Oocyte maturation in the sloth’s giant tick Amblyomma varium (Acari: Ixodidae) in an ecological context

The sloth’s giant tick Amblyomma varium Koch, which is a neotropical species that inhabits tropical rainforests, is the largest tick reported to date. The adult stage of this tick parasitizes mammals from the families Bradypodidae and Magalonychidae (Xenarthra) nearly exclusively. This study aimed to describe morphological and histological features of the reproductive system and the oocyte maturation process of this tick species. The ovary of A. varium is a long single tubular organ that is horseshoe-shaped, winding and arranged in the posterior part of the body. Two oviducts are connected to the ovary on each side; these thicken at certain region forming the uterus (common oviduct), followed by a muscular connecting tube, vagina and genital aperture. A large number of oocytes at different stages of development are attached to the ovary wall by the pedicel, as they reach maturity they are released into the ovary lumen and from there to the genital aperture. These oocytes develop simultaneously and asynchronically along the ovary. Amblyomma varium oocytes were classified into five development stages (i.e., I–V), and specific characteristics were observed; the processes of yolk and chorion deposition begin early in oocytes stage II, and oocytes V exhibit a very thick chorion and eggs of a large size. These characteristics are likely adaptations that enhance the survival and the reproductive success of this extremely host-specific tick, which is limited to a particular environment.     

Feral pigs as hosts for Amblyomma sculptum (Acari: Ixodidae) populations in the Pantanal,Mato Grosso do Sul,Brazil

The Pantanal in Brazil is the largest floodplain of the world. This ecosystem, rich in wildlife, has a large feral pig population. Such a large host biomass must have a strong influence on the parasite fauna. In this work, we evaluated the role of feral pigs in the maintenance of Amblyomma sculptum (formerly Amblyomma cajennense), the most prevalent tick species in the Pantanal. Tick infestations were evaluated on 243 feral pigs and their environment. The suitability of domestic pigs, representing their feral relatives, to A. sculptum adults and nymphs was assessed experimentally. Tick infestation of feral pigs was strongly associated with that of the environment: 96 and 97 % of the ticks, respectively, were A. sculptum. The infestation prevalence on this host species was close to 90 % in the dry season and 100 % in the wet season and mean infestation intensity was above 30 ticks in both seasons. Suitability of pigs as hosts for A. sculptum was shown by the high proportion of nymphs and female ticks found engorging on captured feral pigs and adequate biological parameters displayed by ticks from experimental infestations of domestic pigs. Other tick species on feral pigs, albeit in much lower numbers, were Amblyomma parvum and Ornithodorus rostratus. Results show that feral pigs feed a high proportion of the A. sculptum adults and nymphs in their territories and should be a target for tick-borne diseases studies. This is particularly relevant to public health because all the main tick species found on feral pigs are aggressive to humans as well.     

Amblyomma testudinarium infestation on a brown bear (Ursus arctos yesoensis) captured in Hokkaido,a northern island of Japan

The tick Amblyomma testudinarium Koch, 1844 (Acari: Ixodidae) is known as a vector of several pathogens such as Rickettsia tamurae and severe fever with thrombocytopenia syndrome (SFTS) virus. This tick species is present in many Asian countries, including Japan, where its distribution is limited to the warm areas of Kanto region and the southwestern region. The present study reports the recovery of a partially engorged A. testudinarium from a wild brown bear captured in Shari town, Hokkaido. In addition to morphological identification, the specimen was genetically characterized by the complete mitochondrial genome sequencing. The results showed that the length of the obtained mitogenome is 14,835 bp that encodes 13 protein-coding, two ribosomal RNA (rRNA) (12S and 16S), and 22 transfer RNA genes with two non-coding control regions. The phylogenetic analysis indicated that our sample clustered with A. testudinarium from Nara, Japan, but separated from A. testudinarium from China. Although the introduction of the tick through livestock transportation cannot be ruled out, the detection of A. testudinarium in Hokkaido prefecture, which is separated from the main island where A. testudinarium is present in the south, may suggest the introduction by migratory birds. This study provides important insights on the distribution and host range of A. testudinarium. This will be useful for the future taxonomic analysis of ticks based on the complete mitogenome sequencing. To our knowledge, this is the northernmost detection point of the tropical tick A. testudinarium.     

Avian Migrants Facilitate Invasions of Neotropical Ticks and Tick-Borne Pathogens into the United States

Migratory birds have the potential to transport exotic vectors and pathogens of human and animal health importance across vast distances. We systematically examined birds that recently migrated to the United States from the Neotropics for ticks. We screened both ticks and birds for tick-borne pathogens, including Rickettsia species and Borrelia burgdorferi. Over two spring seasons (2013 and 2014), 3.56% of birds (n = 3,844) representing 42.35% of the species examined (n = 85) were infested by ticks. Ground-foraging birds with reduced fuel stores were most commonly infested. Eight tick species were identified, including seven in the genus Amblyomma, of which only Amblyomma maculatum/Amblyomma triste is known to be established in the United States. Most ticks on birds (67%) were neotropical species with ranges in Central and South America. Additionally, a single Ixodes genus tick was detected. A total of 29% of the ticks (n = 137) and no avian blood samples (n = 100) were positive for infection with Rickettsia species, including Rickettsia parkeri, an emerging cause of spotted fever in humans in the southern United States, a species in the group of Rickettsia monacensis, and uncharacterized species and endosymbionts of unknown pathogenicity. No avian tick or blood samples tested positive for B. burgdorferi, the etiologic agent of Lyme disease. An extrapolation of our findings suggests that anywhere from 4 to 39 million exotic neotropical ticks are transported to the United States annually on migratory songbirds, with uncertain consequences for human and animal health if the current barriers to their establishment and spread are overcome.     

Topographical and numerical study of the idiosomal integumentary structures of the larva of four Neotropical species of <Emphasis Type="Italic">Amblyomma</Emphasis> Koch, 1844 (Acari: Ixodidae)

Integumentary structures of the larvae of Amblyomma longirostre (Koch, 1844), A. parvum Aragão, 1908, A. rotundatum Koch, 1844 and from three populations of A. cajennense (Fabricius, 1787) were studied using light microscopy. A new nomenclature for the localisation of the integumentary structures is proposed. Three types of integumentary structures were identified in the larval idiosoma of the four Amblyomma species: lyrifissures, small glands and large wax glands. These structures were observed isolated or associated over the entire idiosoma, except in the scutum, which lacked lyrifisures and large wax glands. Large wax glands were the most stable within and between the tick species, followed by lyrifissures and small glands. Small glands, although relatively stable, showed the highest number of numerical variations within and between the tick species. Even though there were intra-population variations in the topographical and numerical pattern of some integumentary structures of A. cajennense larvae, there was a definitive pattern for most of the specimens, as showed by the similar modal and mean numbers of integumentary structures per tick side. The patterns of lyrifissures, small glands and large wax glands showed little differences when compared between the four Amblyomma species; however, a few differences were well evident. These differences were sufficient to differentiate larvae of the four species. Thus, we expect that the study of integumentary structures on the larvae of other Amblyomma species will be useful in future taxonomic keys for the identification of Amblyomma larvae from the Neotropical region.     

Perianal Tick-Bite Lesion Caused by a Fully Engorged Female Amblyomma testudinarium

A perianal tick and the surrounding skin were surgically excised from a 73-year-old man residing in a southwestern costal area of the Korean Peninsula. Microscopically a deep penetrating lesion was formed beneath the attachment site. Dense and mixed inflammatory cell infiltrations occurred in the dermis and subcutaneous tissues around the feeding lesion. Amorphous eosinophilic cement was abundant in the center of the lesion. The tick had Y-shaped anal groove, long mouthparts, ornate scutum, comma-shaped spiracular plate, distinct eyes, and fastoons. It was morphologically identified as a fully engorged female Amblyomma testudinarium. This is the third human case of Amblyomma tick infection in Korea.     

Localization and Visualization of a Coxiella-Type Symbiont within the Lone Star Tick, Amblyomma americanum

A Coxiella-type microbe occurs at 100% frequency in all Amblyomma americanum ticks thus far tested. Using laboratory-reared ticks free of other microbes, we identified the Amblyomma-associated Coxiella microbe in several types of tissue and at various stages of the life cycle of A. americanum by 16S rRNA gene sequencing and diagnostic PCR. We visualized Amblyomma-associated Coxiella through the use of a diagnostic fluorescence in situ hybridization (FISH) assay supplemented with PCR-based detection, nucleic acid fluorescent staining, wide-field epifluorescence and confocal microscopy, and transmission electron microscopy (TEM). Specific fluorescent foci were observed in several tick tissues, including the midgut and the Malpighian tubules, but particularly bright signals were observed in the granular acini of salivary gland clusters and in both small and large oocytes. TEM confirmed intracellular bacterial structures in the same tissues. The presence of Amblyomma-associated Coxiella within oocytes is consistent with the vertical transmission of these endosymbionts. Further, the presence of the Amblyomma-associated Coxiella symbiont in other tissues such as salivary glands could potentially lead to interactions with horizontally acquired pathogens.     

Microbial community structure reveals instability of nutritional symbiosis during the evolutionary radiation of Amblyomma ticks

Mutualistic interactions with microbes have facilitated the adaptation of major eukaryotic lineages to restricted diet niches. Hence, ticks with their strictly blood‐feeding lifestyle are associated with intracellular bacterial symbionts through an essential B vitamin supplementation. In this study, examination of bacterial diversity in 25 tick species of the genus Amblyomma showed that three intracellular bacteria, Coxiella‐like endosymbionts (LE), Francisella‐LE and Rickettsia, are remarkably common. No other bacterium is as uniformly present in Amblyomma ticks. Almost all Amblyomma species were found to harbour a nutritive obligate symbiont, Coxiella‐LE or Francisella‐LE, that is able to synthesize B vitamins. However, despite the co‐evolved and obligate nature of these mutualistic interactions, the structure of microbiomes does not mirror the Amblyomma phylogeny, with a clear exclusion pattern between Coxiella‐LE and Francisella‐LE across tick species. Coxiella‐LE, but not Francisella‐LE, form evolutionarily stable associations with ticks, commonly leading to co‐cladogenesis. We further found evidence for symbiont replacements during the radiation of Amblyomma, with recent, and probably ongoing, invasions by Francisella‐LE and subsequent replacements of ancestral Coxiella‐LE through transient co‐infections. Nutritional symbiosis in Amblyomma ticks is thus not a stable evolutionary state, but instead arises from conflicting origins between unrelated but competing symbionts with similar metabolic capabilities.     

New foci of <Emphasis Type="Italic">Rhipicephalus microplus</Emphasis> in West Africa

The invasive character of Rhipicephalus microplus was observed in Benin, the second West-African country from which this ticks species has been collected after the initial confirmed record in Ivory Coast in 2007. A cross-sectional study was carried out in the Department of Mono to examine the presence of the tick R. microplus. The survey covered 9 herds (villages) in an agro-ecological zone inhabited by agro-pastoralists, including the State Farm of Kpinnou that imported Girolando cattle from Brazil. Almost 800 ticks were sampled from 36 cattle, on average four cattle per village. The morphological identification revealed ticks of two different genera: Rhipicephalus and Amblyomma. Rhipicephalus microplus was the only representative of the species previously known as Boophilus or blue ticks. Its taxonomic identity was confirmed molecularly by PCR–RFLP. A comparison was made with the situation of R. microplus in Brazil.     

Cattle ticks of the genera <Emphasis Type="Italic">Rhipicephalus</Emphasis> and <Emphasis Type="Italic">Amblyomma</Emphasis> of economic importance in Tanzania: distribution assessed with GIS based on an extensive field survey

In order to implement a robust integrated tick and tick-borne disease control programme in Tanzania, based on ecological and epidemiological knowledge of ticks and their associated diseases, a national tick and sero-surveillance study was carried out in all 21 regions of the mainland, as well as on Mafia Island, between 1998 and 2001. The current distributions of Rhipicephalus appendiculatus, R. pravus, Amblyomma variegatum, A. gemma, and A. lepidum are illustrated and discussed. Tick distribution maps were assessed using the Weights-of-Evidence method (WofE), and employing temperature, humidity, NDVI, rainfall, and land-cover predictive data. Ground-truthing was done to check correspondence both of the data employed in prediction with land-cover characteristics discerned in the field as well as of the surveyed and predicted tick distributions. Statistical methods were used to analyse associations of the tick species with their environment, cattle density, and other ticks. Except for R. appendiculatus, no appreciable changes were demonstrated in the predicted and observed tick distributions compared to the existing maps that originated in the 1950–1960s. Cattle density influenced the distribution of A. variegatum and, to a certain extent, of A. lepidum, but had no appreciable influence on the distribution of any of the other ticks discussed in this paper, neither did livestock movement. Distinct differences for environmental requirements where observed between different tick species within the same genus. The predictive maps of R. appendiculatus and R. pravus suggest their mutually exclusive distribution in Tanzania, and simultaneous statistical analysis showed R. pravus as a greater specialist. Of the three Amblyomma species, A. variegatum is the most catholic tick species in Tanzania, while both A. gemma and A. lepidum belong to the more specialized species. Despite dissimilar habitat preferences, all three Amblyomma spp. co-exist in central Tanzania, where very heterogeneous habitats may simultaneously satisfy the environmental requirements of all three species. The current study, conducted about 4 decades after the last major survey activities, has shown that changing livestock policies, unrestricted livestock movement and a continuous change in climatic/environmental conditions in Tanzania have brought about only limited changes in the distribution patterns of R. appendiculatus, R. pravus and the three Amblyomma species investigated. Whether this observation indicates a relative indifference of these ticks to environmental and/or climate changes allows room for speculation.     

Community-Based Control of the Brown Dog Tick in a Region with High Rates of Rocky Mountain Spotted Fever, 2012–2013

Rocky Mountain spotted fever (RMSF) transmitted by the brown dog tick (Rhipicephalus sanguineus sensu lato) has emerged as a significant public health risk on American Indian reservations in eastern Arizona. During 2003–2012, more than 250 RMSF cases and 19 deaths were documented among Arizona''s American Indian population. The high case fatality rate makes community-level interventions aimed at rapid and sustained reduction of ticks urgent. Beginning in 2012, a two year pilot integrated tick prevention campaign called the RMSF Rodeo was launched in a ∼600-home tribal community with high rates of RMSF. During year one, long-acting tick collars were placed on all dogs in the community, environmental acaricides were applied to yards monthly, and animal care practices such as spay and neuter and proper tethering procedures were encouraged. Tick levels, indicated by visible inspection of dogs, tick traps and homeowner reports were used to monitor tick presence and evaluate the efficacy of interventions throughout the project. By the end of year one, <1% of dogs in the RMSF Rodeo community had visible tick infestations five months after the project was started, compared to 64% of dogs in Non-Rodeo communities, and environmental tick levels were reduced below detectable levels. The second year of the project focused on use of the long-acting collar alone and achieved sustained tick control with fewer than 3% of dogs in the RMSF Rodeo community with visible tick infestations by the end of the second year. Homeowner reports of tick activity in the domestic and peridomestic setting showed similar decreases in tick activity compared to the non-project communities. Expansion of this successful project to other areas with Rhipicephalus-transmitted RMSF has the potential to reduce brown dog tick infestations and save human lives.     

Molecular basis of anticoagulant and anticomplement activity of the tick salivary protein Salp14 and its homologs

During feeding, a tick''s mouthpart penetrates the host''s skin and damages tissues and small blood vessels, triggering the extrinsic coagulation and lectin complement pathways. To elude these defense mechanisms, ticks secrete multiple anticoagulant proteins and complement system inhibitors in their saliva. Here, we characterized the inhibitory activities of the homologous tick salivary proteins tick salivary lectin pathway inhibitor, Salp14, and Salp9Pac from Ixodesscapularis in the coagulation cascade and the lectin complement pathway. All three proteins inhibited binding of mannan-binding lectin to the polysaccharide mannan, preventing the activation of the lectin complement pathway. In contrast, only Salp14 showed an appreciable effect on coagulation by prolonging the lag time of thrombin generation. We found that the anticoagulant properties of Salp14 are governed by its basic tail region, which resembles the C terminus of tissue factor pathway inhibitor alpha and blocks the assembly and/or activity of the prothrombinase complex in the same way. Moreover, the Salp14 protein tail contributes to the inhibition of the lectin complement pathway via interaction with mannan binding lectin–associated serine proteases. Furthermore, we identified BaSO₄-adsorbing protein 1 isolated from the tick Ornithodoros savignyi as a distant homolog of tick salivary lectin pathway inhibitor/Salp14 proteins and showed that it inhibits the lectin complement pathway but not coagulation. The structure of BaSO₄-adsorbing protein 1, solved here using NMR spectroscopy, indicated that this protein adopts a noncanonical epidermal growth factor domain–like structural fold, the first such report for tick salivary proteins. These data support a mechanism by which tick saliva proteins simultaneously inhibit both the host coagulation cascade and the lectin complement pathway.     

Genomic evidence for rod monochromacy in sloths and armadillos suggests early subterranean history for Xenarthra

Rod monochromacy is a rare condition in vertebrates characterized by the absence of cone photoreceptor cells. The resulting phenotype is colourblindness and low acuity vision in dim-light and blindness in bright-light conditions. Early reports of xenarthrans (armadillos, sloths and anteaters) suggest that they are rod monochromats, but this has not been tested with genomic data. We searched the genomes of Dasypus novemcinctus (nine-banded armadillo), Choloepus hoffmanni (Hoffmann''s two-toed sloth) and Mylodon darwinii (extinct ground sloth) for retinal photoreceptor genes and examined them for inactivating mutations. We performed PCR and Sanger sequencing on cone phototransduction genes of 10 additional xenarthrans to test for shared inactivating mutations and estimated the timing of inactivation for photoreceptor pseudogenes. We concluded that a stem xenarthran became an long-wavelength sensitive-cone monochromat following a missense mutation at a critical residue in SWS1, and a stem cingulate (armadillos, glyptodonts and pampatheres) and stem pilosan (sloths and anteaters) independently acquired rod monochromacy early in their evolutionary history following the inactivation of LWS and PDE6C, respectively. We hypothesize that rod monochromacy in armadillos and pilosans evolved as an adaptation to a subterranean habitat in the early history of Xenarthra. The presence of rod monochromacy has major implications for understanding xenarthran behavioural ecology and evolution.     

Ticks (Acari: Ixodidae) of the state of Amazonas,Brazil

The tick fauna of Brazil is currently composed by 72 species. The state of Amazonas is the largest of Brazil, with an area of ≈ 19% of the Brazilian land. Besides its vast geographic area, only 19 tick species have been reported for Amazonas. Herein, lots containing ticks from the state of Amazonas were examined in three major tick collections from Brazil. A total of 5933 tick specimens were examined and recorded, comprising 2693 males, 1247 females, 1509 nymphs, and 484 larvae. These ticks were identified into the following 22 species: Amblyomma cajennense sensu lato, Amblyomma calcaratum, Amblyomma coelebs, Amblyomma dissimile, Amblyomma dubitatum, Amblyomma geayi, Amblyomma goeldii, Amblyomma humerale, Amblyomma latepunctatun, Amblyomma longirostre, Amblyomma naponense, Amblyomma oblongoguttatum, Amblyomma ovale, Amblyomma rotundatum, Amblyomma scalpturatum, Amblyomma varium, Dermacentor nitens, Haemaphysalis juxtakochi, Ixodes cf. Ixodes fuscipes, Ixodes luciae, Rhipicephalus microplus, Rhipicephalus sanguineus sensu lato. Ticks were collected from 17 (27.4%) out of the 62 municipalities that currently compose the state of Amazonas. The following four species are reported for the first time in the state of Amazonas: A. coelebs, A. dubitatum, H. juxtakochi, and Ixodes cf. I. fuscipes. The only tick species previously reported for Amazonas and not found in the present study is Amblyomma parvum. This study provides a great expansion of geographical and host records of ticks for the state of Amazonas, which is now considered to have a tick fauna composed by 23 species. It is noteworthy that we report 1391 Amblyomma nymphs that were identified to 13 different species.     

Birds as hosts of ixodid ticks (Acarina,Ixodidae)

rophic relations between ixodid ticks of the world fauna parasitizing as adults on birds are analyzed. These ticks include 60 out of 650 species of the family Ixodidae and belong to the genera Ixodes, Haemaphysalis, and Amblyomma. Relations between birds and ticks of the genus Ixodes are most ancient. Colonial marine birds seem to be the first hosts of ixodid ticks in the class of birds. Trophic specialization of ticks is not associated with bird taxa, because in most cases no tick specificity is caused by either phylogenetic features of hosts, or specific environmental conditions.     

Microbial Population Analysis of the Salivary Glands of Ticks; A Possible Strategy for the Surveillance of Bacterial Pathogens

Ticks are one of the most important blood-sucking vectors for infectious microorganisms in humans and animals. When feeding they inject saliva, containing microbes, into the host to facilitate the uptake of blood. An understanding of the microbial populations within their salivary glands would provide a valuable insight when evaluating the vectorial capacity of ticks. Three tick species (Ixodes ovatus, I. persulcatus and Haemaphysalis flava) were collected in Shizuoka Prefecture of Japan between 2008 and 2011. Each tick was dissected and the salivary glands removed. Bacterial communities in each salivary gland were characterized by 16S amplicon pyrosequencing using a 454 GS-Junior Next Generation Sequencer. The Ribosomal Database Project (RDP) Classifier was used to classify sequence reads at the genus level. The composition of the microbial populations of each tick species were assessed by principal component analysis (PCA) using the Metagenomics RAST (MG-RAST) metagenomic analysis tool. Rickettsia-specific PCR was used for the characterization of rickettsial species. Almost full length of 16S rDNA was amplified in order to characterize unclassified bacterial sequences obtained in I. persulcatus female samples. The numbers of bacterial genera identified for the tick species were 71 (I. ovatus), 127 (I. persulcatus) and 59 (H. flava). Eighteen bacterial genera were commonly detected in all tick species. The predominant bacterial genus observed in all tick species was Coxiella. Spiroplasma was detected in Ixodes, and not in H. flava. PCA revealed that microbial populations in tick salivary glands were different between tick species, indicating that host specificities may play an important role in determining the microbial complement. Four female I. persulcatus samples contained a high abundance of several sequences belonging to Alphaproteobacteria symbionts. This study revealed the microbial populations within the salivary glands of three species of ticks, and the results will contribute to the knowledge and prediction of emerging tick-borne diseases.