Specimens of Contracaecum rudolphii Hartwich, 1964 (Nematoda: Anisakidae) from Phalacrocorax aristotelis (Linnaeus) from the Archipelago of La Maddalena (Sardinia, western Mediterranean Sea) were characterised genetically and compared with C. rudolphii A sensu D'Amelio et al. 1990 and C. rudolphii B sensu D'Amelio et al. 1990 from Phalacrocorax carbo sinensis (Blumenbach) from north-eastern Italy, and with C. rudolphii C sensu D'Amelio et al. 2007 from Phalacrocorax auritus (Lesson) from west-central Florida, USA. The sequencing of the small subunit of the mitochondrial ribosomal RNA gene (rrnS) and by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the same gene and of the internal transcribed spacers (ITS) of nuclear ribosomal DNA (rDNA) allowed the identification of all specimens of C. rudolphii from P. aristotelis as C. rudolphii A. The results confirmed that the definition of genetic markers, following the analysis of nuclear ribosomal and mitochondrial DNA, provides quick and practical diagnostic tools for the detection of the 3 sibling species of C. rudolphii. The occurrence of C. rudolphii in P. aristotelis is reported for the first time from the Mediterranean area, improving the picture of the dispersal patterns of the populations of these piscivorous birds, and confirming the existence of different and isolated populations between the North and South European waters.