Background: Dragonflies and damselflies (Odonata) are important components in biomonitoring due to their amphibiotic lifecycle and specific habitat requirements. They are charismatic and popular insects, but can be challenging to identify despite large size and often distinct coloration, especially the immature stages. DNA-based assessment tools rely on validated DNA barcode reference libraries evaluated in a supraregional context to minimize taxonomic incongruence and identification mismatches.Methods: This study reports on findings from the analysis of the most comprehensive DNA barcode dataset for Central European Odonata to date, with 103 out of 145 recorded European species included and publicly deposited in the Barcode of Life Data System (BOLD). The complete dataset includes 697 specimens (548 adults, 108 larvae) from 274 localities in 16 countries with a geographic emphasis on Central Europe. We used BOLD to generate sequence divergence metrics and to examine the taxonomic composition of the DNA barcode clusters within the dataset and in comparison with all data on BOLD.Results: Over 88% of the species included can be readily identified using their DNA barcodes and the reference dataset provided. Considering the complete European dataset, unambiguous identification is hampered in 12 species due to weak mitochondrial differentiation and partial haplotype sharing. However, considering the known species distributions only two groups of five species possibly co-occur, leading to an unambiguous identification of more than 95% of the analysed Odonata via DNA barcoding in real applications. The cases of small interspecific genetic distances and the observed deep intraspecific variation inCordulia aenea(Linnaeus, 1758) are discussed in detail and the corresponding taxa in the public reference database are highlighted. They should be considered in future applications of DNA barcoding and metabarcoding and represent interesting evolutionary biological questions, which call for in depth analyses of the involved taxa throughout their distribution ranges.

Monitoring of odonates has become an important instrument for ecological status assessment of (semi-)aquatic habitats. Besides information on presence and abundance, knowledge about a species´ autochthony at the surveyed waterbody is a significant information within the assessment process. Here, the finding of exuviae represents the ultimate proof of successful reproduction. Although feasible for most odonate species, morphological identification of exuviae is often time consuming, as it relies on small, fragile structures. To facilitate species identification of exuviae, a DNA barcoding approach was developed, including (1) non-destructive extraction of DNA using whole exuviae or their tracheal tubes, and (2) primer systems for long (< 600 bp) and short (< 200 bp) CO1 fragments. A total of 85 exuviae from 33 species were analysed and compared to results of morphological identification. Additionally, factors potentially influencing DNA quality and quantity, as well as PCR and sequencing success were investigated. Eighty-two exuviae matched the morphologically identified genus, and 60 matched at species level. Of the 33 species present in the data set, 82% could be identified to species level via DNA barcoding. The results show how DNA-based approaches can support fast and accurate species identification and therefore enhance monitoring of an ecologically important taxonomic group, with high relevance for conservation and habitat restoration. Moreover, the use of exuviae as DNA resource once more shows that non-invasive sampling offers great potential for molecular species identification, which is essential when studying rare and endangered species. Implications for insect conservation: Our results show how molecular tools, here DNA barcoding of odonate exuviae, can support species monitoring without the need of catching individuals, harming, or even killing them. Obtaining DNA from non-invasive sources can thus be a direct advantage to the conservation of insects, especially when dealing with rare and endangered species and/or populations. Using the example of odonates as bioindicator organisms for aquatic and semi-aquatic habitats, we highlight the importance of non-invasive genetic approaches for population studies and monitoring of insect species and/or species communities for ecosystem assessments and conservation management.

In the original publication of the article, the Table 3 was published incorrectly. The correct Table 3 is given in this correction.

Dragonflies and damselflies are important indicator species for quality and health of (semi-)aquatic habitats. Hitherto, 78 species of Odonata have been reported for Austria. Ecological data, Red List assessments, and a dragonfly association index exist, but population- and species-level genetic data are largely lacking. In this study, we establish a comprehensive reference DNA barcode library for Austrian dragonflies and damselflies based on the standard barcoding marker COI. Because of the increasing significance of environmental DNA (eDNA) analyses, we also sequenced a segment of the mitochondrial 16S rRNA gene, a marker often used in eDNA metabarcoding approaches. In total, we provide 786 new COI barcode sequences and 867 new 16S sequences for future applications. Sequencing success was >90 percent for both markers. Identification success was similar for both markers and exceeded 90 percent. Difficulties were only encountered in the genera Anax Leach, 1815, Chalcolestes Kennedy, 1920, Coenagrion Kirby, 1890 and Somatochlora Selys, 1871, with low interspecific genetic distances and, consequently, BIN (barcode index number) sharing. In Anax, however, individual sequences clustered together in species-specific groups in the COI tree. Irrespective of these challenges, the results suggest that both markers perform well within most odonate families in terms of sequencing success and species identification and can be used for reliably delimiting Austrian species, monitoring, and eDNA approaches.