This study compared serological and molecular approaches for detecting Leptospira spp. using paired blood and urine samples from 124 asymptomatic stray dogs. Blood sera were screened via the microscopic agglutination test (MAT) for ten serovars. Urine samples were analyzed using four PCR assays: a commercial quantitative PCR (qPCR), a lipL32 qPCR, a conventional rrs PCR, and a multiplex PCR (mPCR). Serological exposure was detected in 26 (20.97%) dogs, while commercial qPCR, rrs PCR, lipL32 qPCR, and mPCR assays yielded positive results in 31 (25.00%), 34 (27.42%), 25 (20.16%), and 23 (18.55%) dogs, respectively. Compared with commercial qPCR, the lipL32 qPCR, rrs PCR, and mPCR demonstrated Positive Percentage Agreement (PPA) values of 80.65%, 87.10%, and 67.74%, and Negative Percentage Agreement (NPA) values of 100.00%, 92.47%, and 97.85%. Nineteen urine samples were positive across all four PCR assays. Overall, 42.74% (53/124) of dogs had at least one positive serological or molecular finding, including nine additional conventional PCR-positive findings without reliable sequence confirmation. Sequencing and phylogenetic analysis of 24 samples identified L. interrogans and L. borgpetersenii, representing the first molecular characterization of Leptospira in Bosnia and Herzegovina. The findings demonstrate the complementary value of serological and molecular approaches for Leptospira surveillance while emphasizing the need for cautious interpretation of unconfirmed molecular findings.

Canine filariosis is an emerging vector-borne parasitic disease of veterinary and public health importance, caused primarily by D. immitis and D. repens. The present study investigated the prevalence, risk factors, diagnostic performance of different methods, and molecular characteristics of filarial infections in dogs from the Republic of Srpska (Bosnia and Herzegovina). Blood samples from 418 dogs collected between 2020 and 2024 were examined using a modified Knott test, antigen enzyme-linked immunosorbent assay, and polymerase chain reaction-based methods. Overall, 21.3% of dogs were positive for at least one filarial species. D. repens was the most prevalent species (13.40%), followed by D. immitis (10.8%) and A. reconditum (1.4%). Coinfections with D. immitis and D. repens were detected in 3.35% of dogs, while triple infections occurred in 0.5%. Several individual and environmental factors were associated with an increased risk of infection, including poor body condition, mixed-breed status, inadequate parasite control, and specific movement histories. PCR showed the highest diagnostic performance for all detected species, outperforming both microscopy and antigen testing. Phylogenetic analysis revealed genetic diversity among the detected isolates and demonstrated their relationship to strains previously reported from other European countries. These findings confirm the endemic presence of canine filarial infections in the Republic of Srpska and emphasize the importance of molecular diagnostics, surveillance programs, and preventive measures for effective disease control.