In several urban and peri‑urban areas of Brazil, populations of Amblyomma sculptum and Amblyomma dubitatum ticks are maintained by capybaras (Hydrochoerus hydrochaeris). In some of these areas, this host and these tick species are associated with Brazilian spotted fever (BSF), a lethal human disease caused by the bacterium Rickettsia rickettsii. In this work, we evaluated the risk of human exposure to these tick species using four collection techniques to discern host-seeking behavior. The study was carried out in 10 urban sites inhabited by capybaras in Uberlândia, a BSF-free municipality in southeastern Brazil. Ticks were collected in areas of 400 m² at each site and at three seasons. Within the same municipality, the distance and speed of A. sculptum nymphs moving towards the CO₂ traps were evaluated. In a sample of ticks Rickettsia DNA was investigated. During the study period, 52,953 ticks were collected. Among these, 83.4 % were A. sculptum (1,523 adults, 10,545 nymphs and 32,104 larvae) and 16.6 % were A. dubitatum (464 adults, 2,153 nymphs and 6,164 larvae). An average annual questing tick density of 4.4/m² was observed, with the highest density recorded at one site in autumn (31.8/m²) and the lowest in summer at another site (0.03/m²). The visual search yielded the highest proportion of A. sculptum larvae, constituting 47 % of the total and 63.6 % of all A. sculptum larvae. In contrast, CO₂ traps collected a greater proportion of nymphs and adults of A. sculptum ticks. In the case of A. dubitatum, the CO₂ trap was the most efficient technique with 57.7 % of captures of this species, especially of nymphs (94.5 % of captures) and adults (97.8 % of captures). Ticks' ambush height on vegetation (9 to 77 cm), observed by visual search 30 times, yielded a total of 20,771 ticks. Of these, 28 (93 %) were A. sculptum ticks, with only two (7 %) identified as A. dubitatum ticks. Among A. sculptum ticks, the nymph was the most attracted stage to humans and larva in the case of A. dubitatum. Amblyomma sculptum adults and nymphs were significantly more attracted to humans than those of A. dubitatum, but A. dubitatum larvae were significantly more attracted than the same stage of A. sculptum. The maximum distance and speed of horizontal displacement for A. sculptum nymphs were five meters and 2.0 m/h, respectively. The only species of Rickettsia detected in ticks, exclusively in A. dubitatum, was R. bellii. Importantly, it was observed that the higher the proportion of A. sculptum in the community of ticks, the lower the rate of infection of A. dubitatum by R. bellii. In conclusion, host-seeking behavior differed between the two tick species, as well as between stages of the same species. A greater restriction of A. dubitatum ticks to the soil was observed, while larvae and nymphs of A. sculptum dispersed higher in the vegetation. The behavior presented by A. sculptum provides greater opportunities for contact with the hosts, while A. dubitatum depends more on an active search for a host, the hunter behavior. Taken together, these observations show that a human being crossing an area infested with A. sculptum and A. dubitatum ticks will have almost exclusive contact with A. sculptum larvae and/or nymphs. Humans in a stationary position (sitting, lying or immobile) are exposed to both tick species, but they are more attractive to adults and mainly nymphs of A. sculptum compared to the corresponding stages of the tick A. dubitatum. The negative effect of A. sculptum on A. dubitatum infection by R. bellii deserves further studies.

Diagnosing canine visceral leishmaniasis (CVL) in Brazil faces challenges due to the limitations regarding the sensitivity and specificity of the current diagnostic protocol. Therefore, it is urgent to map new antigens or enhance the existing ones for future diagnostic techniques. Immunoinformatic tools are promising in the identification of new potential epitopes or antigen candidates. In this study, we evaluated peptides selected by epitope prediction for CVL serodiagnosis in ELISA assays. Ten B-cell epitopes were immunogenic in silico, but two peptides (peptides No. 45 and No. 48) showed the best performance in vitro. The selected peptides, both individually and in combination, were highly diagnostically accurate, with sensitivities ranging from 86.4% to 100% and with a specificity of approximately 90%. We observed that the combination of peptides showed better performance when compared to peptide alone, by detecting all asymptomatic dogs, showing lower cross-reactivity in sera from dogs with other canine infections, and did not detect vaccinated animals. Moreover, our data indicate the potential use of immunoinformatic tools associated with ELISA assays for the selection and evaluation of potential new targets, such as peptides, applied to the diagnosis of CVL.

Dogs accessing natural areas in Brazil are exposed to tick species typically associated with wild carnivores, some of which are vectors of pathogenic Rickettsia. Infested dogs may transport ticks into households and act as amplifying hosts for these agents. Iguaçu National Park (INP), a major inland Atlantic Forest reserve in Brazil, harbors at least nine tick species and five Rickettsia species. This study evaluated tick infestation in 116 rural dogs from properties along most of the 420 km perimeter of INP and 22 urban dogs from forest-adjacent areas in Foz do Iguaçu. Sera from 39 dogs were tested against five Rickettsia antigens using an indirect immunofluorescence assay. A total of 169 ticks were collected, including Rhipicephalus linnaei (n=87), Amblyomma ovale (n=62), Amblyomma coelebs (n=5), and Rhipicephalus microplus (n=3), with an overall infestation prevalence of 28.3%. Seroprevalence against Rickettsia spp. was 42.9% in rural dogs and 18.1% in urban dogs. The highest titers were 1:512 in rural dogs (against R. parkeri, R. amblyommatis, and R. rhipicephali) and 1:256 in urban dogs (against R. rickettsii), while median titers were zero. Both tick infestation and Rickettsia exposure were lower than those reported in dogs from other Atlantic Forest regions. These findings suggest an overall limited exposure of free-ranging dogs to ticks and Rickettsia spp. Further investigation is needed to understand the causes of this low exposure, including the potential influence of the jaguar population and other wild carnivores in INP, which may restrict dog access to forested areas.