A new Rickettsia honei-related genotype, two novel soft tick haplotypes and first records of three mite species associated with bats in Pakistan

Bats are well adapted to inhabit human settlements and are suitable reservoirs of a high number of vector-borne pathogens with veterinary-medical importance. Owing to these eco-epidemiological traits, the importance of studying bat ectoparasites is increasingly recognized. However, relevant molecular-phylogenetic data are missing from several countries of southern Asia, including Pakistan. In this study 11 ectoparasites, collected from bats in northern Pakistan, were analyzed morphologically and/or molecularly, phylogenetically from a taxonomic point of view. In addition, soft ticks were screened for pathogen DNA. Three mesostigmatid mite species were identified: Steatonyssus occidentalis evansi Micherdzinski, 1980 and Ancystropus taprobaniusTurk, 1950 from Rousettus leschenaultii (Desmarest 1820) and two specimens of Spinturnix americanus (Banks 1902) from Pipistrellus cf. javanicus (Gray 1838). Six soft tick (Carios vespertilionis Latreille, 1802) larvae were also removed from Scotophilus kuhlii Leach, 1821. Morphometric comparison revealed minor differences in comparison with C. vespertilionis larvae from Europe (i.e., narrower scutum and longer 4th posterolateral setae, while scutal length and shape index were not significantly different in this context). When molecularly analyzed, C. vespertilionis larvae from Pakistan showed the highest, 94.1% cox1 sequence identity with a sample from Kenya, while in their 16S rRNA gene these had the highest, 96.2-96.4% identity with samples from Europe and central Asia (northwestern China). These findings were confirmed in phylogenetic analyses. A further soft tick larva, collected from R. leschenaultii and therefore provisionally called Argas sp. "rousetti", yielded sequences with only 86.2% and 91% similarities in its cox1 and 16S rRNA genes, respectively, to the genetically closest species, A. boueti. Both Argas species belonged to the phylogenetic group of the subgenus Chiropterargas. Molecular screening of two C. vespertilionis larvae for a broad range of tick-borne pathogens yielded negative results. However, the larva of Argas sp. "rousetti" showed the presence of a spotted fever group (SFG) Rickettsia sp., which (based on four genetic markers) was closely related to two Rickettsia species reported from southeastern Asia (i.e., R. honei and Rickettsia sp. IG-1), but differed more significantly from other rickettsiae. In conclusion, the three mite species identified here are new records for their host species and for Pakistan. The present findings support that despite the observed genetic differences, C. vespertilionis in southern Asia (Pakistan) and Europe belong to the same species and share common ancestry with C. vespertilionis in east Africa (Kenya). The soft tick species, Argas sp. "rousetti" is most likely a not yet described species within the subgenus Chiropterargas Hoogstraal, 1955, because it was clearly separated from A. boueti Roubaud & Colas-Belcour, 1933 and A. confusus Hoogstraal, 1955 in the phylogenetic analysis (also taking into account that other known species of the subgenus are unlikely to occur in Pakistan). The novel Rickettsia-genotype from Argas sp. "rousetti", represents the first molecular identification of a Rickettsia sp. phylogenetically close to R. honei, and the first Rickettsia sp. from any bat-associated soft tick species in southern Asia

Transmission of Rickettsia raoultii and Rickettsia massiliae DNA by Dermacentor reticulatus and Rhipicephalus sanguineus (s.l.) ticks during artificial feeding

BACKGROUND : Tick-borne rickettsial pathogens are emerging worldwide and pose an increased health risk to both humans and animals. A plethora of rickettsial species has been identified in ticks recovered from human and animal patients. However, the detection of rickettsial DNA in ticks does not necessarily mean that these ticks can act as vectors for these pathogens. Here, we used artificial feeding of ticks to confirm transmission of Rickettsia massiliae and Rickettsia raoultii by Rhipicephalus sanguineus (sensu lato) and Dermacentor reticulatus ticks, respectively. The speed of transmission was also determined. METHODS : An artificial feeding system based on silicone membranes were used to feed adult R. sanguineus (s.l.) and D. reticulatus ticks. Blood samples from in vitro feeding units were analysed for the presence of rickettsial DNA using PCR and reverse line blot hybridisation. RESULTS : The attachment rate of R. sanguineus (s.l.) ticks were 40.4% at 8 h post-application, increasing to 70. 2% at 72 h. Rickettsia massiliae was detected in blood samples collected 8 h after the R. sanguineus (s.l.) ticks were placed into the in vitro feeding units. D. reticulatus ticks were pre-fed on sheep and subsequently transferred to the in vitro feeding system. The attachment rate was 29.1 % at 24 h post-application, increasing to 43.6 % at 96 h. Rickettsia raoultii was detected in blood collected 24 h after D. reticulatus was placed into the feeding units. CONCLUSIONS : Rhipicephalus sanguineus (s.l.) and D. reticulatus ticks are vectors of R. massiliae and R. raoultii, respectively. The transmission of R. massiliae as early as 8 h after tick attachment emphasises the importance of removing ticks as soon as possible to minimise transmission. This study highlights the relevance of in vitro feeding systems to provide insight into the vectorial capacity of ticks and the dynamics of tick-borne pathogen transmission.http://www.parasitesandvectors.comam2018Veterinary Tropical Disease

First broad-range molecular screening of tick-borne pathogens in Ixodes (Pholeoixodes) kaiseri, with special emphasis on piroplasms

Recently, the occurrence of Ixodes (Pholeoixodes) kaiseri has been reported for the first time in several European countries, but data on the molecular analysis of this hard tick species are still lacking. Therefore, in this study DNA extracts of 28 I. kaiseri (collected from dogs and red foxes in Germany, Hungary and Romania) were screened with reverse line blot hybridisation (RLB), PCR and sequencing for the presence of 43 tick-borne pathogens or other members of their families from the categories of Anaplasmataceae, piroplasms, rickettsiae and borreliae. Rickettsia helvetica DNA was detected in one I. kaiseri female (from a red fox, Romania), for the first time in this tick species. Six ticks (from red foxes, Romania) contained the DNA of Babesia vulpes, also for the first time in the case of I. kaiseri. Molecular evidence of R. helvetica and B. vulpes in engorged I. kaiseri does not prove that this tick species is a vector of the above two pathogens, because they might have been taken up by the ticks from the blood of foxes. In addition, one I. kaiseri female (from a dog, Hungary) harboured Babesia sp. badger type-B, identified for the first time in Hungary and Central Europe (i.e. it has been reported previously from Western Europe and China). The latter finding can be explained by either the susceptibility of dogs to Babesia sp. badger type-B, or by transstadial survival of this piroplasm in I. kaiseri

No evidence of African swine fever virus replication in hard ticks

African swine fever (ASF) is caused by African swine fever virus (ASFV), a tick-borne DNA virus. Soft ticks of the genus Ornithodoros are the only biological vectors of ASFV recognized so far. Although other hard ticks have been tested for vector competence, two commonly found tick species in Europe, Ixodes ricinus and Dermacentor reticulatus, have not been assessed for their vector competence for ASFV. In this study, we aimed to determine whether virus replication can occur in any of these two hard tick species (I. ricinus and/or D. reticulatus), in comparison with O. moubata (the confirmed vector), after feeding them blood containing different ASFV isolates using an improved in vitro system. DNA quantities of ASFV in these infected hard ticks were measured systematically, for 6 weeks in I. ricinus, and up to 8 weeks in D. reticulatus, and the results were compared to those obtained from O. moubata. There was evidence of virus replication in the O. moubata ticks. However, there was no evidence of virus replication in I. ricinus or D. reticulatus, even though viral DNA could be detected for up to 8 weeks after feeding in some cases. This study presents the first results on the possible vector competence of European hard (ixodid) ticks for ASFV, in a validated in vitro feeding setup. In conclusion, given the lack of evidence for virus replication under in vitro conditions, D. reticulatus and I. ricinus are unlikely to be relevant biological vectors of ASFV.http://www.elsevier.com/locate/ttbdishb201

Novel foci of Dermacentor reticulatus ticks infected with Babesia canis and Babesia caballi in the Netherlands and in Belgium

BACKGROUND : Autochthonous populations of Dermacentor reticulatus ticks in the Netherlands were discovered after fatal cases of babesiosis occurred in resident dogs in 2004. The presence of D. reticulatus in the Netherlands has also linked with the emergence of piroplasmosis in the resident horse population. The aim of this study was to put together results of continued surveillance of field sites and hosts for this tick in the Netherlands and also in Belgium and determine their infection status for Babesia and Theileria species. METHODS : Ticks were collected from the vegetation at 11 locations between 2011 and 2013. D. reticulatus ticks were also collected from different hosts between 2007 and 2013. Ticks were screened by PCR and reverse line blot (RLB). RESULTS : A total of 1368 D. reticulatus ticks were collected from 4 previously known field locations and from 5 new locations in the Netherlands and from 2 sites in Belgium (one old and one new location). A total of 855 ticks collected from 8 locations in the Netherlands and 2 locations in Belgium were tested. Fourteen ticks (1,64%) collected at 4 field locations (Dintelse Gorzen, Rozenburg, Slikken van de Heen and St. Philipsland) were positive for Babesia canis, whereas two ticks were positive for Babesia caballi, one tick in the Dintelse Gorzen in the Netherlands and one tick was found positive in De Panne in Belgium. A further 1092 D. reticulatus ticks were collected between 2007 and 2013 from 40 dogs (132 ticks), two ticks from two humans, 51 ticks from 15 horses, two ticks from two cats, one tick from a roe deer, whereas most ticks (904) were collected from cattle (n = 25). Ticks were found throughout the year on dogs in nearly all provinces of the Netherlands. None of the ticks collected from these hosts were infected. CONCLUSIONS : D. reticulatus is continuing its spread into novel areas. The finding that some autochthonous ticks are infected with B. canis and B. caballi poses a threat to the resident dog and horse population and justifies year-round tick control measures.Prof. Frederic Beugnet and Dr. Fabien Danlois of Merial are thanked for their continued interest and financial support for this study.http://www.parasitesandvectors.comam201

Allergenomics of the tick Ixodes ricinus reveal important α-Gal-carrying IgE-binding proteins in red meat allergy

Supplementary material: [http://cherry.chem.bg.ac.rs/handle/123456789/3864]This is the peer-reviewed version of the following article: Apostolovic, D.; Mihailovic, J.; Commins, S. P.; Wijnveld, M.; Kazimirova, M.; Starkhammar, M.; Stockinger, H.; Platts-Mills, T. A. E.; Cirkovic Velickovic, T.; Hamsten, C.; et al. Allergenomics of the Tick Ixodes Ricinus Reveals Important α-Gal–Carrying IgE-Binding Proteins in Red Meat Allergy. Allergy: European Journal of Allergy and Clinical Immunology 2020, 75 (1), 217–220. [https://doi.org/10.1111/all.13978

Tick-borne pathogens of zoonotic and veterinary importance in Nigerian cattle

Additional file 1: Multiple infections by tick-borne pathogens according to age classes and overall number of animals. (PDF 19 kb)BACKGROUND : Ticks and tick-borne diseases undermine cattle fitness and productivity in the whole of sub-Saharan Africa, including Nigeria. In this West African country, cattle are challenged by numerous tick species, especially during the wet season. Consequently, several TBDs are known to be endemic in Nigerian cattle, including anaplasmosis, babesiosis, cowdriosis and theilerioris (by Theileria mutans and Theileria velifera). To date, all investigations on cattle TBDs in Nigeria have been based on cytological examinations and/or on serological methods. This study aimed to ascertain the occurrence of tick-borne pathogens of veterinary and zoonotic importance in cattle in Nigeria using molecular approaches. METHODS : In October 2008, 704 whole blood samples were collected from indigenous cattle in the Plateau State, Nigeria. Analysis for tick-borne pathogens was conducted by means of PCR-based reverse line blotting (RLB) and sequencing targeting a panel of five genera of microorganisms (i.e. Babesia, Theileria, Anaplasma, Ehrlichia and Rickettsia spp.). RESULTS : In total, 561/704 (82.6 %) animals were found infected, with 465 (69.6 %) of them being infected by two or more microorganisms, with up to 77 possible combinations of pathogens detected. Theileria mutans was the most prevalent microorganism (66.3 %), followed by Theileria velifera (52.4 %), Theileria taurotragi (39.5 %), Anaplasma marginale (39.1 %), Anaplasma sp. (Omatjenne) (34.7 %), Babesia bigemina (7.9 %), Anaplasma centrale (6.3 %), Anaplasma platys (3.9 %), Rickettsia massiliae (3.5 %), Babesia bovis (2.0 %) and Ehrlichia ruminantium (1.1 %). Calves were found significantly less infected than juvenile and adult cattle. CONCLUSIONS : This study provides updated, molecular-based information on cattle TBDs in Nigeria. The molecular approach employed allowed the diagnosis of numerous positive cases including carrier statuses, multiple infections and novel pathogen detections within the indigenous cattle population. Moreover, the RLB method here described enabled the detection of veterinary agents not only pertaining to bovine health, including also those of zoonotic importance. The high prevalence recorded for T. mutans, T. velifera, A. marginale, T. taurotragi and Anaplasma sp. (Omatjenne), suggests they may be endemically established in Nigeria, whereas the lower prevalence recorded for other microorganisms (i.e. A. centrale and B. bovis) highlights a less stable epidemiological scenario, requiring further investigations.The UK’s Biotechnology and Biological Sciences Research Council (BBSRC) under the ‘Combating Infectious Diseases in Livestock for International Development’ (CIDLID) scheme, and the European Union’s Seventh Framework Program (FP7/2007–2013) under grant agreement n° 221948, Integrated Control of Neglected Zoonoses (ICONZ).http://www.parasitesandvectors.comam2016Veterinary Tropical Disease

Canine and ovine tick-borne pathogens in camels, Nigeria

AbstractIn April 2008, whole blood samples were collected from 36 dromedary camels in Sokoto, North-western Nigeria. Following PCR and reverse line blotting, twenty-two samples (61%) resulted positive for Ehrlichia/Anaplasma spp. and three (8%) for Theileria/Babesia spp., with three (8%) cases of co-infections being found. Both sequence and BLAST analyses identified Ehrlichia/Anaplasma spp. and Theileria/Babesia spp. positive cases as Anaplasma platys and Theileria ovis, respectively.This is the first report of the detection of A. platys and T. ovis in camels from sub-Saharan Africa. The epidemiological relevance of this finding is enhanced by the close living of these animals with both dogs and small ruminants. The high prevalence detected for A. platys suggests a possible role of camels as carriers of this infection

Novel Rickettsia raoultii strain isolated and propagated from Austrian Dermacentor reticulatus ticks

Abstract Background Continuous culture of tick cell lines has proven a valuable asset in isolating and propagating several different vector-borne pathogens, making it possible to study these microorganisms under laboratory conditions and develop serological tests to benefit public health. We describe a method for effective, cost- and labor-efficient isolation and propagation of Rickettsia raoultii using generally available laboratory equipment and Rhipicephalus microplus cells, further demonstrating the usefulness of continuous tick cell lines. R. raoultii is one of the causative agents of tick-borne lymphadenopathy (TIBOLA) and is, together with its vector Dermacentor reticulatus, emerging in novel regions of Europe, giving rise to an increased threat to general public health. Methods Dermacentor reticulatus ticks were collected in the Donau-Auen (Lobau) national park in Vienna, Austria. The hemolymph of ten collected ticks was screened by PCR-reverse line blot for the presence of rickettsial DNA. A single tick tested positive for R. raoultii DNA and was used to infect Rhipicephalus microplus BME/CTVM2 cells. Results Sixty-five days after infection of the tick-cell line with an extract from a R. raoultii-infected tick, we observed intracellular bacteria in the cultured cells. On the basis of microscopy we suspected that the intracellular bacteria were a species of Rickettsia; this was confirmed by several PCRs targeting different genes. Subsequent sequencing showed 99–100 % identity with R. raoultii. Cryopreservation and resuscitation of R. raoultii was successful. After 28 days identical intracellular bacteria were microscopically observed. Conclusions R. raoultii was successfully isolated and propagated from D. reticulatus ticks using R. microplus BME/CTVM2 cells. The isolated strain shows significant molecular variation compared to currently known sequences. Furthermore we show for the first time the successful cryopreservation and resuscitation of R. raoultii