Over the last two decades, the use of DNA barcodes has transformed our ability to identify and assess life on our planet. Both strengths and weaknesses of the method have been exemplified through thousands of peer-reviewed scientific articles. In the light of novel sequencing approaches, currently capable of generating millions of reads at low cost, we reflect on the questions: what will the future bring for DNA barcoding? Will identification of species using short, standardized fragments of DNA stand the test of time? We present reflected opinions of early career biodiversity researchers in the form of a SWOT-analysis and discuss answers to these questions.

Aim The Neotropics, particularly South America, holds unparalleled high levels of species richness, when compared to other major biomes. Some neotropical areas are hotspots of a fragmentary known diversity of insects and are under manifest danger of biodiversity loss and climate change. Therefore, prompt estimates methods of its diversity are urgently required to complement slower traditional taxonomic approaches. Despite a variety of algorithms for delimiting species through single-locus DNA barcodes having been developed and applied for rapid estimates of species diversity in a wide array of taxa; however, tree-based and distance-based methods may lead to different group assignments, either overestimating or underestimating the number of putative species. Here, we investigate the performance of different DNA-based species delimitation approaches for a rapid biodiversity estimate of the diversity of Polypedilum (Chironomidae, Diptera) in South America. Location Worldwide Methods We analyze a mtDNA dataset comprising 1,492 specimens from 598 locations worldwide. Molecular operational taxonomic units (MOTUs) ranged from 267 to 520, based on the Barcode Index Number (BIN), Bayesian Poisson tree processes (bPTP), multi-rate Poisson tree processes (mPTP), single-rate Poisson tree processes (sPTP), and generalized mixed Yule coalescent (sGMYC) approaches. Results Our results highlight Polypedilum as a species-rich genus, yet incompletely documented, and found the sGMYC method to be the most adequate to estimate putative species in our dataset. Furthermore, based on these data, we describe the distribution of diversity and some biogeographical patterns of Polypedilum. Main Conclusions Findings imply the genus exhibited high levels of endemism and richness of species in the Neotropics, which confirmed our hypothesis that there are substantial differences in community structure between the Polypedilum fauna in South America and the neighboring regions.

Biodiversity studies greatly benefit from molecular tools, such as DNA metabarcoding, which provides an effective identification tool in biomonitoring and conservation programmes. The accuracy of species-level assignment, and consequent taxonomic coverage, relies on comprehensive DNA barcode reference libraries. The role of these libraries is to support species identification, but accidental errors in the generation of the barcodes may compromise their accuracy. Here we present an R-based application, BAGS (Barcode, Audit & Grade System), that performs automated auditing and annotation of cytochrome c oxidase subunit I (COI) sequences libraries, for a given taxonomic group of animals, available in the Barcode of Life Data System (BOLD). This is followed by implementing a qualitative ranking system that assigns one of five grades (A to E) to each species in the reference library, according to the attributes of the data and congruency of species names with sequences clustered in Barcode Index Numbers (BINs). Our ultimate goal is to allow researchers to obtain the most useful and reliable data, highlighting and segregating records according to their congruency. Different tests were performed to perceive its usefulness and limitations. BAGS fulfils a significant gap in the current landscape of DNA barcoding research tools by quickly screening reference libraries to gauge the congruence status of data and facilitate the triage of ambiguous data for posterior review. Thereby, BAGS have the potential to become a valuable addition in forthcoming DNA metabarcoding studies, in the long term contributing to globally improve the quality and reliability of the public reference libraries.