Among the palaemonid shrimp species of the genus Palaemon known from Japan, Palaemon paucidens is the only species inhabiting freshwater areas. Two genetically distinct types (designated by A and B) were found by allozyme electrophoresis analysis three decades ago. We developed a simple multiplex-PCR assay to discriminate the two types based on 18S rDNA sequences. A total of 422 individuals collected at 152 localities in Japan were analyzed by the multiplex-PCR assay. No heterozygote was observed, indicating that these two types have been reproductively isolated. Both types were distributed throughout Japan; type A was observed in ponds, lakes and rivers, while type B was observed only in rivers.