DNA barcoding technologies have provided a powerful tool for the fields of ecology and systematics. Here, we present a part of the InBIO Barcoding Initiative Database: contribution to the knowledge on DNA barcodes of cuckoo wasps (Hymenoptera, Chrysididae) dataset representing 144 specimens and 103 species, covering approximately 44% of the Iberian and 21% of the European fauna. The InBIO Barcoding Initiative (IBI – DNA Barcoding Portuguese terrestrial invertebrate biodiversity) aims to fill the barcoding gap for the terrestrial invertebrate taxa. All DNA extractions are deposited in the IBI collection at CIBIO, Research Center in Biodiversity and Genetic Resources and specimens are deposited in the University of Mons collection (Belgium) and in the Natur-Museum in Lucerne (Switzerland).This dataset increases the knowledge on the DNA barcodes and distribution of 102 species of cuckoo wasps. A total of 52 species, from 11 different genera, were new additions to the Barcode of Life Data System (BOLD), with DNA barcodes for another 44 species added from under-represented taxa in BOLD. All specimens have their DNA barcodes publicly accessible through the BOLD online database. Nine cuckoo wasp species are newly recorded for Portugal. Additionally, two new species for science are described:Chrysis crossiRosa, sp. nov. from southern Portugal andHedychridium calcariumRosa, sp. nov. from eastern Spain. Several taxonomic changes are proposed andHedychrum rutilansDahlbom, 1845 is found to consist of two different taxa that can be found in sympatry,Hedychrum rutilanss. str. andHedychrum viridaureumTournier, 1877 stat. nov.Stilbum westermanniDahlbom, 1845 stat. nov. is confirmed as distinct fromStilbum calens(Fabricius, 1781), with the latter species not confirmed as present in Iberia; barcodedStilbummaterial from Australia is distinct and representsStilbum amethystium(Fabricius, 1775) sp. resurr.; Portuguese material identified asHedychridium chloropygumBuysson, 1888 actually belongs toHedychridium caputaureumTrautmann & Trautmann, 1919, the first confirmed record of this species from Iberia.Philoctetes parvulus(Dahlbom, 1845) is confirmed to be a synonym ofPhiloctetes punctulatus(Dahlbom, 1845).Chrysis lusitanicaBischoff, 1910 is confirmed as a valid species.Chrysis hebraeicaLinsenmaier, 1959 stat. nov. is raised to species status.