Red deer (Cervus elaphus) antler myiasis caused by Prochyliza nigrimanus (Meigen 1826).

Antlers are bony structures that undergo regular annual growth, mineralisation and casting phases, representing only mammalian organs capable of full regeneration. Myiasis is infestation of live vertebrates with dipterous larvae. We sampled mineralised antlers from a red deer spiker stag 2 months after velvet shedding, divided it into three segments and fixed in 10% neutral buffered formalin. After demineralisation, samples were embedded in paraffin and sliced to a thickness of 6 µm. Tissue sections were stained with hematoxylin-eosin (HE), a modified staining to show the ossification process, toluidin blue and Masson's trichrome staining. Smears of liquid content from the antlers were made and stained with May-Grünwald Giemsa. Larvae were separated from segments and preserved in 70% ethanol for identification. Macroscopically, some parts of the antler tips were lacking the compact part. Microscopically, within the Haversian and Volkmann canals, a large number of bacteria, scarce protein content and remnants of red blood corpuscles were visible. In the area of cancellous bone, cross-sectioned larvae were present. A large quantity of bacteria and a few degraded red blood corpuscles were visible on the smear made of liquid from the antlers. For morphological identification, three larvae were examined: two were third-instar larvae (L3), while one was a first instar larva (L1). Based on the shape of the cephaloskeleton, L3 was identified as Prochyliza nigrimanus and confirmed using molecular tools. To the extent of the authors' knowledge, this is the first record of Prochyliza nigrimanus in non-casted hard red deer antlers and the first description of this species in Croatia.