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Microbiology Concept Inventory
Please note, you must be an educator in higher ed or maybe high school to qualify to recieve the MCI
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|Preliminary identification of bacteria is usually based upon their cell morphology and grouping and the manner in which they react to certain staining procedures. The purpose of this section is to demonstrate some common staining reactions used to categorize microorganisms.
Figure 3.2. An unstained bacterial smear. Unstained bacteria are mostly made of water and are nearly transparent when viewed through a light microscope (pictured on the left). Note that most of the microbes are not visible, but a dust spec in the center of the field of view is visible. Stains cling to the positive and negative charges of bacteria, but do not bind as readily to the background of a slide. They therefore differentiate microbes from their surroundings. Stained bacteria are shown at 40X and 100X in the center and right panels.
Unstained bacteria are practically transparent when viewed using the light microscope and thus are difficult to see as shown in Figure 3-2. The development of dyes to stain microorganisms was a significant advance in microbiology. Stains serve several purposes:
Stains differentiate microorganisms from their surrounding environment
They allow detailed observation of microbial structures at high magnification
Certain staining protocols can help to differentiate between different types of microorganisms.
Most dyes consist of two functional chemical groups as shown in Figure 3-3. The chromophore group, which give dyes their characteristic color; and the auxochrome group, containing an ionizable chemical structure, which helps to solubilize the dye and facilitates binding to different parts of microorganisms. Previously, dyes were classified as acidic or basic, depending upon whether the pigment was negatively or positively charged at neutral pH. More accurately, dyes can be referred to as anionic (-) or cationic (+) and this is the convention that will be used in this manual. Cationic dyes (crystal violet, methylene blue) will react with groups on bacteria that have a negative charge. Anionic dyes (eosin, nigrosine) will react with groups that have a positive charge. Since most bacteria have many positive and negative groups in their cell walls and other surfaces, they will react with both cationic and anionic dyes.
Figure 3.3. The structure of crystal violet. The auxochrome groups of crystal violet is the charged carbon in the center of the molecule. This is typically neutralized by a Cl- ion. The chromophore group consists of the three benzene rings and the central carbon. These structures readily absorb light.
Staining protocols can be divided into 3 basic types, simple, differential, and specialized. Simple stains react uniformly with all microorganisms and only distinguish the organisms from their surroundings. Differential stains discriminate between various bacteria, depending upon the chemical or physical composition of the microorganism. The Gram stain is an example of a differential stain. Specialized stains detect specific structures of cells such as flagella and endospores.