DNA barcode reference library of bush-crickets (Orthoptera, Tettigoniidae) from the Iberian Peninsula.

Curated DNA barcode reference libraries are crucial for advancing environmental DNA (eDNA) studies, monitoring biological invasions, reliable biodiversity assessments, accurate species identification, etc. However, DNA barcode databases remain highly incomplete for most invertebrate taxa. In this study, we present the most comprehensive reference library to date for the family Tettigoniidae (Orthoptera) from the Iberian Peninsula-the most species-rich orthopteran family globally, with over 8,000 valid species. We generated 402 new DNA barcodes from at least 121 tettigoniid species from the Iberian Peninsula and integrated these with 169 previously published sequences. The resulting dataset comprises 571 barcoded specimens, representing 49 genera and 123 species, including many recently described taxa. Notably, we provide DNA barcodes for at least 68 described species that previously lacked them. Our dataset covers 85% of the tettigoniid species in the Iberian Peninsula and approximately 25% of European bush-cricket species. Furthermore, our analyses show that most tettigoniid species (95%) can be reliably identified using DNA barcoding. However, mitochondrial introgression events were detected in several species of the subfamilies Bradyporinae and Tettigoniinae, highlighting the need for cautious application of this molecular identification tool.