We report a new case of cutaneous leishmaniosis caused by Leishmania (Mundinia) martiniquensis in a horse in Florida, USA. A 10-year-old neutered male Quarter horse was presented with multifocal to coalescing, raised, ulcerated and oozing, non-healing wounds on both pinnae of several weeks’ duration. After a few months, the lesions regressed spontaneously. Biopsies of the lesions were performed with microscopical findings of epidermal hyperplasia with multifocal ulceration and focally extensive, dermal pyogranulomatous inflammation with numerous intact and degenerate neutrophils being surrounded by epithelioid macrophages, lymphocytes and plasma cells, as well as rare eosinophils. Within the macrophages, and freely within the inflammatory infiltrate, were small (2e4 mm) round, basophilic protozoal organisms. Immunohistochemistry and colourimetric in-situ hybridization were positive for amastigote forms of Leishmania spp. The species L. martiniquensis was identified by polymerase chain reaction targeting the ITS-1 gene performed with extracts from formalin-fixed and paraffin wax-embedded samples of skin lesions. L. martiniquensis causes an ulcerative pyogranulomatous dermatitis in horses with spontaneous healing. This second autochthonous case in Florida, 5 years after the first case, suggests that this parasite may have become endemic in this state.