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Strain-Testing to Determine Genera
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Cytoplasm is generally transparent, making it difficult to study under a microscope. Various methods of staining microbes are used to differentiate them and draw attention to their distinguishing features. In the case of a patient whose symptoms are consistent with an infection of the Bacillus, Escherichia and Mycoplasma genera, a sputum sample can be analyzed with staining procedures to narrow down possibilities to one genera.
All tests begin by applying a sample to a slide and allowing it to dry. Most tests then require that the culture be passed over a flame to kill the cells and allow them to accept dyes. Some tests don’t require heat fixing due to the damage it causes to fine structures.
The Bacillus, Escherichia and Mycoplasma genera can be distinguished by their distinct features anatomically as well as their reactions to various solutions. The acid-fast test, spore test, flagella test, and the negative strain technique, taken together, will eliminate two of the possibilities and leave one. Mycoplasma can be distinguished from other genera using the acid-fast test. In this process, sputum is applied to a slide and an initial bright red dye is applied to the sample via heat or through a solvent. The dye is then rinsed off with an acid-alcohol solution; Mycoplasma cells retain the red dye after being washed, while Escherichia and Bacillus cells would take on a blue stain.
If Mycoplasma is ruled out, a spore test can be carried out. The presence of spores found with this test would indicate the presence of Bacillus, while no spores would indicate that the final option, Escherichia, is the culprit. The spore test is carried out by applying malachite green along with the application of heat to the sputum, which kills the cells and forces them to accept the dye. Safranin is applied to color...

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