A total of 836 unfed Ixodes ricinus ticks were collected from 2 forested areas of the Lublin region in eastern Poland. Of these, 540 ticks were collected in area ‘A’, exposed to flooding from the Vistula river, while the remaining 296 ticks were collected in suburban area ‘B’, not exposed to flooding. Ticks were examined by nested-PCR for the presence of DNA of Leptospira spp. and of Borrelia burgdorferi sensu lato, including its genospecies. The presence of the Leptospira spp. DNA was found in the examined specimens of Ixodes ricinus. The infection rate was much greater in area ‘A’ exposed to flooding, compared to unexposed area ‘B’ (15.6% vs. 1.4%, p<0.0001). A significant difference was noted in the case of all developmental stages. For the total results, the prevalence of Leptospira spp. in nymphs (16.9%) was two-fold greater (p<0.01) than in females and males (7.9% and 7.1%, respectively). The total prevalence of B. burgdorferi sensu lato in examined ticks amounted to 24.3%. Altogether, the genospecies Borrelia burgdorferi sensu stricto was detected most often. No correlation was found to exist between the presence of Leptospira spp. and B. burgdorferi sensu lato in the examined ticks, which indicates that the detection of Leptospira in ticks was not due to a false-positive cross-reaction with DNA of B. burgdorferi. In conclusion, this study shows for the first time the presence of Leptospira spp. in Ixodes ticks and marked frequency of the occurrence of these bacteria in ticks. This finding has significant epidemiological implications by indicating the possibility of the transmission of leptospirosis by Ixodes ricinus, the commonest tick species in Europe and most important vector of numerous pathogens.