Although they represent powerful genetic markers in many fields of biology, microsatellites have been isolated in few fungal species. The aim of this study was to assess whether obtaining microsatellite markers with an acceptable level of polymorphism is generally harder from fungi than in other organisms. We therefore surveyed the number, nature and polymorphism level of published microsatellite markers in fungi from the literature and from our own data on seventeen fungal microsatellite-enriched libraries, and in five other phylogroups (angiosperms, insects, fishes, birds and mammals). Fungal microsatellites indeed appeared both harder to isolate and to exhibit lower polymorphism than in other organisms. This appeared to be due, at least in part, to genomic specificities, such as scarcity and shortness of fungal microsatellite loci. A correlation was observed between mean repeat number and mean allele number in the published fungal microsatellite loci. The cross-species transferability of fungal microsatellites also appeared lower than in other phylogroups. However, microsatellites have been useful in some fungal species. Thus, the considerable advantages of these markers make their development worthwhile, and this study provides some guidelines for their isolation. © 2007 Elsevier Inc. All rights reserved.