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Finally, the barcoding approach first examines species delineation through COI barcodes for species established generally through a traditional approach of taxonomy using phenotypes. Some of the pairs with overlapping barcodes, however, may be a single species. In the present case, 34 species would have been undetected by using a 1% threshold. Providing that seven species share polymorphism or harbour mixed genealogy, 24 species with monophyletic COI lineages would have been overlooked with a 1% threshold. The present study is the first to assess the resolution of barcoding for freshwater fish species from a variety of primary freshwater groups. 0.4% while the average divergence reached 9.9% between congeneric species. However, the average distance between conspecifics and congeneric species reached 0.3% and 8.3%, respectively, for freshwater fishes in this study, a pattern strikingly similar to that of marine fishes. Although geographic structure was often detected here among populations, the present survey suggests that the higher geographic structure of freshwater fishes is not necessarily reflected in deeper intraspecific and interspecific divergence than marine species.
Although, we failed to capture a substantial amount of population diversity through the present sampling, it remains unlikely that sampling artefacts alone can account for similar intraspecific divergences found among freshwater and marine species. In summary, most of the North American freshwater fish species analysed here exhibit a similar pattern of genetic diversity at COI, each being a single cluster of tightly related mtDNA sequences distinct from all other species. Therefore, the present survey supports the view that the use of COI barcodes is a powerful tool for species identification. Using this method would clearly allow the identification of individually isolated freshwater fish eggs, larvae, fillets and fins, hence providing many news tools useful for the practice of conservation and forensics genetic in these freshwater fishes. From a systematic perspective, COI barcodes provide a new and fast approach for screening the real number of species characterised by private sets of diagnostic characters. The present data set coupled with the functionality in BOLD provides a tool that is already operational for molecular assisted identification of the Canadian species.
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