Classification of Bacteria

Identification By Phenotypic Characteristics

Historically, bacterial classification has been based on phenotypic characteristics. A useful first approach to classification was developed in 1884 by Christian Gram, who observed that some bacteria retain crystal violet dye after decolorization with ethanol and others do not. Using what is now called Gram staining, bacteria that retain crystal violet remain blue and are called gram positive , whereas bacteria that are decolorized appear red and are called gram negative . These staining characteristics reflect differences in cell wall structure. As shown in Fig. 114.1 , gram-positive organisms have a relatively simple cell wall composed of an inner membrane and a surrounding peptidoglycan layer between 30 and 200 molecules thick. The gram-positive cell wall also contains teichoic acids and lipoteichoic acids, which are water-soluble polymers of polyol phosphates. Gram-negative organisms have a more complex cell wall, characterized by the presence of an inner membrane, a surrounding peptidoglycan layer one to two molecules thick, and an outer membrane ( Fig. 114.1 ). The outer membrane contains lipopolysaccharide (endotoxin), which is unique to gram-negative organisms. The compartment between the inner and outer membranes is called the periplasm and contains a number of degradative enzymes and transport-related proteins.

Whereas most bacteria are gram-positive or gram-negative, some stain poorly or fail to stain at all with Gram reagents. Examples are mycobacteria, some actinomycetes, treponemes, rickettsiae, anaplasmae, chlamydiae, and mycoplasmas. Mycobacteria possess a complex cell wall that is rich in lipids and have been referred to as acid fast because of their resistance to decolorization with acid solutions. These organisms are best visualized microscopically with Ziehl-Neelsen or Kinyoun acid-fast stain. Most actinomycetes are gram positive, but Nocardia and Rhodococcus characteristically take up Gram stain irregularly. They have a cell wall structure similar to that of mycobacteria, and, like the mycobacteria, they are acid fast.

The family Treponemataceae contains five genera, namely Treponema, Borrelia, Leptospira, Brachyspira, and Spirillum. The treponemes generally do not stain with Gram reagents or other standard laboratory stains and usually are best seen with darkfield microscopy. Members of the family Rickettsiaceae and the family Anaplasmataceae ( Ehrlichia, Anaplasma, Wolbachia, and Neorickettsia ) possess a cell wall typical of gram-negative bacilli but react weakly with Gram stain and are visualized optimally with Giemsa or Gimenez stain. Members of the family Chlamydiaceae ( Chlamydia trachomatis, Chlamydophila psittaci, and Chlamydophila pneumoniae ) are obligate intracellular organisms that possess inner and outer membranes similar to those of gram-negative bacteria, but these organisms lack a peptidoglycan layer and do not take up Gram stain. The two important members of the family Mycoplasmataceae are Mycoplasma and Ureaplasma. Mycoplasmas lack a cell wall and enclose their cytoplasmic contents only by a well-developed plasma membrane.

Cellular morphology also plays an important role in classification. Bacteria can be separated into five major groups on the basis of morphology as viewed through the light microscope. Bacteria that are spherical or oval in appearance are described as cocci. Those that are rod-like or cylindrical are referred to as bacilli. Those with a comma-like or curved rod appearance are referred to as vibrios . Bacteria with a helical or more undulating appearance are called spirochetes if they are flexible or spirilla if they are rigid.

After cell division, the cell wall between daughter cells in some species may not separate completely, giving rise to various cell arrangements. These cell arrangements are typically distinctive and may be helpful in identifying related organisms. Common arrangements are cocci in pairs, cocci in chains, cocci in irregular clusters, cocci in packets of four or eight (called sarcina ), bacilli in pairs, bacilli in chains, unusually short bacilli (called coccobacilli ), and bacilli with tapered ends (called fusiform bacilli ).

A wide range of other phenotypic characteristics are also employed for the purpose of bacterial classification. For example, aerobic and facultative anaerobic organisms are distinguished from strictly anaerobic organisms, and spore-forming bacteria are distinguished from non–spore-forming species. Additional useful properties are carbohydrate fermentation ability, susceptibility to antibiotics and inorganic compounds, and reactivity with well-defined serologic reagents.