| Sažetak: | Leptospirosis is a re-emerging, worldwide zoonosis, and wild boar (<i>Sus scrofa</i>) are involved in its epidemiology as the reservoir. The aim of this study was to investigate the prevalence of <i>Leptospira</i> with serological, bacteriological, and molecular assays in wild boar hunted in Tuscany (Italy) during two hunting seasons. In total, 287 specimens of sera, kidneys, and liver were collected to perform microscopic agglutination tests (MATs), isolation, and RealTime PCR to detect pathogenic (<i>lipL32</i> gene), intermediate (<i>16S rRNA</i> gene), and saprophytic (<i>23S rRNA</i> gene) <i>Leptospira</i>. Within sera, 39 (13.59%) were positive to the MAT, and Australis was the most represented serogroup (4.88%), followed by Pomona (4.18%), and Tarassovi (3.14%). Moreover, four <i>Leptospira</i> cultures were positive, and once isolates were identified, one was identified as <i>L. borgpetersenii</i> serovar Tarassovi, and three as <i>L. interrogans</i> serovar Bratislava. Pathogenic <i>Leptospira</i> DNA were detected in 32 wild boar kidneys (11.15%). The characterization through the amplification of the <i>rrs2</i> gene highlighted their belonging to <i>L. interrogans</i> (23 kidneys), <i>L. borgpetersenii</i> (four), and <i>L. kirschneri</i> (one), while nine kidneys (3.14%) were positive for intermediate <i>Leptospira</i>, all belonging to <i>L. fainei.</i> The results of this study confirmed the importance of wild boar in the epidemiology of leptospirosis among wildlife in Central Italy.
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