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Background: The immunocompetence handicap hypothesis suggests that males with a higher testosterone level should be better at developing male secondary traits, but at a cost of suppressed immune performance. As a result, we should expect that males with an increased testosterone level also possess a higher parasite load. However, previous empirical studies aimed to test this prediction have generated mixed results. Meanwhile, the effect of testosterone level on parasite load in female hosts remains poorly known., Methods: In this study, we tested this prediction by manipulating testosterone level in Daurian ground squirrels (Spermophilus dauricus), a medium-sized rodent widely distributed in northeast Asia. S. dauricus is an important host of ticks and fleas and often viewed as a considerable reservoir of plague. Live-trapped S. dauricus were injected with either tea oil (control group) or testosterone (treatment group) and then released. A total of 10 days later, the rodents were recaptured and checked for ectoparasites. Fecal samples were also collected to measure testosterone level of each individual., Results: We found that testosterone manipulation and sex of hosts interacted to affect tick load. At the end of the experiment, male squirrels subjected to testosterone implantation had an averagely higher tick load than males from the control group. However, this pattern was not found in females. Moreover, testosterone manipulation did not significantly affect flea load in S. dauricus., Conclusions: Our results only lent limited support for the immunocompetence handicap hypothesis, suggesting that the role of testosterone on regulating parasite load is relatively complex, and may largely depend on parasite type and gender of hosts., (© 2024. The Author(s).)

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., (© 2024. The Author(s).)

Background: Amblyomma americanum, the lone star tick, is an aggressive questing species that harbors several pathogens dangerous to humans in the United States. The Southeast in particular has large numbers of this tick due to the combined suitable climate and habitats throughout the region. No studies have estimated the underlying distribution of the lone star tick across the state of Georgia, a state where it is the dominant species encountered., Methods: Ticks were collected by flagging 198 transects of 750 m 2 at 43 state parks and wildlife management areas across the state from March to July of 2022. A suite of climate, landscape, and wildlife variables were assembled, and a logistic regression model was used to assess the association between these environmental factors and the presence of lone star ticks and to predict the distribution of these ticks across the state., Results: A total of 59/198 (30%) transects sampled contained adult or nymph A. americanum, with the majority of transects containing these ticks (54/59, 91.5%) in forested habitats. The presence of A. americanum was associated with elevation, normalized difference vegetation index (NDVI) on January 1, isothermality, temperature seasonality, and precipitation in the wettest quarter. Vast regions of central, eastern, and southern coastal Georgia (57% of the state) were categorized as suitable habitat for the lone star tick., Conclusions: This study describes the distribution of the lone star tick across the state of Georgia at a finer scale than the current county-level information available. It identifies specific variables associated with tick presence and provides a map that can be used to target areas for tick prevention messaging and awareness., (© 2024. The Author(s).)

Background: In Europe, canine babesiosis is most frequently caused by Babesia canis and Babesia vogeli, and occasionally by Babesia gibsoni.. In Germany, B. canis is recognized as endemic. The aims of this study were to assess how often Babesia spp. infections were diagnosed in a commercial laboratory in samples from dogs from Germany, and to evaluate potential risk factors for infection., Methods: The database of the LABOKLIN laboratory was screened for Babesia spp.-positive polymerase chain reaction (PCR) tests for dogs for the period January 2007-December 2020. Sequencing was performed for positive tests from 2018 and 2019. Binary logistic regression analysis was performed to determine the effects of sex, season, and year of testing. Questionnaires were sent to the submitting veterinarians to obtain information on travel abroad, tick infestation, and ectoparasite prophylaxis of the respective dogs. Fisher's exact test was used to calculate statistical significance and P < 0.05 was considered statistically significant., Results: In total, 659 out of 20,914 dogs (3.2%) tested positive for Babesia spp. by PCR. Of 172 sequenced samples, B. canis was identified in 156, B. vogeli in nine, B. gibsoni in five, and B. vulpes in two. Season had a statistically significant impact on test results when summer/winter (1.6% tested positive) was compared to spring/autumn (4.7%), with peaks in April (5.2%) and October (7.4%) [P < 0.001, odds ratio (OR) = 3.16]. Sex (male 3.5%, female 2.8%; P = 0.012, OR = 1.49) and age (< 7 years old 4.0%, ≥ 7 years old 2.3%; P < 0.001, OR = 1.76) of the tested dogs also had a statistically significant effect. A statistically significant impact was demonstrated for observed tick attachment (P < 0.001, OR = 7.62) and lack of ectoparasite prophylaxis (P = 0.001, OR = 3.03). The frequency of positive Babesia spp. tests did not significantly differ between the 659 dogs that had never left Germany and the 1506 dogs with known stays abroad (P = 0.088)., Conclusions: The possibility of canine infection with B. canis needs to be especially taken into consideration in spring and autumn in Germany as the activity of the tick Dermacentor reticulatus, a potential vector for canine babesiosis, is highest in these seasons. Travel and importation of dogs are considered major factors associated with canine babesiosis in Germany. However, autochthonous Babesia spp. infections also occur in a considerable number of dogs in Germany., (© 2023. The Author(s).)

Background: Ticks are obligate bloodsucking parasites responsible for significant economic losses and concerns with human and animal health, mainly due to the transmission of pathogens. Entomopathogenic fungi have been intensively studied as an alternative strategy for tick control that can be used in combination with synthetic acaricides in the integrated management of ticks. Here, we investigated how the gut bacterial community of Rhipicephalus microplus is shaped after Metarhizium anisopliae treatment and how the tick susceptibility to the fungus is affected after disrupting gut bacterial microbiota. Methods: Partially engorged tick females were artificially fed with pure bovine blood or blood plus tetracycline. Two other groups received the same diet and were topically treated with M. anisopliae. The guts were dissected, and the genomic DNA was extracted 3 days after the treatment; the V3–V4 variable region of the bacterial 16S rRNA gene was amplified. Results: The gut of ticks that received no antibiotic but were treated with M. anisopliae exhibited lower bacterial diversity and a higher occurrence of Coxiella species. The Simpson diversity index and Pielou equability coefficient were higher in the gut bacterial community when R. microplus were fed with tetracycline and fungus-treated. Ticks from fungus-treated groups (with or without tetracycline) exhibited lower survival than untreated females. Previous feeding of ticks with the antibiotic did not change their susceptibility to the fungus. Ehrlichia spp. were not detected in the gueated groups. Conclusions: These findings suggest that myco-acaricidal action would not be impacted if the calf hosting these ticks is under antibiotic therapy. Moreover, the hypothesis that entomopathogenic fungi can affect the bacterial community in the gut of R. microplus engorged females is endorsed by the fact that ticks exposed to M. anisopliae exhibited a dramatic reduction in bacterial diversity. This is the first report of an entomopathogenic fungus affecting the tick gut microbiota. [ABSTRACT FROM AUTHOR]

Background: Over 60 years ago clinical patterns resembling tick-borne rickettsioses have been described for the first time in Kazakhstan. Since 1995 the incidence of clinical cases of tick-borne rickettsioses in humans seems to be rising but studies on epidemiological data regarding the occurring etiological agents, tick vector species, prevalence and distribution throughout Kazakhstan are still scarce to date. The aim of the study was molecular investigation of ticks for spotted-fever group rickettsiae in the endemic Kyzylorda region and the so far considered as non-endemic Almaty region. A total of 2341 ticks was collected in the two regions in Kazakhstan and sorted in 501 pools: Ixodes persulcatus (243); Dermacentor marginatus (129); Haemaphysalis punctata (104); Hyalomma asiaticum (17); Dermacentor reticulatus (3); and Rhipicephalus turanicus (5). Pools were tested for Rickettsia spp. using real-time PCR. For positive samples multilocus sequence typing (MLST) was performed., Results: The calculated minimum infection rate (MIR) for rickettsiae in the investigated ticks in Almaty region varied between 0.4-15.1% and 12.6-22.7% in the Kyzylorda region. At least four different Rickettsia species were identified in the two selected regions of Kazakhstan. Two of these are already known to science: Rickettsia raoultii and R. slovaca, the latter being reported for the first time in Almaty region One new form, "Candidatus R. yenbekshikazakhensis", was described by MLST of six gene fragments in Almaty region and one new genotype, "genotype R. talgarensis" was detected using three gene fragments., Conclusions: Kazakh physicians should be aware of rickettsioses after tick bites in both regions studied. Both, R. raoultii and R. slovaca should be included in the diagnostics. The role for human diseases has further to be investigated for the newly described rickettsiae, "Candidatus R. yenbekshikazakhensis" and "Genotype R. talgarensis".

Background: Coxiella bacteria were identified from various tick species across the world. Q fever is a zoonotic disease caused by the bacteria Coxiella burnetii that most commonly infects a variety of mammals. Non-mammalian hosts, such as birds, have also been reported to be infected with the pathogenic form of "Candidatus Coxiella avium". This research increases the list of tick species that have been found with Coxiella-like bacteria in Thailand., Methods: A total of 69 ticks were collected from 27 domestic fowl (Gallus gallus domesticus), 2 jungle fowl (Gallus gallus) and 3 Siamese firebacks (Lophura diardi) at 10 locations (provinces) in Thailand. Ticks were identified and PCR was used to amplify Coxiella bacteria 16S rRNA, groEL and rpoB genes from the extracted tick DNA. MEGA6 was used to construct phylogenetic trees via a Maximum Likelihood method., Results: The phylogenetic analysis based on the 16S rRNA gene showed that the Coxiella sequences detected in this study grouped in the same clade with Coxiella sequences from the same tick genus (or species) reported previously. In contrast, rpoB gene of the Coxiella bacteria detected in this study did not cluster together with the same tick genus reported previously. Instead, they clustered by geographical distribution (Thai cluster and Malaysian cluster). In addition, phylogenetic analysis of the groEL gene (the chaperonin family) showed that all Coxiella bacteria found in this study were grouped in the same clade (three sister groups)., Conclusions: To our knowledge, we found for the first time rpoB genes of Coxiella-like bacteria in Haemaphysalis wellingtoni ticks forming two distinct clades by phylogenetic analysis. This may be indicative of a horizontal gene transfer event.

Background: The blacklegged tick, Ixodes scapularis, transmits most vector-borne diseases in the US. It vectors seven pathogens of public health relevance, including the emerging human pathogen Anaplasma phagocytophilum. Nevertheless, it remains critically understudied compared to other arthropod vectors. Ixodes scapularis releases a variety of molecules that assist in the modulation of host responses. Recently, it was found that extracellular vesicles (EVs) carry several of these molecules and may impact microbial transmission to the mammalian host. EV biogenesis has been studied in mammalian systems and is relatively well understood, but the molecular players important for the formation and secretion of EVs in arthropods of public health relevance remain elusive. RabGTPases are among the major molecular players in mammalian EV biogenesis. They influence membrane identity and vesicle budding, uncoating, and motility. Methods: Using BLAST, an in silico pathway for EV biogenesis in ticks was re-constructed. We identified Rab27 for further study. EVs were collected from ISE6 tick cells after knocking down rab27 to examine its role in tick EV biogenesis. Ixodes scapularis nymphs were injected with small interfering RNAs to knock down rab27 and then fed on naïve and A. phagocytophilum-infected mice to explore the importance of rab27 in tick feeding and bacterial acquisition. Results: Our BLAST analysis identified several of the proteins involved in EV biogenesis in ticks, including Rab27. We show that silencing rab27 in I. scapularis impacts tick fitness. Additionally, ticks acquire less A. phagocytophilum after rab27 silencing. Experiments in the tick ISE6 cell line show that silencing of rab27 causes a distinct range profile of tick EVs, indicating that Rab27 is needed to regulate EV biogenesis. Conclusions: Rab27 is needed for successful tick feeding and may be important for acquiring A. phagocytophilum during a blood meal. Additionally, silencing rab27 in tick cells results in a shift of extracellular vesicle size. Overall, we have observed that Rab27 plays a key role in tick EV biogenesis and the tripartite interactions among the vector, the mammalian host, and a microbe it encounters. [ABSTRACT FROM AUTHOR]

Background: Rodents play an important role in the life cycle of ixodid and argasid ticks, particularly as hosts of larvae and nymphs. The great gerbil (Rhombomys opimus), the preferred prey item of several carnivores (e.g. the red fox and marbled polecat), is the dominant rodent species in the Gurbantunggut Desert in northwestern China. The aim of this study was to investigate tick species associated with different hosts in the habitat of great gerbils, including wildlife and livestock. Methods: During 2018–2023, ticks were removed from 326 great gerbils, two red foxes (Vulpes vulpes), three marbled polecats (Vormela peregusna), 35 pastured sheep (Ovis aries), and one long-eared desert hedgehog (Hemiechinus auritus) in the Gurbantunggut Desert. Ticks were identified according to standard morphological keys. Then, they were further analyzed by molecular and phylogenic methods based on two mitochondrial markers, 16S rDNA and cytochrome c oxidase subunit I (COI) genes. Results: A total of 889 ticks were collected, representing five species. These included Hyalomma asiaticum (n = 425: 24 larvae, 79 nymphs and 322 adults), Rhipicephalus turanicus (n = 153: 2 nymphs and 151 adults), Haemaphysalis erinacei (n = 298: 4 larvae, 7 nymphs and 287 adults), Ixodes acuminatus (n = 7: 4 nymphs and 3 adults) and Ornithodoros tartakovskyi (6 adults). Based on COI sequences, molecular and phylogenetic analyses showed that (i) I. acuminatus from great gerbils and marbled polecats clustered with I. acuminatus reported from Europe; (ii) O. tartakovskyi found in northwestern China belonged to an independent clade; (iii) Hy. asiaticum, R. turanicus and Ha. erinacei had 100% sequence identities to conspecific ticks sampled previously in China. Conclusions: The great gerbil is an important host for the developmental stages of I. acuminatus, O. tartakovskyi, Ha. erinacei, Hy. asiaticum and R. turanicus, thus supporting the life cycle of several tick species which, as adults, parasitize predators (red fox and marble polecat) as well as pastured sheep and hedgehogs in the Gurbantunggut Desert. Ixodes acuminatus and O. tartakovskyi were found for the first time on great gerbil and marbled polecat, respectively. [ABSTRACT FROM AUTHOR]

Background: The speed with which acaricides paralyze and kill ticks is relevant to impeding pathogen transmission. The objective of this study was to assess early-onset lotilaner effects on the motility and weights of Amblyomma americanum ticks collected from treated dogs. Methods: Twelve healthy dogs were randomized between two groups to receive either lotilaner (Credelio®) on Day 0 or to be sham treated. On Day 7, 25 male and 25 female A. americanum were placed under bandages, two on each flank of each dog. After 30 or 45 min, all unattached ticks were removed and T = 0 was set. At T = 2, 4, 8 and 24 h post attachment, 5 attached ticks removed from each bandage on each dog were weighed, assessed by blinded observers for righting ability and movement recorded. Results: After the infestation period significantly fewer treated than control dogs had 20 ticks attached (50.0% versus 91.7%, P = 0.0015). At 24 h post attachment, mean weights of ticks from treated dogs (males 1.69 mg; females 2.72) were significantly less than ticks from controls (males 2.66 mg; females 4.67) (Pmale = 0.0002; Pfemale < 0.0001). Mean tick weights from the treated group were significantly lower at 24 h than at earlier time points (Pmale < 0.0307; Pfemale = 0.0021). At 4 and 8 h, significantly fewer ticks from treated (14.3%, 0.0%, respectively) than from control dogs could right (73.3%, 70.0%) (P4h < 0.0001; P8h = 0.0024) (at 24 h, all ticks from treated dogs were dead), and distance moved was significantly less at all time points (P2h = 0.0413; P4h, P8h < 0.0001). Mean and maximum velocity of ticks from treated dogs were significantly lower, relative to controls, at 4 and 8 h (P ≤ 0.0001). Within the treated group, collected ticks had significantly lower mean and maximum velocities at 4 and 8 h compared to 2 h (Pmean < 0.0042; Pmax < 0.0194). Conclusion: The observed changes indicate that lotilaner may disrupt tick attachment. In ticks that attached, a progressive impairment of neuromuscular processes began within 2 h. Those irreversible changes could substantially reduce the risk of pathogen transmission from tick to host. [ABSTRACT FROM AUTHOR]

Bartonella bacilliformis has recently been described in Amblyomma scalpturatum, Amblyomma ovale and Rhipicephalus microplus collected from wild animals in the Peruvian region of Madre de Dios. In this communication, I will discuss the results of a recent study by del Valle-Mendoza et al. together with the B. bacilliformis epidemiology. Following my argumentation, I consider the presence of this microorganism in the above ticks improbable.

Background: A subset of Australians who have been bitten by ticks experience a complex of chronic and debilitating symptoms which cannot be attributed to the known pathogenic species of bacteria present in Australia. As a result, there has been a renewed effort to identify and characterise viruses in Australian terrestrial ticks. Recent transcriptome sequencing of Ixodes and Amblyomma ticks has revealed the presence of multiple virus sequences. However, without virus isolates our ability to understand the host range and pathogenesis of newly identified viruses is limited. We have established a successful method for high-throughput virus discovery and isolation in mosquitoes using antibodies to double-stranded RNA. In this study we sought to characterise five archival tick-borne viruses to adapt our virus discovery protocol for Australian ticks. Methods: We performed virus characterisation using a combination of bioinformatic sequence analysis and in vitro techniques including replication kinetics, antigenic profiling, virus purification and mass spectrometry. Results: Our sequence analysis of Nugget virus, Catch-me-Cave virus and Finch Creek virus revealed marked genetic stability in isolates collected from the same location approximately 30 years apart. We demonstrate that the Ixodes scapularis-derived ISE6 cell line supports replication of Australian members of the Flaviviridae, Nairoviridae, Phenuiviridae and Reoviridae families, including Saumarez Reef virus (SREV), a flavivirus isolated from the soft tick Ornithodoros capensis. While antibodies against double-stranded RNA could be used to detect replication of a tick-borne reovirus and mosquito-borne flavivirus, the tick-borne flaviviruses Gadgets Gully virus and SREV could not be detected using this method. Finally, four novel virus-like sequences were identified in transcriptome sequencing of the Australian native tick Ixodes holocyclus. Conclusions: Genetic and antigenic characterisations of archival viruses in this study confirm that three viruses described in 2002 represent contemporary isolates of virus species first identified 30 years prior. Our findings with antibodies to double-stranded RNA highlight an unusual characteristic shared by two Australian tick-borne flaviviruses. Finally, comparative growth kinetics analyses of Australian tick-borne members of the Flaviviridae, Nairoviridae, Phenuiviridae and Reoviridae families in ISE6 and BSR cells will provide a useful resource for isolation of Australian tick-borne viruses using existing cell lines. [ABSTRACT FROM AUTHOR]

Background: Adult, nymph, and larval Rhipicephalus sanguineus sensu lato infest dogs and thrive in premises including homes and kennels. Ticks emerge from hiding to seek and attach to dogs, engorge, then leave their hosts to hide then molt or oviposit. This study evaluated the effect of either external or systemic canine treatment on R. sanguineus s.l. populations in premises. Methods: Thirty-two dogs in eight kennels were divided into three groups; one group (eight dogs in two kennels) served as untreated controls; one group (12 dogs in three kennels) received oral fluralaner (25–56 mg/kg); and one group (12 dogs in three kennels) received topical flumethrin/imidacloprid impregnated collars. Treatments were administered once on day 0. Prior to treatment, R. sanguineus s.l. infestations were established in kennels holding dogs, by placing ticks every 2 weeks from day −84 through day −14. Kennel surfaces (walls and floors) were smooth except for uniform "hideouts" to permit precise off-host tick counting. Results: Control dog kennel mean tick counts (all life stages) increased from 737 ticks/kennel at day −7 to 2213 at day 63 when all control kennel dogs were acaricide-treated as a humane endpoint. Kennels housing dogs subsequently treated with systemic fluralaner had a mean of 637 counted ticks/kennel on study day −7 (7 days before treatment). One fluralaner treatment eliminated all premises ticks (100% reduction) by day 70, and these kennels remained tick-free through study completion (day 84). Kennels housing dogs subsequently treated with an external imidacloprid/flumethrin collar had a mean of 614 counted ticks/kennel at study day −7. Collar treatment reduced counts by 90% on day 63, with kennel tick counts climbing after this and ending the study with a 75% reduction. Systemic fluralaner treatment was significantly (P = 0.003) more effective at reducing engorged adult female tick counts than external imidacloprid/flumethrin treatment. Conclusions: Fluralaner treatment eliminated off-host R. sanguineus life stages in infested kennels by day 70 following treatment and was significantly more effective than imidacloprid/flumethrin collar treatment in reducing the premises population of engorged female ticks. Imidacloprid/flumethrin treatment did not eliminate premises tick populations, with populations increasing before the study end. [ABSTRACT FROM AUTHOR]

Background: Glutathione S-transferases (GSTs) are a superfamily of multifunctional enzymes in living organisms with metabolic and detoxification functions, which can detoxify exogenous and endogenous compounds and thereby reduce the damage caused by toxic substances to the body. Ticks are obligate blood-sucking ectoparasites that can transmit various pathogens, and the characterization of tick-derived GSTs may help improve current understanding of the molecular mechanism of tick resistance to insecticides. In this study, a novel GST gene, named HrGSTm1, was identified from Hyalomma rufipes., Methods: Sequence analysis was performed by using bioinformatics techniques. A prokaryotic expression system was used to obtain the recombinant expression protein rHrGSTm1. Detection of spatiotemporal expression patterns of target genes and their response to the toxicity of cyhalothrin on female H. rufipes was performed by using a quantitative PCR platform. The optimal enzymological parameters of rHrGSTm1 using glutathione as substrate were calculated. The antioxidant capacity of the recombinant protein was evaluated by DPPH• (1,1-Diphenyl-2-picrylhydrazyl radical 2,2-Diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl). Knockdown of the HrGSTm1 genes through RNA interference was used to analyze their effects on the physiological parameters of ticks. The changes in HrGSTm1 messenger RNA expression patterns under cypermethrin stress were analyzed., Results: The complementary DNA sequence of HrGSTm1 contained a 672-bp open reading frame, which potentially encoded 223 amino acids. The predicted molecular weight was 25.62 kDa, and the isoelectric point 8.22. HrGSTm1 is a Mu-class GST, belonging to the cytoplasmic GSTs with no signal peptide observed. The V max and K m of rHrGSTm1 were 3.367 ± 0.81 uM and 2.208 ± 0.76 uM, respectively, and its activities were dependent on different temperatures and pH conditions; the scavenging rate of rHrGSTm1 to DPPH• reached 76.4% at 1.25 mg/ml. Variable expressions of HrGSTm1 were observed under various treatment periods and in different tissues, with the highest appearing in eggs (analysis of variance [ANOVA], F (2, 9)  = 279.9, P < 0.0001) and Malpighian tubules (ANOVA, F (3, 12)  = 290.5, P < 0.0001). After knockdown of HrGSTm1, compared with the control group, the mortality in the treatment group was increased by 16.7%, the average oviposition rate decreased by 33.9%, the average engorged body weight decreased by 287.38 mg and egg weight decreased by 127.46 mg, although only the engorged body weight was significantly different (t-test, t (44)  = 2.886, P = 0.006). After exposure to three sublethal concentrations (LC 05 , LC 10 , LC 50 ) of cyhalothrin, the expression level of HrGSTm1 in the midgut, ovary and salivary gland was upregulated, whereas in Malpighian tubules, it showed a trend of upregulation at first and then downregulation, implying different functions during the detoxification in different tissues., Conclusions: In this study, a novel GST of the Mu-class was successfully isolated from H. rufipes and systematically subjected to bioinformatic analysis and recombination identification. The variation trend of HrGSTm1 expression level in different tissues suggests that the gene has different detoxification functions in different tissues. The potential function of this gene was analyzed to provide basic research for further investigation of its detoxification mechanism., (© 2023. The Author(s).)

Background: Migratory birds can cross geographical and environmental barriers and are thereby able to facilitate transmission of tick-borne pathogens both as carriers of infected ticks and as reservoirs of pathogenic microorganisms. Ixodes ricinus is one of the most abundant tick species in the Northern Hemisphere and a main vector of several Babesia species, some which pose a potential threat to human and animal health. At present only two cases of overt babesiosis in humans have so far been reported in Sweden. To better understand the potential role of birds as disseminators of zoonotic Babesia protozoan parasites, we investigated the presence of Babesia species in ticks removed from migratory birds. Methods: Ticks were collected from birds captured at Ottenby Bird Observatory, south-eastern Sweden, from March to November 2009. Ticks were molecularly identified to species, and morphologically to developmental stage, and the presence of Babesia protozoan parasites was determined by real-time PCR. Results: In total, 4601 migratory birds of 65 species were examined for tick infestation. Ticks removed from these birds have previously been investigated for the presence of Borrelia bacteria and the tick-borne encephalitis virus. In the present study, a total of 1102 ticks were available for molecular analysis of Babesia protozoan parasites. We found that 2.4% of the ticks examined, all I. ricinus, were positive for mammal-associated Babesia species. Out of all Babesia-positive samples, Babesia venatorum was the most prevalent (58%) species, followed by Babesia microti (38%) and Babesia capreoli (4.0%). B. venatorum and B. capreoli were detected in I. ricinus larvae, whereas B. microti was only present in I. ricinus nymphs. This supports the view that the two first-mentioned species are vertically (transovarially) transmitted in the tick population, in contrast to B. microti. The largest number of Babesia-infected ticks was removed from the common redstart (Phoenicurus phoenicurus) and European robin (Erithacus rubecula). Conclusions: This study reveals that Babesia protozoan parasites are present in ticks infesting migratory birds in south-eastern Sweden, which could potentially lead to the dissemination of these tick-borne microorganisms into new areas, thus posing a threat to humans and other mammals. [ABSTRACT FROM AUTHOR]

Background: Ticks are blood-sucking arthropods that can transmit diseases to humans and animals. These arthropods are the second most important vectors of pathogens. MicroRNAs are a class of conserved small noncoding RNAs that play regulatory roles in gene expression at the post-transcriptional level. Molting is an important biological process in arthropods. Research on the molting process is important for understanding tick physiology and control., Methods: Dual-luciferase reporter assays were used to assess the role of miRNA let-7 in ecdysteroid receptor (ECR) biology. The expression levels of ECR and let-7 were measured by real-time qPCR before and after tick molting. To explore the function of let-7 and ECR, we performed overexpression and knocking down of let-7 and RNAi of ECR in tick nymphs. The biological function of let-7 in molting was explored by injecting nymphs, ten days after engorgement, with let-7 agomir for overexpression and let-7 antagomir for knocking down. The rate of molting was then determined. ECR dsRNA was injected into ticks to evaluate the function of ECR by gene silencing. The expression of ECR and let-7 was measured using RT-qPCR. All data were analyzed using GraphPad Prism v.6., Results: The results of the luciferase assay using a eukaryotic expression system revealed that ECR was a natural target of let-7. Let-7 overexpressed by agomir affected the rate of molting (P < 0.01) and the period of molting (P < 0.01). Let-7 antagomir for knockdown affected the period of molting (P < 0.01), but there was no effect on the rate of molting (P = 0.27). ECR dsRNA gene silencing significantly affected the rate of molting (P < 0.05)., Conclusions: This study demonstrated that let-7 can regulate the expression of ECR and that let-7 can affect molting in ticks. Our results help to understand the regulation of let-7 by 20-hydroxyecdysone (20E) and will provide a reference for functional analysis studies of microRNAs in ticks.

Background: The Lone Star tick, Amblyomma americanum is important to human health because of a variety of pathogenic organisms transmitted to humans during feeding events, which underscores the need to identify novel approaches to prevent tick bites. Thus, the goal of this study was to test natural and synthetic molecules for repellent activity against ticks in spatial, contact and human fingertip bioassays. Methods: The efficacy of essential oils and naturally derived compounds as repellents to Am. americanum nymphs was compared in three different bioassays: contact, spatial and fingertip repellent bioassays. Results: Concentration response curves after contact exposure to 1R-trans-chrysanthemic acid (TCA) indicated a 5.6 μg/cm2 concentration required to repel 50% of ticks (RC50), which was five- and sevenfold more active than DEET and nootkatone, respectively. For contact repellency, the rank order of repellency at 50 μg/cm2 for natural oils was clove > geranium > oregano > cedarwood > thyme > amyris > patchouli > citronella > juniper berry > peppermint > cassia. For spatial bioassays, TCA was approximately twofold more active than DEET and nootkatone at 50 μg/cm2 but was not significantly different at 10 μg/cm2. In spatial assays, thyme and cassia were the most active compounds tested with 100% and 80% ticks repelled within 15 min of exposure respectively and was approximately twofold more effective than DEET at the same concentration. To translate these non-host assays to efficacy when used on the human host, we quantified repellency using a finger-climbing assay. TCA, nootkatone and DEET were equally effective in the fingertip assay, and patchouli oil was the only natural oil that significantly repelled ticks. Conclusions: The differences in repellent potency based on the assay type suggests that the ability to discover active tick repellents suitable for development may be more complicated than with other arthropod species; furthermore, the field delivery mechanism must be considered early in development to ensure translation to field efficacy. TCA, which is naturally derived, is a promising candidate for a tick repellent that has comparable repellency to commercialized tick repellents. [ABSTRACT FROM AUTHOR]

Background: The tick Amblyomma cajennense sensu stricto (A. cajennense s.s.) frequently parasitizes animals and humans in the Amazon biome, in addition to being a vector of Rickettsia amblyommatis. In the present study, we evaluated both the population dynamics of A. cajennense s.s. in a degraded area of the Amazon biome and the presence of rickettsial organisms in this tick population., Methods: The study was carried out in a rural area of the Santa Inês municipality (altitude: 24 m a.s.l.), Maranhão state, Brazil. Ticks were collected from the environment for 24 consecutive months, from June 2021 to May 2023. The region is characterized by two warm seasons: a rainy season (November-May) and a dry season (June-October). We characterized the temporal activity of A. cajennense s.s. on the vegetation by examining questing activity for each life stage (larvae, nymphs, adults [males and females]) in relation to the dry and rainy season. Ticks collected in this study were randomly selected and individually tested by a TaqMan real-time PCR assay that targeted a 147-bp fragment of the rickettsial gltA gene., Results: Overall, 1843 (62.4%) adults (52.6% females, 47.4% males), 1110 (37.6%) nymphs and 398 larval clusters were collected. All adult females and nymphs were morphologically identified as A. cajennense s.s. Larval activity was observed from April to December, with a peak from June to September (dry season); nymph abundance peaked from September to November (transition period between dry and rainy seasons); and adult ticks were abundant from October to May (spring/summer/early autumn). The infection rate by R. amblyommatis in A. cajennense s.s. ticks was at least 7% (7/99)., Conclusion: Our data suggest a 1-year generation pattern for A. cajennense s.s., with a well-defined seasonality of larvae, nymphs and adults in the Amazon biome. Larvae predominate during the dry season, nymphs are most abundant in the dry-rainy season transition and adults are most abundant in the rainy season. The presence of R. amblyommatis in adult ticks suggests that animals and humans in the study region are at risk of infection by this species belonging to the spotted fever group of Rickettsia., (© 2023. The Author(s).)

Background: Anaplasma phagocytophilum is characterized by a worldwide distribution and distinguished from other Anaplasmataceae by the broadest range of mammalian hosts and high genetic diversity. The role carnivores play in the life cycle of A. phagocytophilum in Europe is uncertain. Currently, only the red fox is considered a suitable reservoir host. In this study, we focused on native and invasive medium-sized carnivore species that live in sympatry and represent the most abundant species of wild carnivores in Poland., Methods: A total of 275 individual spleen samples from six carnivore species (Vulpes vulpes, Meles meles, Procyon lotor, Nyctereutes procyonoides and Martes spp.) were screened combining nested PCR and sequencing for A. phagocytophilum targeting a partial groEL gene with subsequent phylogenetic analysis inferred by the maximum likelihood method., Results: The DNA of A. phagocytophilum was detected in 16 of 275 individuals (5.8%). Eight unique genetic variants of A. phagocytophilum were obtained. All detected haplotypes clustered in the clade representing European ecotype I. Three variants belonged to the subclade with European human cases together with strains from dogs, foxes, cats, and wild boars., Conclusions: While carnivores might have a restricted role in the dissemination of A. phagocytophilum due to their relatively low to moderate infection rates, they hold significance as hosts for ticks. Consequently, they could contribute to the transmission of tick-borne infections to humans indirectly, primarily through tick infection. This underscores the potential risk of urbanization for the A. phagocytophilum life cycle, further emphasizing the need for comprehensive understanding of its ecological dynamics., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Canine babesiosis is a disease caused by a parasite of the genus Babesia which destroys red blood cells. Previous studies have shown the presence of Babesia vogeli in rural areas in Costa Rica using molecular techniques. The objective of the present study was to determine the seroprevalence and prevalence of B. vogeli in clinically healthy dogs and their ticks at the national level, both within and outside the Central Valley. Blood samples and ticks from 482 dogs were collected between June 2011 and May 2014, and analyzed by immunofluorescence assay (IFA) and real-time polymerase chain reaction (qPCR); two protocols of endpoint PCR and sequencing were used to confirm qPCR-positive samples. Seroprevalence of canine babesiosis of 5.3% (24/453) was determined at the national level, specifically 2.0% (5/253) within and 9.5% (19/200) outside the Central Valley, respectively. Real-time PCR determined a global prevalence of B. vogeli of 31.3% (125/400): 21.4% (47/220) within the Central Valley and 43.3% (78/180) outside the Central Valley. The endpoint PCR amplified only 10 of the 125 blood samples identified as positive in qPCR. One sample amplified by endpoint PCR was sequenced and identified as B. vogeli. Twelve canines were identified with past infections, seven canines with active infection, and 111 canines with early infection. Two species of ticks were found with B. vogeli: Rhipicephalus sanguineus sensu lato (n = 40) and Amblyomma ovale (n = 1). The prevalence of canine babesiosis at the national level, both within and outside the Central Valley, is reported here for the first time, determining the presence of the piroplasmid throughout the country, with a higher circulation of the agent outside the Central Valley. Only one species, B. vogeli, was detected in the blood of dogs and their ticks. Therefore, veterinarians should consider using qPCR to determine the presence of the parasite in blood donors and before starting treatment of vector-borne disease in dogs., (© 2021. The Author(s).)

Background: Babesia is a protozoal, tick-borne parasite that can cause life-threatening disease in humans, wildlife and domestic animals worldwide. However, in Southeast Asia, little is known about the prevalence and diversity of Babesia species present in wildlife and the tick vectors responsible for its transmission. Recently, a novel Babesia species was reported in confiscated Sunda pangolins (Manis javanica) in Thailand. To investigate the presence of this parasite in Singapore, we conducted a molecular survey of Babesia spp. in free-roaming Sunda pangolins and their main ectoparasite, the Amblyomma javanense tick., Methods: Ticks and tissue samples were opportunistically collected from live and dead Sunda pangolins and screened using a PCR assay targeting the 18S rRNA gene of Babesia spp. DNA barcoding of the cytochrome oxidase subunit I (COI) mitochondrial gene was used to confirm the species of ticks that were Babesia positive., Results: A total of 296 ticks and 40 tissue samples were obtained from 21 Sunda pangolins throughout the 1-year study period. Babesia DNA was detected in five A. javanense ticks (minimum infection rate = 1.7%) and in nine different pangolins (52.9%) located across the country. Phylogenetic analysis revealed that the Babesia 18S sequences obtained from these samples grouped into a single monophyletic clade together with those derived from Sunda pangolins in Thailand and that this evolutionarily distinct species is basal to the Babesia sensu stricto clade, which encompasses a range of Babesia species that infect both domestic and wildlife vertebrate hosts., Conclusions: This is the first report documenting the detection of a Babesia species in A. javanense ticks, the main ectoparasite of Sunda pangolins. While our results showed that A. javanense can carry this novel Babesia sp., additional confirmatory studies are required to demonstrate vector competency. Further studies are also necessary to investigate the role of other transmission pathways given the low infection rate of ticks in relation to the high infection rate of Sunda pangolins. Although it appears that this novel Babesia sp. is of little to no pathogenicity to Sunda pangolins, its potential to cause disease in other animals or humans cannot be ruled out., (© 2023. The Author(s).)

Background: Rhipicephalus (Boophilus) microplus (Canestrini, 1888), the Asian blue tick, is a highly invasive and adaptable ectoparasite. This tick species has successfully established itself in most regions of the world, with movement of cattle being a major driver for its spread. In the recent past, R. microplus ticks have been reported in three districts of Uganda. Information on its spread and distribution are vital in deepening our understanding of the ecological scenarios that lead to tick persistence and in the formulation of control strategies. This is especially important in the cattle-dense districts. Methods: We randomly collected tick specimens from 1,461cattle spread across seven cattle dense districts located in the Central, Karamoja and West Nile regions of Uganda from January to September 2020. The ticks were identified using standard morpho-taxonomic keys and the R. microplus tick species identities were confirmed by sequencing of the ITS2 region, 12S rRNA and 16S rRNA genes and phylogenetic analyses. Results: Adult ticks (n = 13,019) were collected from 1,461 cattle. Seventeen tick species were identified based on morpho-taxonomic keys and the majority (47.4%; n=6184) of these were R. appendiculatus. In total, 257 R. microplus ticks were found infesting cattle in 18 study sites in the districts of Amudat, Kaabong, Napak (Karamoja region) and Arua (West Nile region). The identity of R. microplus was confirmed using molecular technics. No R. microplus tick was recorded in the districts of Lyantonde and Nakaseke (Central region). Arua district accounted for 82.1% (n=211) of the R. microplus ticks recorded followed by Napak district at 16.3% (n=42), while Amudat and Kaabong districts accounted for 1.5% (n=4). Rhipicephalus microplus and R. decoloratus co-existed in 6 of the 13 study sites in Arua district, while in another 6 study sites, no R. decoloratus was recorded. In the Karamoja region districts R. decoloratus co-existed with R.microplus. Of the total 618 ticks belonging to four species of the subgenus Boophilus recorded in this study, R. decoloratus accounted for 50.04% (n=334), followed by R. microplus at 41.58% (n=257), R. geigyi at 2.75% (n=17) and R. annulatus at 1.61% (n=10). In the districts of Amudat, Kaabong and Napak, R. decoloratus was more dominant (76.1%; n=179) of the three Rhipicephalus (Boophilus) tick species recorded, followed by R. microplus (19.5%; n=46) and R. geigyi (4.2%; n=10). Contrariwise, R. microplus was more dominant (84%; n=211) in Arua district followed by R. decoloratus (10.7%; n=27), R. annulatus (3.9%; n=10) and R. geigyi (1.1%; n=3). Phylogenetic analyses of the ITS2 region, 12S rRNA and 16S rRNA genes revealed subgrouping of the obtained sequences with the previously published R. microplus sequences from other parts of the world. Conclusion: Rhipicephalus microplus ticks were found infesting cattle in four districts of Uganda. The inability to find R. decoloratus, an indigenous tick, from six sites in the district of Arua is suggestive of its replacement by R. microplus. Rhipicephalus microplus negatively affects livestock production, and therefore, there is a need to determine its distribution and to deepen the understanding of the ecological factors that lead to its spread and persistence in an area. [ABSTRACT FROM AUTHOR]

Background: Tick-borne diseases are common throughout Europe. Ticks transmit pathogens to the host while feeding and together with mosquitoes, they are major vectors of infectious agents worldwide. In recent years, there has been a marked increase in the incidence of tick-bite events and tick-borne disease in northwest Italy, but information on the prevalence of tick-borne pathogens in ticks removed from humans remains scarce. To fill this gap, we report here the prevalence of tick bites and tick-borne pathogens documented for humans in Piedmont, northwest Italy, in the 3-year period 2017–2019. Methods: Ticks attached to humans during 2017–2019 were collected from residents of urban and rural area by physicians and veterinarians working with local veterinary agencies. All ticks (n = 1290) were morphologically identified to the species level. A subset of ticks removed from children (age 0–18 years) and the elderly (> 70 years), both age groups considered to be at-risk populations, was screened by biomolecular analysis to detect pathogens (e.g. Rickettsia spp., Borrelia spp., Anaplasma spp.). Pathogen identity was confirmed by Sanger sequencing. Results: Ticks were taxonomically assigned to ten species of six genera (Amblyomma, Dermacentor, Haemaphysalis, Hyalomma, Ixodes and Rhipicephalus). Most belonged to the genus Ixodes: 1009 ticks (78.22%) were classified as Ixodes ricinus. A subset of 500 ticks collected from the two at-risk populations were subjected to PCR assay to determine the presence of Rickettsia spp., Borrelia spp., and Anaplasma spp. The overall prevalence of infection was 22.8% (n = 114; 95% confidence interval [CI]: 19.19–26.73%), meaning that at least one pathogen was detected: Rickettsia spp. (prevalence 15%, n = 76; 95% CI 12.17–18.65%); Borrelia spp. (prevalence 6.4%, n = 32; 95% CI 4.42–8.92%); and Anaplasma spp. (prevalence 1.2%, n = 6; 95% CI 0.44–2.6%). Conclusions: Our data underline the importance of surveillance in the epidemiology of tick-borne diseases and the implementation of strategies to control tick infestation and associated pathogens. [ABSTRACT FROM AUTHOR]

Background: The ixodid tick genera Rhipicephalus and Haemaphysalis contain several species of medical and/or veterinary importance, but their diversity in some regions of the world remains under-explored. For instance, very few modern studies have been performed on the taxonomy of these genera on the Arabian Peninsula. Methods: In this study, we trapped small mammals in the 'Asir Mountains of south-western Saudi Arabia and collected tick specimens for morphological examination and molecular barcoding, targeting three mitochondrial loci: cox1, 16S rRNA and 12S rRNA. Results: We obtained a total of 733 ticks (608 Haemaphysalis spp. and 125 Rhipicephalus spp.) from 75 small mammal hosts belonging to six species. All tick specimens were immature except for nine adults recovered from a hedgehog (Paraechinus aethiopicus). Morphologically, the Rhipicephalus ticks resembled R. camicasi, but the Haemaphysalis ticks showed differences in palp morphology compared with species previously described from Saudi Arabia. Phylogenetic analysis and automatic barcode gap discovery identified a novel clade of Rhipicephalus sp. representing most of the nymphs. This was most closely related to R. leporis, R. guilhoni and R. linnaei. The adult ticks and a small proportion of nymphs clustered with R. camicasi sequences from a previous study. Finally, the Haemaphysalis nymphs formed two distinct clades that were clearly separated from all reference sequences but closest to some African species. Conclusions: This apparent high level of tick diversity observed in a single study site of only ~ 170 km2, on a relatively small number of hosts, highlights the potential for the discovery of new tick species on the Arabian Peninsula. [ABSTRACT FROM AUTHOR]

Background: Lyme borreliosis (LB) is the most common vector-borne disease in Europe. Monitoring changes in the prevalence of different Borrelia species in ticks may be an important indicator of risk assessment and of differences in pathogenicity in humans. The objective of our study was to assess the prevalence, co-infection and distribution of Borrelia and Babesia species in ticks removed from humans in a large sample collected during a study period of 4 years. Methods: The ticks were collected throughout Poland from March to November over 4-year period from 2016 to 2019. All ticks (n = 1953) were morphologically identified in terms of species and developmental stage. Molecular screening for Borrelia and Babesia by amplification of the flagellin gene (flaB) or 18S rRNA marker was performed. Pathogen identity was confirmed by Sanger sequencing or PCR–restriction fragment length polymorphism analysis. Results: The ticks removed from humans in Poland during this study belonged to two species: Ixodes ricinus (97%) and Dermacentor reticulatus (3%). High Borrelia prevalence (25.3%), including B. miyamotoi (8.4%), was confirmed in Ixodes ricinus ticks removed from humans, as was the change in frequency of occurrence of Borrelia species during the 4-year study. Despite Babesia prevalence being relatively low (1.3%), the majority of tested isolates are considered to be pathogenic to humans. Babesia infection was observed more frequently among Borrelia-positive ticks (2.7%) than among ticks uninfected with Borrelia (0.8%). The most frequent dual co-infections were between Borrelia afzelii and Babesia microti. The presence of Borrelia was also confirmed in D. reticulatus (12.7%); however the role of these ticks in spirochete transmission to susceptible hosts is still unclear. Conclusions: Although the overall risk of developing LB after a tick bite is low in Europe, knowledge of the prevalence and distribution of Borrelia and Babesia species in ticks might be an important indicator of the risk of both these tick-borne diseases. [ABSTRACT FROM AUTHOR]

Background: The speed with which acaricides paralyze and kill ticks is relevant to impeding pathogen transmission. The objective of this study was to assess early-onset lotilaner effects on the motility and weights of Amblyomma americanum ticks collected from treated dogs. Methods: Twelve healthy dogs were randomized between two groups to receive either lotilaner (Credelio®) on Day 0 or to be sham treated. On Day 7, 25 male and 25 female A. americanum were placed under bandages, two on each flank of each dog. After 30 or 45 min, all unattached ticks were removed and T = 0 was set. At T = 2, 4, 8 and 24 h post attachment, 5 attached ticks removed from each bandage on each dog were weighed, assessed by blinded observers for righting ability and movement recorded. Results: After the infestation period significantly fewer treated than control dogs had 20 ticks attached (50.0% versus 91.7%, P = 0.0015). At 24 h post attachment, mean weights of ticks from treated dogs (males 1.69 mg; females 2.72) were significantly less than ticks from controls (males 2.66 mg; females 4.67) (Pmale = 0.0002; Pfemale < 0.0001). Mean tick weights from the treated group were significantly lower at 24 h than at earlier time points (Pmale < 0.0307; Pfemale = 0.0021). At 4 and 8 h, significantly fewer ticks from treated (14.3%, 0.0%, respectively) than from control dogs could right (73.3%, 70.0%) (P4h < 0.0001; P8h = 0.0024) (at 24 h, all ticks from treated dogs were dead), and distance moved was significantly less at all time points (P2h = 0.0413; P4h, P8h < 0.0001). Mean and maximum velocity of ticks from treated dogs were significantly lower, relative to controls, at 4 and 8 h (P ≤ 0.0001). Within the treated group, collected ticks had significantly lower mean and maximum velocities at 4 and 8 h compared to 2 h (Pmean < 0.0042; Pmax < 0.0194). Conclusion: The observed changes indicate that lotilaner may disrupt tick attachment. In ticks that attached, a progressive impairment of neuromuscular processes began within 2 h. Those irreversible changes could substantially reduce the risk of pathogen transmission from tick to host. [ABSTRACT FROM AUTHOR]

Background: Ixodes spp. are vectors of zoonotic pathogens. All three active life stages (larvae, nymphs, adults) need to feed on a host in order to develop. Usually ticks parasitize attached to the external surface of their hosts' skin. Interestingly, in some cases ticks can also be found in the subcutaneous tissue in a variety of hosts, such as red foxes (Vulpes vulpes), raccoon dogs (Nyctereutes procyonoides) and dogs. Methods: The visceral side of 126 red fox-furs from Germany was examined visually searching for ticks. The localization of ticks was recorded and assigned to ten specific body parts. Morphological identification of ticks was performed according to standardized taxonomic protocols. Ticks which could not be further identified were examined genetically via conventional PCR targeting the 16S rRNA and cox1 gene. Hematoxylin and eosin (H&E) staining was used for histopathological examination. Results: In 111 out of 126 (88.1%) examined coats, at least one tick was found in the subcutaneous tissue. A total of 1203 ticks were removed from the subcutaneous tissue. Well-preserved ticks could be identified based on morphological criteria, but most ticks were in a progressed state of decomposition. Here, morphological species identification was not successful. Also, PCR methods did not lead to a successful species identification. The following species and development stages were found by morphological identification: Ixodes ricinus (female, n = 289; male, n = 8; nymph, n = 1), I. hexagonus (female, n = 2), I. canisuga (female, n = 1). Male I. ricinus were found individually or copulating in pairs with females. Subcutaneous ticks were localized at three predominant affected body parts: ears, axillar and inguinal region. Histological examination of subcutaneous ticks revealed a granulomatous panniculitis. Conclusions: To the authors' knowledge, this is the first finding of highly prevalent subcutaneous ticks in red foxes from Germany. Subcutaneous location of ticks seems to be very common in red foxes and the rule rather than the exception. Deep embedment of longirostra and long feeding times of females seem to put the subcutaneous location in favor. Most foxes were infested in the inguinal area, where the skin is thin and less hairy. [ABSTRACT FROM AUTHOR]