Nuclear 18S rRNA gene sequences of oyster shell borers, Polydora websteri, P. calcarea and P. haswelli (Polychaeta: Spionidae), were determined for the first time. A wide range of morphological variation, particularly with regard to pigmentation, is observed among these three species. This variation was characterized at the species level. Black pigmentation pattern on the palps is suggested to be a key characteristic for species determination. Polydora websteri and P. haswelli were sister species. This is the first record of P. calcarea from Japan and Australia and P. haswelli from Japan.