Abstract
A DNA barcode is a preferrably short and highly variable region of DNA supposed to facilitate a rapid identification of species. In many protistan lineages, a lack of species-specific morphological characters hampers an identification of species by light or electron microscopy, and difficulties to perform mating experiments in laboratory cultures also do not allow for an identification of biological species. Thus, testing candidate barcode markers as well as establishment of accurately working species identification systems are more challenging than in multicellular organisms. In cryptic species complexes the performance of a potential barcode marker can not be monitored using morphological characters as a feedback, but an inappropriate choice of DNA region may result in artifactual species trees for several reasons. Therefore a priori knowledge of the systematics of a group is required. In addition to identification of known species, methods for an automatic delimitation of species with DNA barcodes have been proposed. The Cryptophyceae provide a mixture of systematically well characterized as well as badly characterized groups and are used in this study to test the suitability of some of the methods for protists. As species identification method the performance of blast in searches against badly to well-sampled reference databases has been tested with COI-5P and 5′-partial LSU rDNA (domains A to D of the nuclear LSU rRNA gene). In addition the performance of two different methods for automatic species delimitation, fixed thresholds of genetic divergence and the general mixed Yule-coalescent model (GMYC), have been examined. The study demonstrates some pitfalls of barcoding methods that have to be taken care of. Also a best-practice approach towards establishing a DNA barcode system in protists is proposed.
Highlights
DNA barcodes are short and highly variable DNA regions that can be used to identify species [1]
Funk and Omland summarized problems in DNA barcoding of animals that will likely cause failures in species identification due to poly- or paraphylies in species that can be roughly assigned to three categories [9]: (a) an artificial systematics combined with an appropriate barcode marker, (b) a consistent systematics, but an inappropriate molecular marker and (c) naturally non-monophyletic taxa
The search strategy is set to ‘‘megablast’’, an algorithm supposed to be used for highly similar sequences, i.e. for intraspecific comparisons with identities equal or higher than 95%
Summary
DNA barcodes are short and highly variable DNA regions that can be used to identify species [1]. In category (b) genes have been either subject to introgression, to incomplete lineage sorting after duplication events or paralogous instead of orthologous gene copies have been used as barcode markers. In these cases systematics may be consistent, but the wrong marker has been chosen for barcoding. Natural non-monophyly may be caused by interspecific hybridization events or by reproductive isolation of a subpopulation from a species. In the latter case, the parent species will be paraphyletic with the monophyletic offspring species being nested inside. Apart from the problems listed by Funk and Omland, molecular traits inherent in a chosen marker may result in artifactual tree topologies or erroneous species identifications, e.g. unequal evolutionary rates across taxa, sites and/or time, heterogenous base compositions or differing codon usage [11,12,13,14,15]
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