Molecular Taxonomy

Each microbial species is presently characterized as having a specific percent molar content of guanosine (G) + cytosine (C) in its DNA. The applicable equation is: 100 mols % = mols % G + mols % C + mols % T + mols % A. Because each guanine nucleotide on one strand of DNA is hydrogen bonded to a cytosine nucleotide on the opposite strand, and because each thymidine nucleotide is hydrogen bonded to a cytosine nucleotide on the opposite strand, the mols % guanine is always equal to the mols % C and the mols % thymine is always equal to the mols % cytosine. By convention, each organism is then defined on the basis its mols % G + C content or GC ratio; 100 mols % = (mols% G + C) + (mols% A + T) or mols % GC = 100 mols % - (mols % A + T). All strains of S. cerevisiae are defined as having a molar GC content of 39%. Any yeast strain that deviates significantly from this value cannot be considered S. cerevisiae. With unknown isolates, the value of the molar GC content is primarily exclusionary. An unknown organism with a molar GC content of 50% is clearly not S. cerevisiae. An unknown isolate with a molar GC content of 39% may be S. cerevisiae, but a molar GC content of 39% is not conclusive evidence of identity. Because all biological species fall within the range of about 10% to 90% GC, numerous unrelated species have the same numerical value for their molar GC DNA content. For example, micrococci and all mammals and fish have a molar GC content of 45%.

The practical definition of a species is that it consists of a collection of strains that share many features in common and that differ considerably from other strains (5). A species is presently defined as encompassing strains with approximately 70% or greater DNA-DNA similarity based on DNA strand hybridization, and with 5% or less Tm (Thermal denaturation temperature) for the hybridized strands (8). Phenotypic characteristics should agree with this definition. This corresponds to a 16S ribosomal ribonucleic acid (rRNA) similarity of 98% or higher (9). The nucleotide sequences of rRNA are far more conserved than DNA among various taxonomic groups. rRNA is capable of hybridizing with DNA; however, RNA-DNA hybridization is far less discriminating in terms of recognizing differences between strains of the same species. It is, however, of utility in discerning the difference between two different species of the same genus. Stated more succinctly, DNA-DNA hybridization experiments are used to detect similarities between closely related organisms, whereas RNA-DNA hybridization experiments are used to detect similarities between more distantly related organisms (10). rRNA sequence data is considered more appropriate for determining inter- and intrageneric relationships than for confirming the species identity of an isolate (11). Several groups of organisms have been found to share almost identical 16S rRNA sequences but a DNA-DNA hybridization significantly lower than 70%, indicating that they represent different species (12).

The early classification of microorganisms was based on the utility of their recognition and identification. What emerged with bacteria, however, was a dual system of classification, one based on metabolism and the other on morphology, which are still with us. In 1910, Orla-Jensen proposed that all lactic-acid-producing bacteria (cocci and rods) be housed in the family Lactobacteriacea. In contrast, the family Micrococcaceae houses the various genera of spherical cells or cocci. The bacterial phylogeny that has emerged from molecular sequence data has little in common with these early concepts regarding the morphological relationships of microbial groups (13). Morphology is no longer the guiding principle regarding phylogenetic relationships, in that most characteristics of bacterial morphology are presently regarded as too simple not to have evolved independently in unrelated organisms (13). However, the concept of morphology as a utilitarian character of an organism can be of great value, if, for example, one suspects that a pure culture of a coccus is contaminated and finds rods present.

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