Prevalence of tick-borne pathogens in ticks collected from the wild mountain ungulates mouflon and chamois in 4 regions of France.

Ticks are major vectors of various pathogens of health importance, such as bacteria, viruses and parasites. The problems associated with ticks and vector-borne pathogens are increasing in mountain areas, particularly in connection with global climate change. We collected ticks (n = 2,081) from chamois and mouflon in 4 mountainous areas of France. We identified 6 tick species: Ixodes ricinus, Rhipicephalus bursa, Rh. sanguineus s.l., Haemaphysalis sulcata, H. punctata and Dermacentor marginatus. We observed a strong variation in tick species composition among the study sites, linked in particular to the climate of the sites. We then analysed 791 ticks for DNA of vector-borne pathogens: Babesia/Theileria spp., Borrelia burgdorferi s.l., Anaplasma phagocytophilum, A. marginale, A. ovis, and Rickettsia of the spotted fever group (SFG). Theileria ovis was detected only in Corsica in Rh. bursa. Babesia venatorum (2 sites), Borrelia burgdorferi s.l. (B. afzelii and B. garinii; 2 sites) and Anaplasma phagocytophilum (3 sites) were detected in I. ricinus. Anaplasma ovis was detected at one site in I. ricinus and Rh. sanguineus s.l. SFG Rickettsia were detected at all the study sites: R. monacensis and R. helvetica in I. ricinus at the 3 sites where this tick is present; R. massiliae in Rh. sanguineus s.l. (1 site); and R. hoogstraalii and Candidatus R. barbariae in Rh. bursa in Corsica. These results show that there is a risk of tick-borne diseases for humans and domestic and wild animals frequenting these mountain areas.

Title: Prévalence d'agents pathogènes vectorisés chez des tiques collectées chez des ongulés sauvages (mouflons, chamois) dans 4 zones montagneuses en France. Abstract: Les tiques sont des vecteurs majeurs de différents agents pathogènes d'importance sanitaire, tels que des bactéries, des virus et des parasites. Les problématiques liées aux tiques et aux pathogènes vectorisés augmentent en zones de montagne, en lien notamment avec le réchauffement climatique. Nous avons collecté des tiques (n = 2 081) sur des chamois et des mouflons dans 4 zones montagneuses en France. Six espèces ont été identifiées : Ixodes ricinus, Rhipicephalus bursa, Rh. sanguineus s.l., Haemaphysalis sulcata, H. punctata et Dermacentor marginatus. Nous avons observé une forte variation de la composition en espèces de tiques entre les sites d'étude, en lien notamment avec le climat des sites. Nous avons ensuite recherché les ADN d'agents pathogènes vectorisés sur 791 tiques : Babesia/Theileria spp, Borrelia burgdorferi s.l., Anaplasma phagocytophilum, A. marginale, A. ovis, et de Rickettsia du groupe des fièvres boutonneuses (SFG). Theileria ovis a été détecté uniquement en Corse chez Rh. bursa. Babesia venatorum (2 sites), Borrelia burgdorferi s.l. (B. afzelii and B. garinii; 2 sites) et Anaplasma phagocytophilum (3 sites) ont été détectés chez I. ricinus. Anaplasma ovis a été détecté dans un site chez I. ricinus et Rh. sanguineus s.l.. Les Rickettsia SFG ont été détectées dans tous les sites d'étude : Rickettsia monacensis et R. helvetica chez I. ricinus dans les 3 sites où cette tique est présente; R. massiliae chez Rh. sanguineus s.l. (1 site); et R. hoogstraalii et Candidatus R. barbariae chez Rh. bursa en Corse. Ces résultats montrent un risque de transmission de maladies par les tiques pour les personnes et les animaux domestiques et sauvages fréquentant ces zones de montagne.

Seasonal dynamics and genetic characterization of bovine arthropod-borne parasites in Nan Province, Thailand with molecular identification of Anaplasma platys and Trypanosoma theileri.

Virulent species or strains of hematophagous borne pathogens such as Anaplasma spp., Babesia spp., Theileria spp., and Trypanosoma spp., are lethal to susceptible animals or reduce their productivity on a global scale. Nonetheless, efforts to diagnose the causative agents and assess the genotypic profiles as well as quantify the parasite burden of aforementioned parasites across seasons remain limited. Therefore, the present investigation sought to elucidate the genotypic composition of Anaplasma spp., Babesia spp., Theileria spp., and Trypanosoma spp. The findings revealed heightened infection rates during the summer, manifesting a correlation between Trypanosoma spp. infection and seasonal fluctuations. Among the identified pathogens, Anaplasma marginale emerged as the most dominant species, while the occurrence of Anaplasma platys in Thai cattle was confirmed via the sequencing of the groEL gene. Moreover, the study successfully identified two lineages of Trypanosoma theileri. The findings of this investigation offer valuable insights that can inform the development of preventive strategies for vector-borne diseases, such as considering the appropriate use of insect repellent, mosquito or insect nets, or eliminating breeding places for insects in each season.

Evaluation of the efficacy of Simparica Trio® in the prevention of the transmission of Babesia canis by infected Dermacentor reticulatus to dogs.

BACKGROUND: Babesia canis is a clinically relevant vector-borne pathogen in dogs, and its presence is expanding. The efficacy of Simparica Trio® (Zoetis) in the prevention of B. canis transmission was evaluated at the minimum recommended label dose of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin and 5 mg/kg pyrantel per kg bodyweight. METHODS: Twenty-four (24) dogs were randomly allocated to either a placebo-treated group or one of two treatment groups with Simparica Trio. Dogs were infested with B. canis-infected Dermacentor reticulatus ticks 21 or 28 days after treatment administration. Blood samples for antibody and DNA detection were collected from each dog prior to tick infestation until 28 days after infestation. A dog was defined as being B. canis positive if it tested positive by both an indirect immunofluorescence assay (IFA) and PCR at any time during the study. RESULTS: No treatment-related adverse reactions were recorded during the study. All placebo-treated animals displayed clinical signs due to babesiosis and tested positive on both IFA and PCR. None of the Simparica Trio-treated animals displayed any clinical symptoms or tested positive, resulting in a 100% efficacy in the prevention of canine babesiosis (P < 0.0001). CONCLUSIONS: A single treatment with Simparica Trio at the minimum recommended label dose of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin and 5 mg/kg pyrantel per kg bodyweight prevents the transmission of B. canis by infected D. reticulatus to dogs for at least 28 days.

Comparative chemical genomics in Babesia species identifies the alkaline phosphatase PhoD as a determinant of antiparasitic resistance.

Babesiosis is an emerging zoonosis and widely distributed veterinary infection caused by 100+ species of Babesia parasites. The diversity of Babesia parasites and the lack of specific drugs necessitate the discovery of broadly effective antibabesials. Here, we describe a comparative chemogenomics (CCG) pipeline for the identification of conserved targets. CCG relies on parallel in vitro evolution of resistance in independent populations of Babesia spp. (B. bovis and B. divergens). We identified a potent antibabesial, MMV019266, from the Malaria Box, and selected for resistance in two species of Babesia. After sequencing of multiple independently derived lines in the two species, we identified mutations in a membrane-bound metallodependent phosphatase (phoD). In both species, the mutations were found in the phoD-like phosphatase domain. Using reverse genetics, we validated that mutations in bdphoD confer resistance to MMV019266 in B. divergens. We have also demonstrated that BdPhoD localizes to the endomembrane system and partially with the apicoplast. Finally, conditional knockdown and constitutive overexpression of BdPhoD alter the sensitivity to MMV019266 in the parasite. Overexpression of BdPhoD results in increased sensitivity to the compound, while knockdown increases resistance, suggesting BdPhoD is a pro-susceptibility factor. Together, we have generated a robust pipeline for identification of resistance loci and identified BdPhoD as a resistance mechanism in Babesia species.

Epidemiology and genetic diversity of Theileria equi and Babesia caballi in Mongolian horses.

Equine piroplasmosis is a tick-borne disease caused by Theileria equi and Babesia caballi in horses. Because of its impact on horse industry, control of this disease is crucial for endemic countries. The control of equine piroplasmosis may be influenced by the genotypic diversity of T. equi and B. caballi. Mongolia, a country with a thriving livestock industry, is endemic for T. equi and B. caballi. However, nationwide epidemiological surveys have not been conducted to determine the current status of infections and genetic diversity of these two parasite species. Therefore, the objective of this research was to investigate the infection rates and genotypes of T. equi and B. caballi in horses across Mongolia. Blood samples were collected from 1353 horses in 15 of Mongolia's 21 provinces, and their DNAs were analyzed with T. equi- and B. caballi-specific PCR assays. Additionally, blood smears were prepared from 251 horses, stained with Giemsa, and examined under a light microscope to identify T. equi and B. caballi. The microscopy revealed that 30 (11.9%) and 4 (1.6%) of the 251 horses were positive for T. equi and B. caballi, respectively. By contrast, PCR assays detected the T. equi and B. caballi in 1058 (78.2%) and 62 (4.6%) horses, respectively. Phylogenetic analysis of 18S rRNA sequences from 42 randomly selected T. equi-positive DNA samples detected the genotypes A and E. On the other hand, the rap-1 sequences from 19 randomly selected B. caballi-positive DNA samples occurred in clades representing the genotypes A and B1, as well as in a distinct clade closely related to the genotype A. Our findings confirm the widespread occurrence of T. equi and B. caballi infections in Mongolian horses, highlighting the need for a comprehensive control approach.

Transmission risk evaluation of transfusion blood containing low-density Babesia microti.

Background: Babesia is a unique apicomplexan parasite that specifically invades and proliferates in red blood cells and can be transmitted via blood transfusion, resulting in transfusion-transmitted babesiosis. However, detecting Babesia in blood before transfusion has not received enough attention, and the risk of transfusing blood containing a low density of Babesia microti (B. microti) is unclear, possibly threatening public health and wellness. Purpose: This study aimed to determine the lower detection limit of B. microti in blood and to evaluate the transmission risk of blood transfusion containing low-density B. microti. Methods: Infected BALB/c mouse models were established by transfusing infected whole blood with different infection rates and densities of B. microti. Microscopic examination, nested Polymerase Chain Reaction (nested PCR), and an enzyme-linked immunosorbent assay (ELISA) were used to evaluate the infection status of the mouse models. Meanwhile, the nested PCR detection limit of B. microti was obtained using pure B. microti DNA samples with serial concentrations and whole blood samples with different densities of B. microti-infected red blood cells. Thereafter, whole mouse blood with a B. microti density lower than that of the nested PCR detection limit and human blood samples infected with B. microti were transfused into healthy mice to assess the transmission risk in mouse models. The infection status of these mice was evaluated through microscopic examination, nested PCR tests, and ELISA. Results: The mice inoculated with different densities of B. microti reached the peak infection rate on different days. Overall, the higher the blood B. microti density was, the earlier the peak infection rate was reached. The levels of specific antibodies against B. microti in the blood of the infected mice increased sharply during the first 30 days of infection, reaching a peak level at 60 days post-infection, and maintaining a high level thereafter. The nested PCR detection limits of B. microti DNA and parasite density were 3 fg and 5.48 parasites/µL, respectively. The whole blood containing an extremely low density of B. microti and human blood samples infected with B. microti could infect mice, confirming the transmission risk of transfusing blood with low-density B. microti. Conclusion: Whole blood containing extremely low density of B. microti poses a high transmission risk when transfused between mice and mice or human and mice, suggesting that Babesia detection should be considered by governments, hospitals, and disease prevention and control centers as a mandatory test before blood donation or transfusion.

Assessment of Babesia spp. prevalence in various domestic animals across Southern Punjab, Pakistan.

Parasitic diseases, notably babesiosis, exert a substantial impact on the global cattle industry, posing challenges to commerce, economies, and human health. This study, conducted in Southern Punjab, Pakistan, aimed to assess the prevalence of Babesia spp. across various livestock species using microscopic and PCR methods. A total of 180 blood samples (60 from each district) were systematically collected from apparently healthy animals, with 36 samples obtained from each domestic animal species, including camel, cattle, buffalo, goat, and sheep, noting that 12 samples were collected from each district for each animal species. Overall prevalence was determined to be 32.8% (59/180), with varying rates among species: 25.0% in cattle, 41.66% in buffalo, 30.55% in goats, 33.3% in sheep, and 33.3% in camels. Microscopic examination revealed slightly varied infection rates among large and small domestic animals (22.2%), while PCR results indicated a 32.8% overall infection rate in both large and small domestic animals, with no statistical significance. District-wise analysis showed regional variations, with Muzaffargarh recording a prevalence rate of 23.33% through microscopic examination, while Lodhran and Bahawalpur recorded 21.67%. PCR results revealed higher rates (38.33%, 26.67%, and 33.33%, respectively), underlining the importance of employing PCR for accurate detection. Examining ruminant types, large ruminants exhibited a 32.4% infection rate, while small domestic animals showed 33.3%, with no significant difference (p=0.897). District-wise prevalence showcased significant variation, with Muzaffargarh demonstrating a 25% prevalence, Lodhran 22%, and Bahawalpur 22%, through microscopic examination. PCR results displayed 38.33%, 27%, and 33.3%, respectively, with no statistical significance. Detailed analysis of individual districts highlighted variations in infection rates among camels, cattle, buffalo, goats, and sheep. The binomial test indicated significant differences through microscopic analysis (P=0.011) but non-significant variations through PCR (P=0.065), emphasizing the precision of PCR. Regional variations in prevalence, notably with Punjab exhibiting the highest frequency (33.87%) and KPK the lowest (13.24%), suggest potential influences from varying veterinary practices and environmental factors. This study underscores the pivotal role of PCR alongside microscopy for accurate babesiosis diagnosis. These findings contribute to the broader understanding of babesiosis prevalence, emphasizing the necessity of advanced molecular techniques for informed control measures.

Expression of sex-specific molecular markers by Babesia bovis gametes.

BACKGROUND: Bovine babesiosis caused by Babesia bovis is one of the most important tick-borne diseases of cattle in tropical and subtropical regions. Babesia bovis parasites have a complex lifecycle, including development within the mammalian host and tick vector. In the tick midgut, extracellular Babesia parasites transform into gametes that fuse to form zygotes. To date, little is known about genes and proteins expressed by male gametes. METHODS AND RESULTS: We developed a method to separate male gametes from in vitro induced B. bovis culture. Separation enabled the validation of sex-specific markers. Collected male gametocytes were observed by Giemsa-stained smear and live-cell fluorescence microscopy. Babesia male gametes were used to confirm sex-specific markers by quantitative real-time PCR. Some genes were found to be male gamete specific genes including pka, hap2, α-tubulin II and znfp2. However, α-tubulin I and ABC transporter, trap2-4 and ccp1-3 genes were found to be upregulated in culture depleted of male gametes (female-enriched). Live immunofluorescence analysis using polyclonal antibodies confirmed surface expression of HAP2 by male and TRAP2-4 by female gametes. These results revealed strong markers to distinguish between B. bovis male and female gametes. CONCLUSIONS: Herein, we describe the identification of sex-specific molecular markers essential for B. bovis sexual reproduction. These tools will enhance our understanding of the biology of sexual stages and, consequently, the development of additional strategies to control bovine babesiosis.

Molecular survey of canine tick-borne pathogens in ticks and stray dogs in Dhaka city, Bangladesh.

Molecular surveillance of canine tick-borne pathogens (TBPs) in Bangladesh has constantly been undervalued. Therefore, the emergence of new pathogens often remains undetected. This study aimed to screen tick-borne pathogens in stray dogs and ticks in the Dhaka metropolitan area (DMA). Eighty-five dog blood and 53 ticks were collected in six city districts of DMA from September 2022 to January 2023. The ticks were identified by morphology. Screening of TBPs was performed by polymerase chain reaction (PCR), followed by sequencing. The PCR assays were conducted to analyze the 18S rRNA (Babesia gibsoni, B. vogeli, and Hepatozoon canis), 16S rRNA (Anaplasma phagocytophilum, A. platys, and A. bovis), gltA (Ehrlichia canis and Rickettsia spp.), flagellin B (Borrelia spp.) and 16-23S rRNA (Bartonella spp.). Three tick species, Rhipicephalus sanguineus (50/53), R. microplus (1/53), and Haemaphysalis bispinosa (2/53), were identified. Babesia gibsoni (38 out of 85) and A. platys (7 out of 85) were detected in dog blood. In contrast, four pathogens, B. gibsoni (1 out of 53), B. vogeli (1 out of 53), H. canis (22 out of 53), and A. platys (1 out of 53), were detected in the ticks. However, the detection rates of TBPs in dog blood and ticks were not correlated in this study. The phylogenetic analyses suggested that a single genotype for each of the four pathogens is circulating in DMA. This study reports the existence of B. vogeli, H. canis, and A. platys in Bangladesh for the first time.

Tick-borne pathogens in questing adults Dermacentor reticulatus from the Eastern European population (north-eastern Poland).

Dermacentor reticulatus is tick species with an expanding geographical range in Europe, which creates the possibility of spreading microorganisms of significant veterinary and medical importance. The study aimed to investigate the prevalence and genetic diversity of Rickettsia spp., Babesia spp., Borrelia spp. and Anaplasma phagocytophilum in adult D. reticulatus ticks from the Eastern European population in the urban and the natural biotopes of north-eastern Poland. Microorganisms were detected by PCR and identified by DNA sequencing. The overall infection rate of at least one of the pathogens was 29.6%. The predominantly was Rickettsia spp. (27.1%) (with R. raoultii-9.1%) followed by Babesia spp. (2.4%) with B. canis (1.5%) as the most frequent. Based on 18S rRNA gene sequence, three B. canis genotypes were revealed. The prevalence of R. raoultii and B. canis was significantly higher in ticks from natural biotopes. The infection rates of B. afzelii and A. phagocytophilum were determined at 0.9% and 0.3%, respectively. Co-infections were detected in 3.8% of infected ticks. In diagnosing tick-borne diseases in humans, tick-borne lymphadenopathy should not be excluded. The prevalence of different genotypes of B. canis suggests differences in the clinical picture of canine babesiosis in the area.

Molecular surveillance of microbial agents from cattle-attached and questing ticks from livestock agroecosystems of Antioquia, Colombia.

Ticks are obligate ectoparasites and vectors of pathogens affecting health, agriculture, and animal welfare. This study collected ticks from the cattle and questing ticks of 24 Magdalena Medio Antioquia region cattle farms. Genomic DNA was extracted from the specimens (individual or pools) of the 2088 adult ticks collected from cattle and 4667 immature questing ticks collected from pastures. The molecular detection of Babesia, Anaplasma, Coxiella and Rickettsia genera was performed by polymerase chain reaction amplification and subsequent DNA sequencing. In a total of 6755 Rhipicephalus microplus DNA samples, Anaplasma marginale was the most detected with a frequency of 2% (Confidence Interval- CI 1.68-2.36), followed by Babesia bigemina with 0.28% (CI 0.16-0.44), Coxiella spp. with 0.15% (CI 0.07-0.27), and Rickettsia spp. with 0.13% (CI 0.06-0.25). Molecular analysis of the DNA sequences obtained from the tick samples revealed the presence of Coxiella-like endosymbiont and R. felis. These results demonstrated the diversity of microorganisms present in R. microplus ticks predominantly associated with cattle and questing ticks from livestock agroecosystems, suggesting their role as reservoirs and potential biological vectors of these microorganisms on the studied sites. Also, it emphasizes the need to combine acarological surveillance with clinical diagnoses and control strategies on regional and national levels.

Detection and quantification of Babesia bovis and Babesia bigemina using different target genes.

Molecular assays have been widely used for the detection and quantification of bovine babesiosis due to their high sensitivity and specificity. However, variations in the sensitivity of pathogen detection may occur depending on the selected target gene. Thus, this study aimed to compare the detection sensitivity (DS) of Babesia bovis and B. bigemina infection levels in artificially and naturally infected cattle using quantitative PCR (qPCR) and six target genes. For B. bovis, the merozoite surface antigen genes 2b and 2c (msa-2b and msa-2c), and the mitochondrial cytochrome b gene (cybmt) were used. For B. bigemina, the genes encoding the proteins associated with rhoptry 1c (rap-1c), rap-1a, and cybmt were used. Six bovines, free of babesiosis, were artificially infected with 1 × 10-8 red blood cells infected (iRBC) with B. bovis (n = 3) or 1 × 10-6B. bigemina iRBC (n = 3). The animals were evaluated daily until parasitemia was confirmed (≥ 2.0%). The quantity of iRBC present in each animal was determined by examining blood smears. Blood samples were then subjected to DNA extraction, serial dilution, and qPCR analysis to determine the DS of each target gene. In addition, 30 calves naturally infected by Babesia spp. were also evaluated using the same six target genes. Regarding the artificial infection, B. bovis cybmt showed 25-fold higher sensitivity than the msa-2b and msa-2c genes, while the B. bigemina cybmt exhibited 5-fold and 25-fold higher sensitivity than the rap-1a and rap-1c genes, respectively. The rap-1a gene was found to be 5 times more sensitive than the rap-1c gene, while the B. bovis msa-2b and msa-2c genes exhibited similar DS. The positive frequencies of naturally infected calves for the target cybmt, msa-2b, and msa-2c genes (B. bovis) were: 100%, 33.3% and 50%, while cybmt, rap-1a, and rap-1c genes (B. bigemina) were 90%, 83.3%, and 63.3%, respectively. This study may contribute to the selection of suitable genes for molecular monitoring of bovine babesiosis. Mitochondrial genes could be considered as an alternative to improve the sensitivity of B. bovis and B. bigemina detection using qPCR.

First report of dog ticks and tick-borne pathogens they are carrying in Malawi.

Ticks are vectors for transmitting tick-borne pathogens (TBPs) in animals and humans. Therefore, tick identification is necessary to understand the distribution of tick species and the pathogens they carry. Unfortunately, data on dog ticks and the TBPs they harbor in Malawi are incomplete. This study aimed to identify dog ticks and the TBPs they transmit in Malawi. One hundred thirty-two ticks were collected from 87 apparently healthy but infested domestic dogs in four districts of Malawi, which were pooled into 128 tick samples. The ticks were morphologically identified under a stereomicroscope using identification keys, and species identification was authenticated by polymerase chain reaction (PCR) through the amplification and sequencing of 12S rRNA and cytochrome c oxidase subunit I (CO1) genes. The tick species identified were Rhipicephalus sanguineus sensu lato (58.3%), Haemaphysalis elliptica (32.6%), and Hyalomma truncatum (9.1%). Screening for TBPs using species-specific PCR assays revealed that 48.4% of the ticks were infected with at least one TBP. The TBP detection rates were 13.3% for Anaplasma platys, 10.2% for Babesia rossi, 8.6% for B. vogeli, 6.3% for Ehrlichia canis, 3.9% for A. phagocytophilum, 3.1% for B. gibsoni, 2.3% for B. canis and 0.8% for Hepatozoon canis. Co-infections of up to three pathogens were observed in 48.4% of the positive samples. This is the first study to identify dog ticks and the TBPs they harbor in Malawi. These findings provide the basis for understanding dog tick distribution and pathogens they carry in Malawi. This study necessitates the examination of ticks from more study locations to have a better picture of tick challenge, and the development of ticks and tick-borne disease control methods in Malawi.

Tick-borne pathogens in camels: A systematic review and meta-analysis of the prevalence in dromedaries.

Published data on tick-borne pathogens (TBPs) in camels worldwide have been collected to provide an overview of the global prevalence and species diversity of camelid TBPs. Several TBPs have been detected in dromedary camels, raising concerns regarding their role as natural or maintenance hosts for tick-borne pathogens. Insubstantial evidence exists regarding the natural infection of camels with Babesia spp., Theileria spp., Anaplasma spp., and Ehrlichia spp., particularly because most of the camels were considered healthy at the time of sampling. Based on polymerase chain reaction (PCR) testing, a pooled prevalence of 35.3% (95% CI: 22.6-48.1%) was estimated for Anaplasma, which was the most frequently tested TBP in dromedaries, and DNA of Anaplasma marginale, Anaplasma centrale, Anaplasma ovis, Anaplasma platys, and A. platys-like were isolated, of which ruminants and dogs are reservoirs. Similarly, the estimated pooled prevalence for the two piroplasmid genera; Babesia and Theileria was approximately equal (10-12%) regardless of the detection method (microscopy or PCR testing). Nevertheless, Babesia caballi, Theileria equi, and Theileria annulata DNA have frequently been detected in camels but they have not yet been proven to be natural hosts. Scarce data detected Babesia microti, Anaplasma phagocytophilum, and Borrelia burgdorferi sensu lato (s.l.) DNA in blood of dromedaries, although ticks of the genus Ixodes are distributed in limited areas where dromedaries are raised. Interestingly, a pooled seroprevalence of 47.7% (26.3-69.2%) was estimated for Crimean-Congo hemorrhagic fever virus, and viral RNA was detected in dromedary blood; however, their contribution to maintain the viral transmission cycles requires further experimental investigation. The substantially low incidence and scarcity of data on Rickettsia and Ehrlichia species could imply that camels were accidentally infected. In contrast, camels may play a role in the spread of Coxiella burnetii, which is primarily transmitted through the inhalation of aerosols emitted by diseased animals and contaminated environments. Bactrian camels showed no symptoms due to the examined TBPs, meanwhile, clinical disease was seen in alpacas infected with A. phagocytophilum. Similar to dromedaries, accidental tick bites may be the cause of TBP DNA found in the blood of Bactrian camels.

Low prevalence of Babesia hongkongensis infection in community and privately-owned cats in Hong Kong.

Domestic cats are susceptible to infection with at least 11 species of Babesia. In Hong Kong, where dogs are commonly infected with B. gibsoni, a single infection in a cat by a novel species, B. hongkongensis, was reported previously. The aim of this study was to investigate the frequency of Babesia spp. detection in cats in Hong Kong. Residual blood-derived DNA from healthy free-roaming community cats (n = 239), and privately-owned cats with and without anaemia undergoing diagnostic investigations (n = 125) was tested for Babesia spp. DNA using a pan-Babesia PCR targeting mitochondrial Cytochrome B, and a B. hongkongensis specific PCR targeting 18S rRNA. Positive samples were confirmed by sequencing and comparative sequence analysis against the GenBank nucleotide database. Babesia hongkongensis was detected in 4/239 (1.7 %) community cats, and 0/125 (0.0 %) privately-owned cats. Babesia gibsoni was detected in 0/239 community cats and 1/125 (0.8 %) privately-owned cats. Cats infected with B. hongkongensis were clinically healthy at the time of sampling. The B. gibsoni-infected cat was anaemic and thrombocytopenic. Cats in Hong Kong can be infected with B. hongkongensis and B. gibsoni, albeit at low frequency. The tick vector for B. hongkongensis is yet to be identified.

Babesia negevi infection in dogs and response to treatment.

Canine babesiosis is an important protozoan tick-borne disease associated with anemia and thrombocytopenia and caused by several different Babesia spp. Babesia negevi was first reported to infect dogs in the Middle East in 2020. This study describes the presentation, clinical signs, parasitemia levels quantified by molecular techniques, laboratory findings and treatment of dogs infected with B. negevi following the first description of this species. Clinical findings in the infected dogs, a 3-year old female and two 8-week old male and female pups, included extreme lethargy and pale mucous membranes, anemia and thrombocytopenia found in all three animals. Fever was present in the older female and icterus in the female pup. Babesia parasites resembling B. negevi were detected by microscopy of blood smears from the dogs. PCR of blood targeting the 18S rRNA and cox1 genes confirmed that babesiosis was caused by B. negevi and PCR targeting the Borrelia flagellin gene indicated co-infection with Borrelia persica in two dogs. Treatment of the dogs with imidocarb dipropionate resulted in clinical improvement and initial decrease in the B. negevi parasite load as detected by quantitative PCR in two dogs, however the female pup continued to deteriorate and died. The parasite load in the 3-year old female decreased from 43,451 parasites/µl blood pre-imidocarb dipropionate treatment to 803 parasites/µl within two weeks. In the surviving pup, it decreased from 3,293,538 parasites/µl pre-treatment to 20,092 parasites/µl after two weeks. Babesia negevi DNA was still recovered from blood samples by PCR despite repeated treatment with imidocarb dipropionate one-month post-treatment in the surviving pup and up to seven months post-treatment in the 3-year old female. Only treatment with atovaquone and azithromycin for ten days eliminated B. negevi in both dogs as confirmed by negative PCR two weeks later. In conclusion, treatment with imidocarb dipropionate was helpful for recovery from clinical disease but did not facilitate parasite elimination, and it is therefore recommended to treat canine B. negevi infection with the combination of atovaquone and azithromycin.

First molecular detection of a novel Babesia species from Haemaphysalis hystricis in Taiwan.

Newly recorded ticks and emerging tick-borne pathogens have recently been reported in subtropical and tropical East Asia. In this study, a total of 1,615 ticks (259 Haemaphysalis hystricis, 1334 Rhipicephalus microplus, 19 H. flava, and 3 R. haemaphysaloides) were collected by flagging from vegetation in Taiwan during 2019-2021. All 1,615 captured tick samples tested negative for SFTSV and Borrelia, but 12 of 356 tick samples tested positive for PCR amplification of a fragment of the 18S rRNA gene of Babesia spp., with an infection rate of 3.37 % (12/356) and a minimum infection rate of 0.74 % (12/1,615). Among the 12 detected Babesia spp., 11 were identified as Babesia bigemina in R. microplus, and the other one, detected in H. hystricis, was classified as an unnamed novel Babesia sp. Interestingly, the 18S rRNA sequence from the isolate detected in H. hystricis shared 98.79 % to 99.50 % identity with those of recent isolates from Japan, China and Nigeria. The exact origin of the Babesia species is not known, but the findings highlight the importance of international cooperation and the exchange of information on ticks and tick-borne pathogens. This represents a rare report of a Babesia sp. identified in H. hystricis, a tick species that has been proposed as a novel vector for some Babesia spp. This study supports H. hystricis as a possible vector of Babesia spp.

Molecular detection of Babesia vesperuginis in bats from Lithuania.

Babesia vesperuginis is an intraerythrocytic protozoan parasite that circulates among bats and ticks in many countries worldwide. However, the distribution of B. vesperuginis in the Baltic region has not been studied. A total of 86 dead bats from eight different species were collected and screened for Babesia spp. using real-time PCR. Overall, 52.3% (45/86) of the bats were found positive for Babesia spp. The prevalence of Babesia spp. in different organs varied, with the highest prevalence observed in heart tissues (37.0%) and the lowest in liver tissues (22.2%). However, the observed differences in prevalence among organs were not statistically significant. Blood samples from 125 bats of nine different species were also analyzed for Babesia spp. prevalence using real-time PCR and nested PCR. The results showed a prevalence of 35.2% and 22.4%, respectively. Moreover, 28.3% (17/60) of the examined blood samples were confirmed positive for Babesia spp. through blood smear analysis. The total of 32 partial sequences of the 18S rRNA gene derived in this study were 100% identical to B. vesperuginis sequences from GenBank. In eight species of bats, Pipistrellus nathusii, Pipistrellus pipistrellus, Pipistrellus pygmaeus, Vespertilio murinus, Eptesicus nilssonii, Eptesicus serotinus, Myotis daubentonii and Nyctalus noctula, Babesia parasites were identified. In E. nilssonii, Babesia spp. was identified for the first time.

High-throughput screening of pathogens in Ixodes ricinus removed from hosts in Lombardy, northern Italy.

Ticks are important vectors of many pathogens in Europe, where the most impactful species is Ixodes ricinus. Recently, the geographical distribution of this tick species has been expanding, resulting in an increased risk of human exposure to tick bites. With the present study, we aimed to screen 350 I. ricinus specimens collected from humans and wild animals (mainly ungulates), to have a broader understanding of the tick-borne pathogens circulating in the Lombardy region, in northern Italy. To do so, we took advantage of a high-throughput real-time microfluidic PCR approach to screen ticks in a cost-effective and time-saving manner. Molecular analysis of the dataset revealed the presence of four genera of bacteria and two genera of protozoa: in ungulates, 77 % of collected ticks carried Anaplasma phagocytophilum, while the most common pathogen species in ticks removed from humans were those belonging to Borrelia burgdorferi sensu lato group (7.6 %). We also detected other pathogenic microorganisms, such as Rickettisa monacensis, Rickettsia helvetica, Neoehrlichia mikurensis, Babesia venatorum, and Hepatozoon martis. Besides, we also reported the presence of the pathogenic agent Borrelia miyamotoi in the area (1.4 % overall). The most common dual co-infection detected in the same tick individual involved A. phagocytophilum and Rickettsia spp. Our study provided evidence of the circulation of different tick-borne pathogens in a densely populated region in Italy.