The genus Perinereis, family Nereididae, consists of approximately 66 worldwide distributed species with marine, estuarine, and freshwater representatives. The objective of this study is to use morphological and molecular traits to evaluate the taxonomic differentiation of the estuarine polychaete P gualpensis. Also we reconstruct the phylogenetic relationships between P gualpensis and congeneric species recorded in Chilean coast: P. falklandica, P. longidonta and, especially with the morphologically most similar species, P vallata. Phylogenetic analyses based on molecular and morphological characters allow us to validate P. gualpensis and P. vallata as two independent and clearly defined species. Consequently, the taxonomic confusion about the possible sibling species P. gualpensis and P. vallata is completely resolved. However, morphological and genetic data are partially incongruent about the phylogenetic relationships of these four Perinereis species. We concluded that different evolutionary process that affect the morphological and genetics data produce different answers. Also we propose the use of new molecular evidence arising from molecular markers with different nucleotide substitution rate to that used here.