Diversity of Haemaphysalis-associated piroplasms of ruminants in Central-Eastern Europe, Hungary.
ABSTRACT: Increasing numbers of genetic variants are being recognized among piroplasms, but the precise taxonomical status, the tick vector and the geographical range of several species or genotypes are still unknown. Bovine piroplasmosis was reported to re-emerge in north-east Hungary. Because Theileria-infection was newly diagnosed in one cattle herd in the same region of the country, the aim of this study was to molecularly identify the relevant agent, to find its local vector tick species, and to examine the range of Babesia/Theileria spp. of ruminants in Haemaphysalis sp. ticks collected previously in Hungary.Blood samples were drawn on two occasions from 90 dairy cattle in northern Hungary, and ticks were collected on their pastures. In addition, questing ticks (315 Haemaphysalis inermis, 259?H. concinna and 22?H. punctata), which originated mainly in the same region of the country from 2007, were included in the study. DNA was extracted from these samples, followed by molecular analysis for piroplasms. In the cattle Theileria orientalis was identified, with 100 % sequence homology to isolates from Japan, China, South-Africa and Australia. Based on GenBank data this genotype has not been previously reported in Europe. The prevalence of infection in the herd remained almost constant in the main tick season, suggesting exposure in previous years. Retrospective analysis of ticks revealed the presence of Babesia crassa in H. inermis, for the first time in Europe and in this tick species. On the other hand, H. concinna carried five different piroplasms, including B. motasi that was also newly detected in Central-Eastern Europe and in this tick species; whereas H. punctata harboured Theileria sp. OT3, hitherto known to occur in the Mediterranean region.Results of this study broaden the range of piroplasms that are infective for ruminants in Central-Eastern Europe. Although bovine babesiosis and theileriosis was known to occur in Hungary, molecular evidence is provided here for the first time on the presence of Babesia and/or Theileria spp. of sheep, goats and cervids in Hungary.
Project description:Natural landscape alterations as a consequence of urbanisation are one of the main drivers in the movements of wildlife into metropolitan and peri-urban areas. Worldwide, these wildlife species are highly adaptable and may be responsible for the transmission of tick-borne pathogens including piroplasms (Babesia, Theileria and Cytauxzoon spp.) that cause piroplasmosis in animals and occasionally in humans. Little is known about piroplasms in the ticks of urban wildlife in Australia. Ticks from long-nosed bandicoots (Perameles nasuta; n?=?71), eastern-barred bandicoots (Perameles gunnii; n?=?41), northern-brown bandicoots (Isoodon macrourus; n?=?19), southern-brown bandicoots (Isoodon obesulus; n?=?4), bandicoot sp. (n?=?2), flying foxes (Pteropus sp.; n?=?3), black rats (Rattus rattus; n?=?7), bush rats (Rattus fuscipes; n?=?4), brushtail possums (Trichosurus vulpecula; n?=?19), ringtail possums (Pseudocheirus peregrinus; n?=?12), short-eared possums (Trichosurus caninus; n?=?6), possum sp. (Trichosurus sp.; n?=?8), and red foxes (Vulpes vulpes; n?=?12) were analysed using piroplasm-specific 18S primers and Sanger sequencing. Seven Ixodes tasmani ticks from long-nosed bandicoots and bandicoots sp., three I. tasmani ticks and one Ixodes holocyclus tick from brushtail possums, and one Haemaphysalis longicornis tick from a red fox were positive for piroplasms. New genotypes, with sequences sharing 98% nucleotide similarities with Theileria sp. K1 detected in a burrowing bettong (Bettongia lesueur), were identified from bandicoot ticks. New genotypes were detected in ticks from brushtail possums, which shared 98% similarity with a Babesia sp. (JQ682877) previously identified in marsupials. Theileria orientalis was identified in the H. longicornis tick from the red fox. Babesia and Theileria spp. in the ticks parasitizing bandicoots and brushtail possums clustered closely with respective Babesia and Theileria clades derived from Australian marsupials. This represents the first detection of piroplasms in ticks parasitizing brushtail possums and a red fox in Australia.
Project description:Several new taxa belonging to the genus Francisella have been described recently. The present study describes the prevalence of Francisella tularensis and Francisella-like endosymbionts (FLE) in ticks collected from Hungary from 2007 to 2009 and characterizes the genetic variability of FLEs. A total of 5402 Ixodid ticks (Ixodes ricinus, I. acuminatus, Dermacentor marginatus, D. reticulatus, Haemaphysalis inermis, H. concinna, H. punctata) were collected from vegetation and animal hosts and tested with conventional PCR, detecting the 16S rRNA and tul4 genes. F. tularensis ssp. holarctica was found in 2 pools of H. concinna and 1 pool of D. reticulatus, both representing minimum prevalence (calculated with 1 infected tick per pool) of 0.27% whereas the sequences of a FLE were detected in 11 pools of D. reticulatus showing a minimum prevalence of 3%. Although the tul4 gene sequence of this FLE was identical to all Hungarian and Portuguese FLEs found earlier, and the 16S rRNA sequence was also identical to the sequence of the endosymbiont of D. reticulatus described in Bulgaria, these 16S rRNA gene coding sequences differed in 2 nucleotides from the one found earlier in this tick species in Hungary. This divergence may appear to be a minor difference between the sequences, potentially even resulting from a technical failure, but it could also indicate a significant difference stemming from the conservative genetic character of Francisellaceae. Thus, it raises a question about the number of FLE variants circulating in D. reticulatus in Europe and indicates the need for further data about the FLEs described in other parts of the continent and new FLE genotyping markers.
Project description:BACKGROUND:Free-living ungulates are hosts of ixodid ticks and reservoirs of tick-borne microorganisms in central Europe and many regions around the world. Tissue samples and engorged ticks were obtained from roe deer, red deer, fallow deer, mouflon, and wild boar hunted in deciduous forests of south-western Slovakia. DNA isolated from these samples was screened for the presence of tick-borne microorganisms by PCR-based methods. RESULTS:Ticks were found to infest all examined ungulate species. The principal infesting tick was Ixodes ricinus, identified on 90.4% of wildlife, and included all developmental stages. Larvae and nymphs of Haemaphysalis concinna were feeding on 9.6% of wildlife. Two specimens of Dermacentor reticulatus were also identified. Ungulates were positive for A. phagocytophilum and Theileria spp. Anaplasma phagocytophilum was found to infect 96.1% of cervids, 88.9% of mouflon, and 28.2% of wild boar, whereas Theileria spp. was detected only in cervids (94.6%). Importantly, a high rate of cervids (89%) showed mixed infections with both these microorganisms. In addition to A. phagocytophilum and Theileria spp., Rickettsia helvetica, R. monacensis, unidentified Rickettsia sp., Coxiella burnetii, "Candidatus Neoehrlichia mikurensis", Borrelia burgdorferi (s.l.) and Babesia venatorum were identified in engorged I. ricinus. Furthermore, A. phagocytophilum, Babesia spp. and Theileria spp. were detected in engorged H. concinna. Analysis of 16S rRNA and groEL gene sequences revealed the presence of five and two A. phagocytophilum variants, respectively, among which sequences identified in wild boar showed identity to the sequence of the causative agent of human granulocytic anaplasmosis (HGA). Phylogenetic analysis of Theileria 18S rRNA gene sequences amplified from cervids and engorged I. ricinus ticks segregated jointly with sequences of T. capreoli isolates into a moderately supported monophyletic clade. CONCLUSIONS:The findings indicate that free-living ungulates are reservoirs for A. phagocytophilum and Theileria spp. and engorged ixodid ticks attached to ungulates are good sentinels for the presence of agents of public and veterinary concern. Further analyses of the A. phagocytophilum genetic variants and Theileria species and their associations with vector ticks and free-living ungulates are required.
Project description:BACKGROUND:Apicomplexan tick-borne pathogens that cause disease in companion animals include species of Babesia Starcovici, 1893, Cytauxzoon Neitz & Thomas, 1948, Hepatozoon Miller, 1908 and Theileria Bettencourt, Franca & Borges, 1907. The only apicomplexan tick-borne disease of companion animals that is known to occur in Australia is babesiosis, caused by Babesia canis vogeli Reichenow, 1937 and Babesia gibsoni Patton, 1910. However, no molecular investigations have widely investigated members of Apicomplexa Levine, 1980 in Australian ticks that parasitise dogs, cats or horses, until this present investigation. RESULTS:Ticks (n = 711) removed from dogs (n = 498), cats (n = 139) and horses (n = 74) throughout Australia were screened for piroplasms and Hepatozoon spp. using conventional PCR and Sanger sequencing. The tick-borne pathogen B. vogeli was identified in two Rhipicephalus sanguineus Latreille ticks from dogs residing in the Northern Territory and Queensland (QLD). Theileria orientalis Yakimov & Sudachenkov, 1931 genotype Ikeda was detected in three Haemaphysalis longicornis Neumann ticks from dogs in New South Wales. Unexpectedly, the exotic tick-borne pathogen Hepatozoon canis James, 1905 was identified in an Ixodes holocyclus Neumann tick from a dog in QLD. Eight novel piroplasm and Hepatozoon species were identified and described in native ticks and named as follows: Babesia lohae n. sp., Babesia mackerrasorum n. sp., Hepatozoon banethi n. sp., Hepatozoon ewingi n. sp., Theileria apogeana n. sp., Theileria palmeri n. sp., Theileria paparinii n. sp. and Theileria worthingtonorum n. sp. Additionally, a novel cf. Sarcocystidae sp. sequence was obtained from Ixodes tasmani Neumann but could not be confidently identified at the genus level. CONCLUSIONS:Novel species of parasites in ticks represent an unknown threat to the health of companion animals that are bitten by these native tick species. The vector potential of Australian ticks for the newly discovered apicomplexans needs to be assessed, and further clinical and molecular investigations of these parasites, particularly in blood samples from dogs, cats and horses, is required to determine their potential for pathogenicity.
Project description:Canine babesiosis is an emerging or re-emerging disease caused by Babesia and Theileria protozoans, also called piroplasms, transmitted by Ixodid ticks. In Europe, four etiological agents have been identified to date, namely Babesia canis, B. vogeli, B. gibsoni and Theileria annae. France has a high prevalence of canine babesiosis and two tick species, Dermacentor reticulatus and Rhipicephalus sanguineus, are supposed to transmit B. canis and B. vogeli respectively. In southern France, where dog infections with B. vogeli were recently confirmed, no comprehensive study was performed to date on piroplasm species infecting dogs. Thus, a large scale survey involving veterinary clinics, kennels and tick collection from the environment was conducted from 2010 to 2012 in this area.From 2010 to 2012, 140 dog blood samples and 667 ticks were collected. All blood and a subset of ticks were screened for the presence of piroplasms by PCR amplification of 18S rDNA. B. vogeli, B. canis and T. annae were detected in 13.6, 12.9 and 0.7 % dogs respectively. B. vogeli and B. canis were detected in 10.5 % and in 1.6 % R. sanguineus ticks including 1.3 % co-infections. B. canis was the only species detected in D. reticulatus ticks (9.7 %). B. canis infections were only recorded in the southwest of France whereas B. vogeli was mainly found in the southeast. Finally, a significantly higher prevalence of B. vogeli infection was found in Gard compared to Corsica and Drôme regions, both in dogs (p < 0.002) and R. sanguineus ticks (p < 0.02) although R. sanguineus was the main ticks species removed from dogs in those three areas.The survey confirmed the circulation of both B. canis and B. vogeli in dogs in southern France with differences in distribution probably linked to the distribution of their respective vectors. It also showed differences in prevalence of B. vogeli infection in areas similar in terms of risk of dogs infestation with R. sanguineus. Further studies focusing on genetic and microbiota of R. sanguineus ticks should be conducted to explore other biological interactions that may explain the differences observed.
Project description:In this study 308 ticks (Ixodes ariadnae: 26 larvae, 14 nymphs, five females; I. vespertilionis: 89 larvae, 27 nymphs, eight females; I. simplex: 80 larvae, 50 nymphs, nine females) have been collected from 200 individuals of 17 bat species in two countries, Hungary and Romania. After DNA extraction these ticks were molecularly analysed for the presence of piroplasm DNA. In Hungary I. ariadnae was most frequently identified from bat species in the family Vespertilionidae, whereas I. vespertilionis was associated with Rhinolophidae. Ixodes ariadnae was not found in Romania. Four, four and one new bat host species of I. ariadnae, I. vespertilionis and I. simplex were identified, respectively. DNA sequences of piroplasms were detected in 20 bat ticks (15 larvae, four nymphs and one female). I. simplex carried piroplasm DNA sequences significantly more frequently than I. vespertilionis. In I. ariadnae only Babesia vesperuginis DNA was detected, whereas in I. vespertilionis sequences of both B. vesperuginis and B. crassa. From I. simplex the DNA of B. canis, Theileria capreoli, T. orientalis and Theileria sp. OT3 were amplified, as well as a shorter sequence of the zoonotic B. venatorum. Bat ticks are not known to infest dogs or ruminants, i.e. typical hosts and reservoirs of piroplasms molecularly identified in I. vespertilionis and I. simplex. Therefore, DNA sequences of piroplasms detected in these bat ticks most likely originated from the blood of their respective bat hosts. This may indicate either that bats are susceptible to a broader range of piroplasms than previously thought, or at least the DNA of piroplasms may pass through the gut barrier of bats during digestion of relevant arthropod vectors. In light of these findings, the role of bats in the epidemiology of piroplasmoses deserves further investigation.
Project description:Ticks and tick-borne diseases (TBDs) have a major economic impact on animal production worldwide. In the present study, 2410 ticks were collected from January to August 2017 from livestock and other domestic animals in North Kordofan and Kassala, Sudan, for species identification and investigation of <i>Rickettsia</i> spp. and piroplasms, either individually or as pools containing up to 10 ticks by molecular methods. In total, 13 tick species were identified by morphology and 16S rDNA sequencing. The most frequent tick species were <i>Hyalomma impeltatum</i> (24.90%), <i>Rhipicephalus evertsi evertsi</i> (18.84%), <i>Amblyomma lepidum</i> (16.06%), and <i>Rhipicephalus camicasi</i> (12.49%). A pan-<i>Rickettsia</i> real-time PCR revealed an overall minimum infection rate (MIR) with <i>Rickettsia</i> spp. of 5.64% (136 positive tick pools/2410 total ticks). <i>Rickettsia africae</i> and <i>Rickettsia aeschlimannii</i> were the most frequently identified species by sequencing. Furthermore, the following highly pathogenic livestock parasites were detected: <i>Theileria annulata</i>, <i>Theileria lestoquardi</i>, <i>Theileria equi</i>, and <i>Babesia caballi</i>. The present study documented <i>Rhipicephalus afranicus</i> as well as <i>Rickettsia conorii israelensis</i>, <i>Rickettsia massiliae</i>, and <i>Babesia pecorum</i> for the first time in Sudan. These findings are significant for the animal production sector as well as in terms of One Health, as the detected <i>Rickettsia</i> spp. can cause serious illness in humans.
Project description:Birds play an important role in short- and long-distance transportation of ticks and tick-borne pathogens. The aim of the present study was to provide comprehensive information on the species and genetic diversity of ixodid ticks transported by migratory and non-migratory bird species in Central Europe, and to evaluate relevant data in a geographical, as well as in an ecological context.During a three year period (2012-2014), altogether 3339 ixodid ticks were collected from 1167 passerine birds (representatives of 47 species) at ringing stations in Hungary. These ticks were identified, and the tick-infestations of bird species were compared according to various traits. In addition, PCR and sequencing of part of the cytochrome oxidase subunit-I (COI) and 16S rDNA genes were performed from representatives of five tick species.The most abundant tick species found were Ixodes ricinus and Haemaphysalis concinna (with 2296 and 989 immature stages, respectively). In addition, 48 I. frontalis (all stages), three Hyalomma rufipes nymphs, one I. lividus and two I. festai females were collected. The majority of I. ricinus and I. frontalis specimens occurred on ground-feeding bird species, as contrasted to Ha. concinna. Hy. rufipes showed the highest degree of sequence identity to an Ethiopian hybrid of the same tick species. Based on both COI and 16S rDNA gene analyses, two genetic lineages of I. frontalis were recognized (with only 91.4 % identity in their partial COI gene). These were highly similar to South-Western European isolates of the same tick species. Phylogenetic analysis of Ha. concinna specimens collected from birds in Hungary also revealed two genetic lineages, one of which showed high (?99 %) degree of 16S rDNA sequence identity to conspecific East Asian isolates.Two genetic lineages of I. frontalis and Ha. concinna are transported by birds in Central Europe, which reflect a high degree of sequence identity to South-Western European and East Asian isolates of the same tick species, respectively. In addition, I. festai was collected for the first time in Hungary. These findings highlight the importance of western and eastern migratory connections by birds (in addition to the southern direction), which are also relevant to the epidemiology of tick-borne diseases.
Project description:Babesiosis and Theileriosis are important worldwide-distributed tick-borne diseases for human and animals. Their presence in a particular area depends on the presence of suitable tick-vector and host species as well as competent reservoirs such as roe deer, one of the most abundant wild cervids in Spain. Spleen samples from 174 roe deer hunted in Spain were analysed to determine the prevalence of Babesia and Theileria species. DNA of both piroplasms was firstly detected using a commercial qPCR. Then, positive samples were molecularly characterized at the 18S rRNA and ITS1 genes of Babesia spp. and Theileria spp. The possible influence of some factors such as ecological area, age and sex was also assessed. Overall, 89.7% of roe deer were positive to any of the two piroplasms. Theileria spp. was more prevalent (60.9%) than Babesia spp. (19.0%); species identification could not be achieved in 17.3% of positive samples. Babesia prevalence was significantly higher in young animals and in roe deer from Oceanic regions, in contrast to Theileria spp. Five species were identified: Theileria sp. OT3 (60.3%), Babesia capreoli (15.5%), Babesia venatorum (2.9%), Theileria sp. 3185/02 (0.6%) and Babesia bigemina (0.6%). The coinfection B. capreoli/T. sp. OT3 was the most common (4.6%) followed by B. venatorum/T. sp. OT3 (0.6%) and B. bigemina/T. sp. OT3 (0.6%). Our results reveal that Theileria spp. and Babesia spp. are prevalent piroplasms in roe deer from Spain. These cervids can act as reservoirs for several Babesia and Theileria species, including the zoonotic B. venatorum. This study represents the first description of B. venatorum and B. bigemina in roe deer from Spain.
Project description:Babesiosis is an emerging and potentially zoonotic disease caused by tick-borne piroplasmids of the Babesia genus. New genetic variants of piroplasmids with unknown associations to vectors and hosts are recognized. Data on the occurrence of Babesia spp. in ticks and wildlife widen the knowledge on the geographical distribution and circulation of piroplasmids in natural foci. Questing and rodent-attached ticks, rodents, and birds were screened for the presence of Babesia-specific DNA using molecular methods. Spatial and temporal differences of Babesia spp. prevalence in ticks and rodents from two contrasting habitats of Slovakia with sympatric occurrence of Ixodes ricinus and Haemaphysalis concinna ticks and co-infections of Candidatus N. mikurensis and Anaplasma phagocytophilum were investigated.Babesia spp. were detected in 1.5 % and 6.6 % of questing I. ricinus and H. concinna, respectively. Prevalence of Babesia-infected I. ricinus was higher in a natural than an urban/suburban habitat. Phylogenetic analysis showed that Babesia spp. from I. ricinus clustered with Babesia microti, Babesia venatorum, Babesia canis, Babesia capreoli/Babesia divergens, and Babesia odocoilei. Babesia spp. amplified from H. concinna segregated into two monophyletic clades, designated Babesia sp. 1 (Eurasia) and Babesia sp. 2 (Eurasia), each of which represents a yet undescribed novel species. The prevalence of infection in rodents (with Apodemus flavicollis and Myodes glareolus prevailing) with B. microti was 1.3 % in an urban/suburban and 4.2 % in a natural habitat. The majority of infected rodents (81.3 %) were positive for spleen and blood and the remaining for lungs and/or skin. Rodent-attached I. ricinus (accounting for 96.3 %) and H. concinna were infected with B. microti, B. venatorum, B. capreoli/B. divergens, Babesia sp. 1 (Eurasia), and Babesia sp. 2 (Eurasia). All B. microti and B. venatorum isolates were identical to known zoonotic strains from Europe. Less than 1.0 % of Babesia-positive ticks and rodents carried Candidatus N. mikurensis or A. phagocytophilum.Our findings suggest that I. ricinus and rodents play important roles in the epidemiology of zoonotic Babesia spp. in south-western Slovakia. Associations with vertebrate hosts and the pathogenicity of Babesia spp. infecting H. concinna ticks need to be further explored.