Molecular evaluation of Hysterothylacium auctum (Nematoda, Ascaridida, Raphidascarididae) taxonomy from fish of the southern Baltic

Molecular investigations of Hysterothylacium auctum from Baltic Sea fish we re performed using multilocus allozyme electrophoresis (MAE) and genomic DN A analysis based on PCR technique. Nematodes from eel-pout, flounder, cod ( MAE and PCR) and herring (PCR) were examined to establish whether one speci es or complex of sibling ones occurs within Baltic H. auctum. Multilocus al lozyme electrophoresis revealed that three enzymatic loci: Got, Adk and Pep -2 were monomorphic, two loci: Mdh-1 and Pgm-2 showed slight polymorphism a nd one locus: Gpi showed strong polymorphism. H. auctum from eel-pout, flou nder and cod caught in the southern Baltic proved to share a common gene po ol; however, an incipient subdivision of this population into specimens con nected with eel-pout and flounder on the one hand and with cod on the other was ascertained with locus coding for Gpi. PCR-RFLP analysis of the fragme nt of ribosomal DNA embracing the 3'-end of the gene coding 18S rRNA, ITS1, gene coding 5.8S rRNA, ITS2 and the 5'-end of the gene coding 28S rRNA (si milar to 1500 bp) was performed using HaeIII, HinfI and PvuII endonucleases . In the case of particular endonuclease, electrophoretic patterns of diges ted PCR products were similar for all individuals of nematodes independent of host species or place of fishing. On the basis of nucleotide sequence of H. auctum obtained from eel-pout, a pair of species-specific primers was d esigned. This pair allows amplification of DNA of Hysterothylacium isolated from eel-pout, flounder, cod and herring and did not give cross-reactions with DNA from other Baltic anisakids: Anisakis simplex sensu stricto, Contr acaecum osculation s. str. and C. rudolphii. The results obtained suggest t hat only one biological species of H. auction occurs in the southern Baltic .