INTRODUCTION. Eustrongylides spp. are parasitic nematodes of piscivorous birds and freshwater fish, which act as their definitive and intermediate/paratenic hosts respectively, and are as well recognized as potentially zoonotic parasites. To date, Eustrongylides has been reported from fish and birds from different areas of central Italy (Dezfuli et al., 2015Parasites Vectors. 8: 227; Mazzone et al., 2019 J Parasitol. 105: 882-89; Guardone et al., 2021 Food Control. 120: 107517); therefore, the purpose of this study was to investigate the presence of the parasite in fish populations from areas not yet studied in northern Italy. MATERIALS AND METHODS. A total of 2130 specimens belonging to a wide range of fish species (Perca fluviatilis, Lepomis gibbosus, Micropterus salmoides, Squalius cephalus, Scardinius erythrophthalmus, Carassius carassius, Rutilus rutilus, Alburnus arborella, Ameiurus melas, Silurus glanis) were collected with the help of local fishermen from six lakes in northern Italy (Garda, Nero, Alice, Pistono, Sirio, Candia) and subjected to a full parasitological examination. Collected nematodes were fixed in 70% ethanol and identified morphologically through light and scanning electron microscopy; a subset of larvae was analyzed with molecular methods (amplification and sequencing of ITS rDNA) for species confirmation. RESULTS AND CONCLUSIONS. Overall, Eustrongylides larvae were recovered in 4.88% of the examined fish and were exclusively found in the species P. fluviatilis, L. gibbosus and M. salmoides, with particularly high prevalence in L. gibbosus (as high as 95.45% in Candia lake). The parasite occurrence was confirmed in all six surveyed lakes with prevalence values higher in Candia lake (65.15%) and lower in Sirio lake (1.42%). Morphological and molecular analyses allowed to confirm that all larvae belonged to the species Eustrongylides excisus. The distribution of E. excisus in all the lakes under study and the high prevalence values reported in some fish species throughout the present investigation highlight a possible emergence of this parasitic infection, which could represent an important health issue for piscivorous birds, being particularly pathogenic for nestlings, as well as a potential zoonotic risk.