The Opisthokonta clade includes Metazoa, Fungi, and several unicellular lineages, such as choanoflagellates, filastereans, ichthyosporeans, and nucleariids. To date, studies of the evolutionary diversity of opisthokonts have focused exclusively on metazoans, fungi, and, very recently, choanoflagellates. Thus, very little is known about diversity among the filastereans, ichthyosporeans, and nucleariids. To better understand the evolutionary diversity and ecology of the opisthokonts, here we analyze published environmental data from nonfungal unicellular opisthokonts and report 18S ribosomal DNA phylogenetic analyses. Our data reveal extensive diversity among all unicellular opisthokonts, except for the filastereans. We identify several clades that consist exclusively of environmental sequences, especially among ichthyosporeans and choanoflagellates. Moreover, we show that the ichthyosporeans represent a significant percentage of overall unicellular opisthokont diversity, with a greater ecological role in marine environments than previously believed. Our results provide a useful phylogenetic framework for future ecological and evolutionary studies of these poorly known lineages.