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The results of the first comprehensive study of myxomycetes from the island of Madagascar, a world biodiversity hotspot, are reported in this paper. The island is of continental origin, the fourth largest in the world, and has been geographically isolated for more than 160 million years, since its separation from Gondwanaland. The isolation, size and topography of Madagascar have triggered the development of a great variety of different habitats and favoured multiple evolutionary pathways, resulting in many animals and plants that exist nowhere else on earth. Fieldwork for the biodiversity survey of the central and southern parts of the island took place in May 2009, to coincide with the end of the rainy season. Tropical moist forest, sclerophyll forest and dry forest were selected for sampling in Ranomafana, Andringitra, Andohahela and L’Isalo National Parks. Some unique vegetation was sampled in the spiny dry forest and succulent scrub with plants from the genera Alluaudia, Euphorbia, Kalanchoe and Pachypodium. The survey produced 124 species from 22 different genera in more than 750 myxomycete collections. In this paper one species, Perichaena madagascariensis, is described as new to science, 21 species are new records for Africa, and 106 are reported for the first time from Madagascar. Some unusual collections included Physarum lakhanpalii that appeared on Ravenala madagascariensis, Fuligo intermedia and Licea nannengae found on Adansonia grandidieri, Perichaena pulcherrima, Physarum dictyosporum, and P. echinosporum on Euphorbia and Licea rufocuprea on bark. The scope, methods and results of this survey are included in this paper, and comments are made on the ecology, distribution and substrate association of the myxomycetes of these areas of Madagascar. Macrographs, micrographs and SEM images of interesting species are included. The results indicate that the island of Madagascar has a unique assemblage of species of myxomycetes, different from neighbouring islands and from similar but distant environments.