Survey on the presence of A. simplex s.s. and A. pegreffii hybrid forms in Central-Western Mediterranean Sea

In this study, 1029 fish and cephalopod samples came from Central-Western Mediterranean (FAO 37.1.1 and FAO 37.1.3) were analysed for Anisakidae larvae research with the aim to identify possible hybridisations between Anisakis pegreffii and Anisakis simplex s.s. species. A total of 1765 larvae were detected, with prevalence values between 8.1% and 100%. The morphologic analysis revealed characters attributable to morphotype I of Anisakis in 98.5% of the examined larvae, while 1.5% belonged to the morphotype II. PCR-based Restriction Fragment Length Polymorphism (PCR–RFLP) analysis of the entire ITS region (ITS1, 5.8S and ITS2) of nuclear ribosomal DNA (rDNA) was performed with HinfI and HhaI restriction enzymes. The majority of the larvae examined by PCR-RFLP were identified as A. pegreffii (71%), with a prevalence on horse mackerel from FAO 37.1.3, while 10% were identified as A. simplex s.s., 2% as A. physeteris and 17% as A. pegreffii × A. simplex s.s. hybrid genotype. The sequence analysis confirmed the hybridisation in the 85% of the larvae recognised as hybrid forms by PCR- RFLP, suggesting this form as the product of natural interspecific recombination due to the presence of sympatry areas. The presence of hybrid forms were mostly found in fish samples from FAO subzone 37.1.1. This is the first report of A. pegreffii x A. simplex s.s. hybrid genotype in fishes caught off the coasts of Sicily (Southern Italy). Finally, this study provided substantial information about the geographical distribution of Anisakidae family in Central-Western Mediterranean Sea.