Genetic barcodes

A more recently established approach to taxonomy seeks to identify species solely on the basis of a genetic barcode (also known as a DNA barcode) consisting of one or a few DNA sequences. For example, a 648 bp region of the mitchondrial cyctochrome c oxidase I gene (COI) is currently being developed as a barcode identifier in animals. In a comparison of 260 bird species, this gene region was found to be species-specific, and was also an average of 18 times more variable between species (7.05--7.93 per cent) than within species (0.27-0.43 per cent) (Figure 7.3; Hebert et al., 2004b). This is one of the findings that led to an international collaboration known as the Consortium for the Barcoding of Life that is currently hosted by the Smithsonian's National Museum of Natural History in Washington, DC, and is promoting the eventual acquisition of genetic barcodes for all living species.

The use of genetic barcodes to identify species has two general applications: the identification of previously characterized species from a comparison of documented DNA sequences, and the discovery of new species on the basis of

Percentage sequence divergence

Figure 7.3 The extent to which the mitochondrial cytochrome oxidase I gene varies among 260 species of North American birds. Comparisons are based on levels of sequence divergence within and among species, genera and families. Data from Hebert et al. (2004b)

Percentage sequence divergence

Figure 7.3 The extent to which the mitochondrial cytochrome oxidase I gene varies among 260 species of North American birds. Comparisons are based on levels of sequence divergence within and among species, genera and families. Data from Hebert et al. (2004b)

novel DNA sequences. The former application is not particularly controversial and, as we saw in the previous chapter, the practice of identifying species or samples by matching up sequences is becoming increasingly widespread. Nevertheless, this approach does assume that sequences are species-specific and we know from Chapter 5 that both hybridization and incomplete lineage sorting mean that this will not always be the case. Because hybridization occurs between species within all major taxonomic groups, and an estimated one-quarter of all animal species have yet to reach the stage of reciprocal monophyly (Funk and Omland, 2003), DNA sequences sometimes will transcend the boundaries of putative species.

The second application of DNA barcodes, which is the identification of new species, is more controversial. This is partly because the range of intraspecific sequence divergence can be difficult to predict. Although Hebert et al. (2004b) found that avian intraspecific divergence was consistently <0.44 per cent and therefore lower than interspecific divergence, a study by Johnson and Cicero (2004) found that interspecific sequence divergences were 0--8.2 per cent in 39 comparisons of avian sister species. Inconsistencies such as these may be the exception rather than the rule, although data from a wider range of taxonomic groups are needed before we can reach this conclusion.

Before such data can be acquired, appropriate genetic regions first must be identified in these other taxonomic groups. Microbes, for example, transfer genes between putative species so often that sequence data from an estimated 6-- 9 genes will be required before closely allied species can be differentiated (Unwin and Maiden, 2003). In plants, hybridization and polyploidy can obscure evolutionary relationships, although proponents of genetic barcodes hope that a region of the chloroplast genome can be found that will reliably distinguish species. They also suggest that COI will be useful for identifying a number of protistan species, although anaerobic species lack mitochondria and therefore will require a different marker. In the meantime, DNA barcodes are becoming an increasingly acceptable tool for identifying species and may well become more widespread in the literature over the next few years (see also Box 7.1).

Box 7.1 Defining species from molecular and ecological data

Some of the potential problems associated with molecular taxonomy, such as incomplete lineage sorting or low sequence divergence between closely related species, sometimes can be overcome if molecular data are combined with ecological studies. The value of this combined approach was illustrated by a recent taxonomic re-evaluation of the neotropical skipper butterfly Astraptes fulgerator. For many years this was described as a single, variable, wide-ranging species that occurred in a variety of habitats distributed between the far southern USA and northern Argentina. The ecology of this species has been studied intensively throughout a long-term project in which the colour patterns and feeding preferences of >2500 wild-caught caterpillars were monitored. Once these had developed into adults, researchers recorded the sizes of the butterflies and their wing shapes, colours and patterns. Overall morphological similarity is high throughout the range because of recent shared ancestry, and also because selection has maintained mimicry of warning colouration against predators. Nevertheless, although morphological differences were subtle, the ecological data suggested that A. fulgerator was in fact a complex of at least six or seven species (Hebert et al., 2004a, and references therein).

As a recent addition to the barcoding project, cytochrome oxidase I sequences were obtained from 465 A. fulgerator individuals. Morphological characters of caterpillars and adults, plus the identity of their food plants, were superimposed onto a neighbour-joining tree that was reconstructed from the COI sequence data. One group was paraphyletic and pseudogenes (nuclear copies of mitochondrial genes; Chapter 2) were amplified from several individuals, but for the most part the combined genetic and ecological data revealed ten distinct clusters suggesting that A. fulgerator is in fact a complex of at least ten distinct species. The sequence divergence between these ten species ranged from 0.32 to 6.58 per cent (Hebert et al., 2004a).

Species are unlikely to be distinguished solely on the basis of sequence divergences as low as 0.32 %, which is why a combination of molecular and ecological data was necessary in this case before realistic species designations could be made. Although the initial investigations were lengthy, the authors suggest that future studies on Astraptes spp. can use the COI barcode as the sole identification tool, thereby bypassing the need for the relatively time-consuming acquisition of ecological and morphological data. In an ideal world, all species would be characterized on the basis of such comprehensive phenotypic and genotypic data, although in many cases this option will be logistically impossible.

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