell by the production of a class of volatile low molecular weight compounds called geosmins.
In the laboratory, different isolates were found to produce numerous compounds capable of inhibiting or killing other microorganisms. Many of these Figure 10-3 shows some examples of streptomycetes
Figure 10.3. An example of streptomycetes. A photomicrograph of Streptomyces vegetative cells and spores.
In this section, we describe the enrichment of this group of organisms and test several isolates for their antibiotic producing ability. Dried soil, a great source of inoculum for Streptomyces, will be plated directly onto Actinomycete Isolation Agar (AIA). Since an enrichment step is not necessary, what does that tell you about the population of Streptomyces in the soil? AIA contains starch and sodium caseinate as sole carbon and energy sources. Only organisms capable of degrading these complex polymers (mostly molds and streptomycetes) are able to grow. Sodium propionate (a preservative sometimes present in commercial bread) is added to inhibit the growth of molds. Incubation at 30°C for more than five days results in the formation of Streptomyces colonies that are recognized by their characteristic appearance.
3 0.85% NaCl dilution blanks (9 ml)
3 plates Actinomycete Isolation Agar (AIA)
Below is shown the appearance of a Streptomyces isolation plate after 5 days of incubation
5 AIA plates
Figure 10.4. AIA after incubation. This AIA plate was inoculated with soil from near Hayward Wisconsin. The plate was incubated at 30Â°C for one week. Streptomyces species are the white and colorful chalky looking colonies.
Figure 10.5. A closeup of a typical streptomyces colony. Note the colorful chalky/dusty appearance. The colony is hard, not gummy, and does not easily lift from the agar.
Figure 10.6. A wet mount of a Streptomyces colony. A Streptomyces colony under the phase microscope. Notice the long branched hyphae in the picture. This picture was taken at 1000x and enlarged 2 fold.
6 plates of Penassay Agar
Culture of Streptomyces griseus, an active antibiotic producer
Figure 10.7. Streaking pattern for the antibiotic test. Diagram of the appropriate streaking technique for the Penassay plates. (A) The placement of the Streptomyces colonies. (B) The placement of the test cultures. Note that there is a 5-7 day incubation between streak A and streak B.
Figure 10.8. Streaks for isolated colonies on penassay agar. The appearance of penassay agar after streaking for isolated colonies of Actinomycetes.
Broth cultures of 5 test organisms
Figure 10.9. An appropriately grown test plate. Example plates after growth of the Streptomyces test strains. Note the full growth completely along the streak line. There is a minor contaminant in the left sample, but it should not interfere with analysis.
Figure 10.10. Antimicrobial production by isolates. Examples of microbes that have been isolated and their inhibition of test microbes. Isolate 3 is secreting a brown metabolite into the medium. Isolate 4 is secreting an antimicrobial into the medium that is inhibiting S. epidermidis, E. faecalis, and B. subtilis. Ec - E. coli, Pf - P. flourescens, Bs - B. subtilis, Se - S. epidermidis, and Ef - E. faecalis.