Vector-borne diseases are endemic in Mediterranean regions, where dogs are frequently exposed to multiple pathogens throughout their lives. Although the immunological consequences of these infections have been widely investigated, less is known about the biochemical changes associated with natural pathogen exposure in clinically healthy autochthonous breeds. To characterize haematological and biochemical alterations associated with exposure to vector-borne pathogens in clinically healthy Mannara dogs living in an endemic Mediterranean area. Thirty-two clinically healthy Mannara dogs were sampled between September 2024 and January 2025. Antibodies against Anaplasma phagocytophilum, Ehrlichia canis, Leishmania infantum and Rickettsia spp. were determined by indirect immunofluorescence assay, whereas L. infantum DNA was assessed by real-time PCR. Complete blood count, serum biochemistry and serum protein electrophoresis were performed. Statistical analyses included parametric or non-parametric tests with false discovery rate correction. All dogs were negative for L. infantum DNA by PCR, although 37.5% were seropositive. Coinfections involving two or more pathogens were detected in 40.6% of the animals. Dogs seropositive for A. phagocytophilum showed significantly lower albumin concentrations and albumin/globulin ratios together with higher β2-globulin concentrations than seronegative dogs. Coinfected animals also exhibited increased β2- and γ-globulin fractions and reduced albumin concentrations. No significant differences were observed according to sex or age after multiple-comparison correction. Natural exposure to vector-borne pathogens was associated with consistent serum protein alterations in clinically healthy Mannara dogs despite the absence of detectable L. infantum parasitaemia. These findings are consistent with persistent antigenic stimulation associated with natural exposure to vector-borne pathogens and provide reference biochemical information for this autochthonous breed and support further studies investigating the biological significance of these alterations under natural exposure conditions.