Enterocytozoon bieneusi is a common human pathogen, responsible for more than 90% of overall human microsporidian infections. The epidemiology of E. bieneusi infections, however, is not well studied, and little is known about its transmission routes. Enterocytozoon bieneusi-like spores have been reported in surface and ground water in the United States and France, and a possible outbreak of microsporidiosis involving 200 persons was associated with drinking water in France. Recently, genotyping tools based on DNA sequencing of the internal transcribed spacer of the rRNA gene have been used in elucidating E. bieneusi transmission. Studies conducted in Europe (Switzerland, France, Germany, and UK) and Uganda identified nine E. bieneusi genotypes in humans. A more recent study in Peru with a larger sample size confirmed the presence of three of the nine genotypes in humans, but also found eight other genotypes that have not been found in humans before. As expected, geographic differences are present in the distribution of E. bieneusi genotypes in humans, with genotype B being the predominant genotype in Europe and genotype A the most common genotype in Peru. Four of the human-pathogenic genotypes have been found in domestic animals and wild mammals, and many of the 40 or so Enterocytozoon genotypes identified in animals are genetically related to the human-pathogenic genotypes, indicating that zoonotic transmission of E. bieneusi is possible. Nevertheless, host-adapted Enterocytozoon spp have been found in dogs, cattle, and some wild mammals (muskrats and raccoons), which do not have apparent public health significance. Further studies of E. bieneusi transmission in humans and animals in other regions should be helpful to elucidate the epidemiology of microsporidiosis.