Natural enemies in the last Central European population of the Danube Clouded Yellow butterfly ( <i>Colias myrmidone</i> )

The Danube Clouded Yellow (Colias myrmidone) is one of the most endangered butterfly species in Europe. Despite growing knowledge about habitat requirements and major threats, little is known about its natural enemies, particularly parasitoids. To fill this gap, we undertook field studies on the last Central European C. myrmidone population which inhabits the Knyszyn Forest (north-eastern Poland). Reared parasitoids were identified based on morphological features and were further DNA-barcoded. At the adult stage, we documented only four cases of predation, all caused by spiders: Araneus quadratus, Argiope bruennichi and Misumena vatia. Egg mortality was primarily due to parasitism by Trichogramma evanescens (Trichogrammatidae), a common parasitoid of Lepidoptera, which is also used for pest control. Surprisingly, larval parasitism was rare, with only a few individuals affected by Cotesia ancilla (Braconidae: Microgastrinae), a regular parasitoid of Colias species. Cases of pseudohyperparasitism by Lysibia nanus (Ichneumonidae: Phygadeuontinae) were recorded. Pupal mortality was considerably higher, and the most common was the solitary parasitoid Brachymeria femorata (Chalcididae: Chalcidinae). Additionally, we reared the gregarious Blapsidotes vicinus (Ichneumonidae: Phygadeuontinae). Both species are widespread and regular parasitoids of exposed butterfly pupae. Furthermore, we obtained Hoplocryptus murarius (Ichneumonidae: Cryptinae) from two chrysalises, which was unexpected due to its usual association with Auplopus carbonarius (Hymenoptera: Pompilidae). Aside from pupal parasitism, we recorded a single case of pupal predation by a Polistes wasp (Vespidae: Polistinae). These findings provide crucial insights for the conservation of C. myrmidone, highlighting potential risks posed by natural enemies, including those manipulated against crop pests.