Capsules are protective structures with various functions, including but not limited to adherence to surfaces and other bacteria, protection from desiccation, and protection from phagocytosis. Some bacteria produce an extracellular viscous outer layer called a capsule (3, 5). A Gram stain of a mixture of Staphylococcus aureus (Gram-positive purple cocci) and Escherichia coli (Gram-negative red rods). Also, shown are two typical bacterial cell shapes: the bacilli (or rods) and the cocci (or spheres).įigure 3: Gram Staining Results. The right column shows how Gram-positive bacteria react. The left column shows how Gram-negative bacteria react at each step of the protocol. Gram staining is used to differentiate cells into two types (Gram-positive and Gram-negative) and is also useful to distinguish cell shape (spheres or cocci, rods, curved rods, and spirals) and arrangement (single cells, pairs, chains, groups, and clusters) (1, 3).įigure 2: Schematic of the Gram Staining Protocol. Gram-positive bacteria stain purple due to the thick peptidoglycan layer which is not easily penetrated by the decolorizer Gram-negative bacteria, with their thinner peptidoglycan layer and higher lipid content, destain with the decolorizer and are counterstained red when Safranin is added (Figure 3). In the final step, a counterstain, Safranin, is added to color cells red (Figure 2). The slide is flushed with an Iodine solution, which fixes the Crystal Violet to the cell wall, followed by a decolorizer (an alcohol) to wash away any non-fixed Crystal Violet. The heat-fixed sample is stained with Crystal Violet, turning the cells purple.
Briefly, a bacterial smear is placed on a microscope slide and then heat-fixed to adhere the cells to the slide and make them more readily accepting of stains (1). The Gram stain, developed in 1884 by the Danish bacteriologist Hans Christian Gram (1), differentiates bacteria based on the composition of the cell wall (1, 2, 3, 4). The most important parts of the microscope are labeled. Here, we will discuss visualizing bacteria with Gram stains, Capsule stains, and Endospore stains.įigure 1: A typical compound microscope.
Typically, compound microscopes have multiple objective lenses of varying powers to allow for different magnification (1, 2). Combining the objective lens and eyepiece allows for higher magnification than using a single lens alone. This is then magnified by the eyepiece (ocular lens) which enlarges the image. Compound microscopes have an objective lens close to the object which collects light to create an image of the object. Simple microscopes (for example a magnifying glass) have only one lens to magnify an object, while compound microscopes have several lenses to enhance magnification (Figure 1). The main difference between them is the number of lenses used to magnify the object. The two main types of light microscopes are simple and compound. Staining bacteria is necessary when distinguishing bacterial types with light microscopy. To observe these properties, one can use light microscopy however, some bacterial characteristics (for example size, lack of coloration, and refractive properties) make it hard to distinguish bacteria solely with a light microscope (1, 2). Many properties help distinguish them from each other, including but not limited to Gram-staining type, shape and arrangement, production of capsule, and formation of spores. DiRita 1ġ Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan, United States of Americaīacteria are diverse microorganisms found nearly everywhere on Earth.
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