The internal transcribed spacer (ITS-1 and ITS-2) regions and the 5·8S ribosomal RNA gene of 2 Perkinsus spp. (G117 and H49) originating from the softshell clam, Mya arenaria, of the Chesapeake Bay were cloned and sequenced to obtain evidence for their genetic divergence. A high level of heterogeneity in both regions, probably resulting from deletions, insertions, and base substitutions, was evident from alignments of the sequences of the 2 isolates with published sequences of other Perkinsus spp. The isolate G117 and other Perkinsus spp. were highly divergent (13–26% and 19–20% sequence divergence in ITS-1 and ITS-2, respectively). These regions in the isolate H49 and Perkinsus marinus were similar (99·07% and 99% for ITS-1 and ITS-2, respectively). Evidence obtained from a phylogenetic analysis using the aligned sequences suggests that G117 and H49 belong to 2 distinct species of Perkinsus. The isolate G117 possibly belongs to an as yet undescribed species of Perkinsus, and H49 belongs to the species P. marinus. The conclusions drawn from the genetic analysis of H49 and G117 are supported by previously reported morphological characteristics (McLaughlin & Faisal, 1998b). Isolates H49 and G117 originated from the same molluscan species demonstrating that at least 2 different species of Perkinsus can co-exist in 1 host.