Several diseases caused by tick-borne pathogens, including Lyme borreliosis (LB) and spotted fever group rickettsioses, are endemic in the Balkan Peninsula, positioned between Central Europe and the Middle East. This cross-sectional study aimed to assess serological exposure to Borrelia spp. and spotted fever group Rickettsiae (SFGR) among individuals with recent tick bites and healthy controls in two Balkan countries—Serbia and North Macedonia. Serum samples from 223 participants were tested for anti-Borrelia and anti-SFGR IgG antibodies. SFGR exposure was significantly higher in tick-exposed individuals from Skopje (North Macedonia) compared to those from Novi Sad (Serbia) (30.9% vs. 8.0%; p = 0.003). In contrast, anti-Borrelia IgG was more frequently detected in Novi Sad, though differences did not reach statistical significance. The findings support a north-to-south gradient in Borrelia exposure and a reverse trend for SFGR, consistent with earlier studies and regional tick infection data. Given the high SFGR exposure and limited clinical reporting in North Macedonia, the results highlight the likelihood that tick-borne rickettsioses remain under-recognized. Additionally, Borrelia exposure in North Macedonia warrants further investigation. These findings emphasize the need for enhanced tick-borne disease surveillance, identification of endemic zones, and improved diagnostic and public health infrastructure in both countries.

Ticks and tick-borne pathogens (TBPs) pose an expanding health threat across Europe, yet few studies link tick infection data directly to human clinical outcomes. We conducted a prospective, cross-border "One Health" study in Serbia and North Macedonia to assess the prevalence, diversity, and potential transmission of TBPs from ticks to humans. A total of 133 tick-infested patients were enrolled at two healthcare centers (112 in Skopje, 26 in Novi Sad), and 135 ticks were collected from these individuals, alongside paired blood and serum samples. Overall, TBP DNA was detected in 66.6 % of ticks, with Rickettsia helvetica (25.9 %) and Rickettsia monacensis (12.5 %) were most common. Mixed infections occurred in 15 % of ticks, with up to three pathogens co-occurring. Other detected TBPs included Borrelia afzelii, Borrelia garinii, Borrelia lusitaniae, Anaplasma phagocytophilum, Babesia microti, and Babesia divergens. Four patients (3 %) had detectable pathogen DNA in blood, primarily Rickettsia spp. and Anaplasma spp., yet remained asymptomatic. Most common clinical outcome during the observation period was local hypersensitivity reaction (21.1 %), while α-Gal sensitization was detected in two cases. Phylogenetic analysis revealed Balkan Rickettsia and Borrelia strains were mostly identical to Western and Central European lineages, with a notable exception: a Bo. garinii strain from North Macedonia clustered with strains from Russia and China, indicating possible long-distance introduction. The present study demonstrates that human exposure to multiple TBPs is common and while asymptomatic bloodstream infections do occur, local hypersensitivity reactions are much more common.

Tick-borne diseases are rising across Europe, yet human-biting tick data from the central Balkans remain limited. We analysed ticks removed from patients seeking care after tick bites at centers in Skopje (North Macedonia) and Novi Sad (Serbia) between January 2022 and December 2024, recording species, developmental stage, month of removal, and anatomical attachment site. A total of 1641 ticks were collected (North Macedonia, n = 157; Serbia, n = 1484) and identified morphologically. Six tick species were detected in North Macedonia and eight in Serbia. Ixodes ricinus was dominant in both countries, comprising ~69% of ticks in North Macedonia and >90% in Serbia, while Hyalomma marginatum was found in North Macedonia. Adult females predominated among ticks in North Macedonia (75.8%), whereas nymphs were most frequent among ticks in Serbia (55.6%); between-center comparisons of developmental stage should be interpreted cautiously given the marked sampling imbalance. In both settings, collections peaked in May–June (~64% of annual ticks), indicating highest bite pressure in late spring/early summer. Attachment sites were most commonly the torso and lower extremities, with significant between-country differences and species-/stage-specific patterns most evident in Serbia. These data refine the epidemiology of human tick exposure in two Balkan countries and support targeted seasonal prevention messaging and strengthened regional surveillance.