Rickettsial diseases present ongoing diagnostic challenges due to antigenic diversity, serological cross-reactivity, and the lack of reliable standardised control reagents. Current serodiagnostic assays frequently rely on convalescent human sera as positive controls, which are inherently variable, limited in supply, and poorly suited to assay standardisation and quality assurance. In this study, we developed and validated human-mouse chimeric monoclonal antibodies targeting the conserved outer membrane protein B (OmpB) of Rickettsia typhi as defined, renewable, positive-control calibrators for existing rickettsial serological platforms. Murine monoclonal antibodies were generated and engineered into chimeric IgG antibodies, which were expressed in HEK293T cells. The chimeric antibodies demonstrated high antigen specificity, bound native OmpB on the surface of R. typhi, and showed no detectable cross-reactivity within the tested antigen panel when evaluated using dot blot, immunofluorescence microscopy, and Luminex-based multiplex assays. Compared with patient-derived positive control sera, the chimeric antibodies demonstrated improved specificity and consistent performance within the evaluated assay conditions. These findings support the potential use of chimeric monoclonal antibodies as standardised positive controls for rickettsial serodiagnostics, with potential applications in assay validation, quality assurance, and inter-laboratory harmonisation.

This study investigated the prevalence and molecular characteristics of canine vector-borne pathogens (CVBPs) circulating in diverse dog populations in Thailand by using molecular diagnostic methods. A total of 400 blood samples were collected from four groups (n = 100 each): stray dogs (Group A), vector-borne disease-suspected companion dogs (Group B), healthy companion dogs presenting for routine examination (Group C), and companion dogs presenting with non-vector-borne illnesses (Group D). The overall infection rate was 46.25%. Ehrlichia spp. were the most frequently detected pathogens (23.5%), followed by Babesia spp. (16.5%), Rickettsia spp. (15.0%), and Anaplasma spp. (11.5%). The prevalence differed markedly among groups, including group A (88.0%), group B (54.0%), group C (27.0%) and group D (16.0%) (p < 0.05). DNA sequence analysis showed 100% identity with GenBank™ reference sequences, confirming the presence of Ehrlichia canis, Rickettsia asembonensis, Babesia vogeli, and Anaplasma platys. The detection of CVBPs across all groups demonstrates free-roaming and owned dogs serve as reservoirs for substantial ongoing infections and pose potential zoonotic implications to humans. Overall, these findings emphasize the importance of sustained molecular surveillance, improved vector control strategies, and proactive monitoring of high-risk dog populations to reduce the burden of CVBPs in Thailand.

Tick-borne diseases are a growing public health concern in the Mediterranean. Brijuni National Park (BNP), a unique, highly visited island ecosystem characterized by increased large game host density and diverse Mediterranean habitats, presents an elevated risk for pathogen co-circulation. This study addresses the lack of spatial and epidemiological data to accurately assess human exposure risk in this environment. We performed a detailed geospatial and epidemiological risk mapping of pathogen co-circulation in BNP. A total of 587 hard ticks were collected across 26 georeferenced micro-locations (2020-2022). Ticks were morphologically identified and subsequently screened for six key zoonotic bacterial pathogens using qPCR. The Minimal Infection Rate (MIR) and a Co-infection Rate (CR) were calculated. Geographic Information System (GIS) mapping was utilized to map ecological determinants of risk. Ixodes ricinus was the overwhelmingly dominant vector (94.0%), peaking in spring, with activity absent in summer. Recorded diverse tick fauna also included Hyalomma marginatum (3%), Haemaphysalis punctata (2%), Ixodes frontalis (0.8%) and Rhipicephalus sanguineus (0.2%). Active circulation of Borrelia burgdorferi s.l. (Bbsl), Anaplasma phagocytophilum, and Ehrlichia canis were confirmed. Bbsl presented the highest MIR (3.05). The Co-infection Rate (CR) was notably high at 29.41%, with triple co-infections (Bbsl, A. phagocytophilum, E. canis) concentrated in cultivated mosaics and holm oak forests (Quercus ilex L.). The highest number of ticks was recovered from ecotone zones, accounting for 50.0% of the total catch, confirming them as high-risk interfaces. The absence of Rickettsia conorii may be attributed to the strict control/absence of its primary host (domestic dogs). The presence of the exotic vector H. marginatum was also confirmed. The high rate of co-infection and the spatial concentration of risk in specific habitats underscore an elevated and complex public health risk in BNP, closely linked to habitat structure and increased game host density. This research provides an essential geospatial framework for targeted 'One Health' management, prioritizing vector control in ecotone zones and dense forest refugia. Urgent surveillance for the exotic H. marginatum is warranted to monitor the potential risk of Crimean-Congo Hemorrhagic Fever.

Leptospirosis, Q fever, scrub typhus, and murine typhus are endemic zoonotic diseases in southern Taiwan that often present as acute undifferentiated febrile illness. We retrospectively compared epidemiological, clinical, laboratory, and outcome characteristics of leptospirosis (n = 41) and rickettsial infections, including Q fever, scrub typhus, and murine typhus (QSM; n = 103), in Kaohsiung City from 2013 to 2024. Leptospirosis occurred mainly during rainy and typhoon seasons and was associated with myalgia, gastrointestinal symptoms, jaundice, conjunctival suffusion, hepatic and renal dysfunction, leukocytosis, elevated C-reactive protein, and higher rates of acute kidney injury and shock. QSM occurred year-round, with eschar characteristic of scrub typhus, while prolonged activated partial thromboplastin time and relative bradycardia were more common in QSM. QSM generally showed a milder clinical course, particularly Q fever. Distinct clinical and seasonal patterns may facilitate early differentiation and risk stratification in endemic areas.

Domestic dogs have been proposed as sentinels for various infectious diseases, including rickettsial infections. In Chile, the only endemic human rickettsiosis is scrub typhus caused by Ca ndidatus Orientia chiloensis, an emerging infectious disease, discovered in recent years in southern Chile. The geographic distribution of scrub typhus and other rickettsial infections across Chile is incompletely understood. This study using domestic dogs as sentinel investigated the potential exposure to members of the three main groups of rickettsiae, scrub typhus orientiae (STGO), spotted fever group rickettsiae (SFGR) and typhus group rickettsiae (TGR), in northern, central, and southern Chile. Serological analyses for STGO, SFGR, and TGR were conducted on 1097 dogs from rural and urban areas, which were sampled during 2010 to 2016 in five Chilean regions using a cross-sectional household-based design. Household and dog-level variables were assessed to investigate retrospectively associated risk factors for rickettsial exposure. Overall seroprevalences were 13.8% for STGO, 1.2% for SFGR, and 1.0% for TGR. Higher seropositivity for STGO was associated with dogs from rural areas, male dogs, older dogs, and dog infestation with ticks. Canine seroreactivity to STGO was found across Chile, including regions not known as endemic for scrub typhus. Absence of external deworming was a significant risk factor for SFGR. The present results suggest that during the study period dogs were exposed to rickettsiae over a broader geographic range than previously recognized. Future epidemiological, ecological, and clinical studies are required to confirm the distribution of Orientia species and potential endemicity of other rickettsiae in Chile.

The objective of this study was to investigated the presence of Rickettsiales bacteria and Piroplasmida protozoa in canine blood samples from different peri-urban/rural localities in eastern Salta Province, Argentina. Complete DNA was extracted from 77 blood samples and analysed by conventional PCR assays targeting fragments of the genes for 16S rRNA (Anaplasmataceae), 18S rRNA (Piroplasmida) and gltA (Rickettsia). Also, a real-time PCR assay for direct detection of Ehrlichia canis and phylogenetic analyses of obtained gene sequences were applied. The analyses resulted in the detection of Anaplasma platys (10.91%), Babesia vogeli (2.6%) and E. canis (14.29%). A co-infection with different microorganism and rickettsial infection was not observed. The detection of B. vogeli represents the northernmost finding of this protozoa in Argentina. These results demonstrate the circulation canine vector-borne diseases agents in eastern Salta Province and expand the knowledge of the geographical distribution of these pathogens.

Rickettsiales bacteria represent a major global public health threat, yet their distribution patterns along ecological gradients remain poorly characterized. The distinctive east-west biome transition in Inner Mongolia provides an ideal model for studying how environmental factors shape the prevalence and diversity of Rickettsiales. Molecular screening for Rickettsia, Anaplasma, and Ehrlichia species was performed on 2372 ticks collected from diverse hosts and environmental sources across five leagues in Inner Mongolia. Nine tick species from four genera were identified, with Dermacentor silvarum being the dominant species, accounting for 24.7% of collected specimens. The overall infection rate of Rickettsia spp. was 59.8%, encompassing seven species. Anaplasma spp. infection was detected at 13.9%, involving five species, while Ehrlichia spp. showed a prevalence of 5.6%, comprising three species. A distinct east-west divergence in pathogen distribution was observed. Eastern forest-steppe regions exhibited higher Rickettsia prevalence at 62.5%, primarily vectored by Ixodes persulcatus. Western arid regions demonstrated elevated infection levels of Anaplasma (26.4%) and Ehrlichia (7.7%), mainly associated with Dermacentor nuttalli and Hyalomma asiaticum. Co-infections occurred in 12.3% of ticks, with Rickettsia raoultii and Anaplasma ovis co-infection peaking at 65.4% in D. nuttalli from Urad Back Banner, indicating a strong association with co-detection patterns in desert-steppe environments. Phylogenetic analyses based on 16S rRNA, gltA, and groEL genes confirmed species identification and revealed geographic genetic variations. These findings provide baseline data on tick-borne pathogens in Inner Mongolia and highlight the influence of ecological heterogeneity on pathogen distribution. Region-specific surveillance and control strategies that consider environmental conditions and dominant tick species may improve the management of tick-borne disease risks in ecologically diverse regions.

Rocky Mountain spotted fever (RMSF) is an important and potentially fatal tick-borne disease impacting people and dogs. RMSF, caused by Rickettsia rickettsii, is frequently reported in parts of the North, Central and South American countries. The treatment for RMSF is limited to tetracyclines, specifically to doxycycline. If untreated, the disease can progress rapidly to fatal outcomes, particularly in children. In recent years, fatality rates ranging from 30% to 80% have been reported throughout South America, Mexico, and in parts of Indian reservations in southwest regions of the USA. To aid in support of monitoring in vivo R. rickettsii infections, we developed a novel quantitative PCR (qPCR) assay to facilitate the sensitive and specific detection of R. rickettsii. The assay was designed and optimized using the surface cell antigen 4 gene as the target. The utility of the assay was defined for its specificity, analytical sensitivity, efficiency, and to monitor infections in the canine host following R. rickettsii infection challenges. With having high specificity and analytical sensitivity, the qPCR assay can be utilized for diverse research applications promoting investigations on R. rickettsii.

Ticks pose an increasing threat to public health due to their geographic expansion and the emergence of tick-borne pathogens. In Nicaragua, information on ticks that infest humans is limited. This study reports human tick bites recorded between March 2023 and September 2025 in rural and peri‑urban areas of the departments of Chontales and Río San Juan. Twenty-five cases were analyzed, with 172 ticks collected (28 females, 6 males, 13 nymphs, and 125 larvae). Morphological identification revealed five species: Amblyomma mixtum, Amblyomma ovale, Amblyomma cf. parvum, Rhipicephalus linnaei, and Rhipicephalus microplus. Amblyomma mixtum was the dominant species, present in 80% of cases, and all developmental stages were found attached to humans. Skin reactions ranged from pruritus and local erythema to intense inflammatory responses associated with massive larval infestations (up to 123 larvae in a single individual). One infested individual presented transient systemic symptoms (fatigue, generalized pruritus, difficulty concentrating, and insomnia) that resolved after tick removal. This is the first report of A. ovale, A. cf. parvum, R. linnaei, and R. microplus infesting humans in Nicaragua. These findings underscore the risk of tick-borne diseases in rural areas of Nicaragua and the need for active surveillance, accurate tick identification, and molecular studies to detect rickettsial agents and other potential pathogens. Information on preferred attachment sites (e.g., lower limbs) is clinically relevant and may help inform preventive measures.