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Urban brown rats (Rattus norvegicus) are frequently implicated to spread of Leptospira spp. Urban brown rats (n = 116) were trapped in different places near the lakes and ponds from Holoseevo and Obolon districts of Kyiv, and tested for Leptospira spp. using an agglutination test (MAT), which was conducted with 21 Leptospira's serological groups. It is identified the circulation of pathogenic Leptospira among urban brown rats population from Kyiv, Ukraine. In total, 65 (56%) rats were positive by MAT. The antibodies against serogroups Grippotyphosa, Mini and Ballum were dominant in positive reactions.