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PubMed2005

Antigens from the midgut membranes of Ornithodoros erraticus induce lethal anti-tick immune responses in pigs and mice.

Ornithodoros erraticus is an argasid tick that can transmit severe diseases such as human relapsing fever and African swine fever. In southern Europe O. erraticus lives in close association with swine on free-range pig farms. Application of acaricides for the eradication of O. erraticus from pig farms is inefficient. This is the reason why we tried to develop an anti-O. erraticus vaccine as alternative method of control. Accordingly, we were prompted to investigate the protective possibilities of a midgut membrane extract from the parasite (GME) that has not been studied hitherto. Administration of the GME with Freund's adjuvants (FAs) to pigs and mice induced a protective response able to kill 80% of the immature forms of the parasite in the first 72 h post-feeding and to reduce the fecundity of females by more than 50%. The action of the vaccine is the result of damage to the midgut wall of the argasid, and, in mice, it has been shown that this damage is mediated by activation of the complement system. In pigs, the administration of GME with alum, instead of with FAs, reduced the degree of protection. The protective antigens of the GME were expressed by all the developmental stages examined and are probably proteins from the luminal membrane of midgut epithelial cells. These antigens were seen to be more abundant in recently fed parasites than in fasting specimens, suggesting that their expression is induced after blood ingestion.

African Swine Fever↗

PubMed2006

The geographic distribution of the putative agent of epizootic bovine abortion in the tick vector, Ornithodoros coriaceus.

Epizootic bovine abortion (EBA), also known as "foothill abortion", is a vector borne disease of beef cattle that graze in the mountainous regions of California, southern Oregon and western Nevada transmitted by the argasid tick Ornithodoros coriaceus. Recently, the putative agent of EBA was identified as a novel Deltaproteobacter in the order Myxococcales. In this study, a TaqMan real-time PCR (TM-PCR) protocol specific to the putative EBA agent was developed. The new real-time TM-PCR assay functioned sensitively and specifically to detect pathogen DNA in field-collected O. coriaceus ticks. The assay had an analytical sensitivity of a single plasmid copy and, when evaluated with a collection of tick-borne pathogens, yielded a positive PCR-result only for the agent of EBA. Use of the TM-PCR represents an effective tool for rapid and highly sensitive assessment of environmental risk and spatial and statistical analysis to highlight areas where there may be increased risk for EBA in susceptible cattle.

Abortion, Veterinary↗

PubMed2006

Ecdysteroid hormone titer and its relationship to vitellogenesis in the soft tick, Ornithodoros moubata (Acari: Argasidae).

A blood meal is required for reproduction in most argasid female ticks. The blood meal appears to stimulate an organ in the posterior end to produce a fat body stimulating factor (FSF), which is thought to be an ecdysteroid, to induce vitellogenin (Vg) synthesis. In this study, the relationship of vitellogenesis and ecdysteroids was investigated by measuring Vg and ecdysteroid titers while observing oocyte development and oviposition in mated and virgin females. Oviposition occurred from day 10 after engorgement in mated females and continued up to 40-50 days, whereas egg maturation and oviposition did not occur in virgin females. Vg titers in the hemolymph peaked on day 6 after engorgement and subsequently declined in mated females. Interestingly, Vg synthesis occurred and ovarian development progressed to the development of early vitellogenic oocytes in virgin females but oocyte maturation and oviposition did not occur. Topical application of ecdysteroids induced oviposition in fed virgin females indicating that ecdysteroids may induce oviposition. Concentrations of ecdysteroids for 20 days after engorgement revealed several peaks in mated female whole body extracts, but no peaks in virgin female extracts. In the hemolymph of only mated females, ecdysteroid titers showed two peaks that followed the early peak of ecdysteroids in the whole body on day 4 and 6 after engorgement. In addition, ecdysteroids in the reproductive tissues increased with the development of the ovary in mated females and this increase coincided with the latter peaks of the whole body. These observations indicate that physiological elevation of ecdysteroids accelerate Vg synthesis, and may induce egg maturation and stimulate oviposition in fed mated Ornithodoros moubata females.

Animals↗

PubMedDate not supplied

Identification of extrinsic blood coagulation pathway inhibitors from the tick Ornithodoros savignyi (Acari: Argasidae).

The salt BaSO(4) selectively adsorbs two proteins from crude Ornithodoros savignyi salivary gland extract. They co-purify during reversed-phase HPLC, but can be separated by hydrophobic-interaction chromatography. Their molecular masses are 9333 and 9173Da. The 9.3kDa protein was designated BSAP1 and the 9.1kDa protein BSAP2. Their amino acid compositions show significant differences, in particular the presence of seven and eight cysteine residues in BSAP1 and BSAP2, respectively. The proteins do not contain gamma-carboxyglutamic acid, hydroxyproline, or hydroxylysine. The proteins do not inhibit the intrinsic coagulation cascade, but inhibit the extrinsic pathway. The observed inhibition is not due to inhibition of factor VII. Both proteins bind to membranes. BSAP1 binds neutral and negatively charged membranes more strongly than BSAP2. Its affinity for negative membranes is, however, much lower than for neutral membranes. In contrast, BSAP2 binds both membranes equally strongly. The binding of the proteins to the membranes was significantly lowered upon pre-incubation with Ca(2+).

Adsorption↗

PubMed2002

Pathogenic mechanisms of sand tampan toxicoses induced by the tick, Ornithodoros savignyi.

The tick, Ornithodoros savignyi has been implicated in inducing paralysis and tampan toxicosis. In this study, a basic toxin (TSGP4) was identified and the presence of an acidic toxin (TSGP2) was confirmed. Both basic and acidic toxins were more lethal than previously described, with TSGP4 (34microg) and TSGP2 (24microg) causing mortality of adult mice within 30min. Pathological effects on the cardiac system, notably of salivary gland extract on an isolated rat heart perfusion system and of purified toxins on mouse electrocardiogram patterns could be observed. TSGP4 caused Mobitz type ventricular block, while TSGP2 induced ventricular tachycardia. Conversely, fractions from reversed phase high performance liquid chromatography preparations caused paralysis-like symptoms of the limbs after only 48h. The toxins also differ from previously described tick paralysis toxins in terms of molecular behavior and properties. These results indicate that tampan toxicoses and tick paralysis are unrelated pathogenic phenomena.

Animals↗

PubMed2003

Lysozyme from the gut of the soft tick Ornithodoros moubata: the sequence, phylogeny and post-feeding regulation.

Sequence of a tick gut lysozyme (TGL) from the soft tick Ornithodoros moubata was determined by cloning and sequencing of overlapping polymerase chain reaction (PCR) and RACE PCR products. It is the first lysozyme sequence representing the subphylum Chelicerata. The resulting open reading frame codes for a putative signal peptide of 22 amino-acid residues and a mature protein composed of 124 amino-acids. Calculated mass of the protein is 14037.75 Da and a theoretical isoelectric point is 8.16. The phylogenetic analysis revealed that the TGL belongs to the c-type lysozymes. It forms a distinct monophyletic group together with multiple lysozyme-like sequences found in the gene products agCP6542 from Anopheles gambiae strain PEST and CG8492-PA from Drosophila melanogaster. This group is referred to as an H-branch due to a unique histidine residue at position 52 which replaces the highly conserved tyrosine present in the vast majority of c-type lysozymes. TGL seems to be an interesting case in which the features of lysozymes with anti-bacterial and digestive function are combined. Semi-quantitative RT-PCR and Northern blotting analysis demonstrated that TGL is strongly up-regulated at the transcriptional level after a bloodmeal. The maximum lysozyme mRNA level was detected 16 h post bloodmeal and the message remained stable for 5 days and then it slowly dropped down to the level of non-fed ticks within 2 weeks.

Amino Acid Sequence↗

PubMed2006

Babesia parasites develop and are transmitted by the non-vector soft tick Ornithodoros moubata (Acari: Argasidae).

Ornithodoros moubata ticks were fed on blood infected with Babesia equi. However, the parasites were quickly cleared as evidenced by the disappearance of B. equi-specific ribosomal RNA from the ticks. We hypothesized that if the Babesia parasite can escape midgut-associated barriers a non-vector tick can become infected with Babesia. To test this hypothesis, B. equi parasite-infected blood from in vitro culture was injected into the haemocoel of ticks. B. equi-specific rRNA was surprisingly detected 45 days after injection even in the eggs. Babesia-free dogs were infested with O. moubata ticks that were infected by inoculation with B. gibsoni-infected red blood cells. Parasitaemia and antibody production against Bg-TRAP of B. gibsoni increased gradually. These results indicate that O. moubata may be a useful vector model for Babesia parasites and also a very important tool for studies on tick immunity against Babesia parasites and tick-Babesia interactions.

Animals↗

PubMed2001

Ornithodoros porcinus ticks, bushpigs, and African swine fever in Madagascar.

African swine fever (ASF) has recently made its appearance in Madagascar. Ticks of the Ornithodoros moubata group, considered to be O. porcinus Walton, 1962 were formerly known to occur in western Madagascar, but seem to have disappeared from that region. However, three new sites where they occur were found in the humid and cool central highlands of Anatananarivo province. These ticks are known to be efficient reservoirs and vectors of ASF and constitute a considerable complication to the control of the disease. The authors also discuss another potentially complicating factor, the presence of a species of African bushpig, Potamochoerus larvatus.

African Swine Fever↗

PubMed2007

In vivo characterization and therapeutic efficacy of a C5-specific inhibitor from the soft tick Ornithodoros moubata.

The involvement of complement (C) in inflammatory diseases has driven the search for agents capable of inhibiting dysregulated complement activation. Many of these reagents inhibit the C3 convertases during the early stages of the cascade. However, a drawback of total systemic C inhibition, particularly in longterm treatment of chronic disease, is potentiation of infection and immune complex disease due to an inability to opsonize complexes and foreign cells and to lyse pathogens. Recent identification of a C5-binding protein in the salivary gland of the soft tick Ornithodoros moubata has enabled development of a terminal pathway-specific reagent, OmCI, with potential to ameliorate disease while leaving key physiological processes unaffected. Here we demonstrated that OmCI has broad cross-species activity. When given intravenously to rodents, OmCI totally ablated complement hemolytic activity, which gradually restored as C5 was resynthesized. The circulating half-life of OmCI was 30 h, demonstrating a much slower clearance than other small, biological agents. Using C5-sufficient and C5-deficient mice we showed that prolonged half-life was due to binding to plasma C5. Surface plasmon resonance analysis of C5 binding to OmCI confirmed a high binding affinity with a slow dissociation rate. OmCI was effective in preventing experimental autoimmune myasthenia gravis induced by passive transfer in normal Lewis rats. OmCI ablated clinical disease, reduced C3 and C9 deposition at the neuro-muscular junction, and effected a marked reduction in cellular infiltration at this site. These data offer exciting prospects for targeted treatment of complement-mediated diseases without the detrimental inhibition of the opsonic roles of complement.

Animals↗

PubMed2006

Molecular characterization and oligosaccharide-binding properties of a galectin from the argasid tick Ornithodoros moubata.

The argasid tick Ornithodoros moubata is a vector of various viral and borrelian diseases in animals and humans. We report here molecular characterization and oligosaccharide-binding properties of a novel galectin (OmGalec) from this tick. OmGalec consisted of 333 amino acids with a predicted molecular weight of 37.4 kDa. Its amino acid sequence did not contain a signal peptide or transmembrane domain. It possessed tandem-repeated carbohydrate recognition domains, in which the typical motifs important for carbohydrate affinity were conserved. OmGalec was expressed both transcriptionally and translationally at all stages of the tick life cycle and in multiple organs and was abundant in hemocytes, midguts, and reproductive organs, which are of importance in immunity, interaction with pathogens, and development, respectively, suggesting that OmGalec is a multifunctional molecule. The oligosaccharide affinity profile analyzed by applying an automated frontal affinity chromatography system revealed that rOmGalec showed a general feature of the galectin family, i.e. significant affinity for lactosamine-type disaccharides, Galbeta1-3(4)Glc(NAc), via recognition of 4-OH and 6-OH of galactose and 3 (4)-OH of Glc(NAc). Its preference for type I saccharides and alpha1-3GalNAc-containing oligosaccharides might provide clues for identifying its ligands and its potential multiple functions. Our results may contribute to the elucidation of galectin functions in the development and immunity of arthropods and/or vector and pathogen interaction and provide valuable information for the development of novel tick control strategies.

Amino Acid Sequence↗

PubMed2005

Genetic structure of the tick Ornithodoros coriaceus (Acari: Argasidae) in California, Nevada, and Oregon.

The argasid tick Ornithodoros coriaceus (Koch) is the only confirmed vector of epizootic bovine abortion (EBA) in the United States. The disease and its tick vector have historically been reported in the foothills of the Sierra Nevada Mountains and coast ranges of California. In the past two decades, the range of EBA has apparently expanded into southern Oregon and northern Nevada. Possible explanations for this expansion include 1) increased recognition and reporting of EBA in these regions; 2) widespread movement of tick-infested and EBA-infected hosts with subsequent colonization of these regions by infected ticks; and 3) widespread movement of the EBA agent, independent of tick movements, into extant tick populations in these new regions. The current study was performed to evaluate these hypotheses by examining patterns of variability in a 420-bp segment of the 16S mitochondrial rDNA gene sequence among 210 O. coriaceus individuals from 14 sites in California, Oregon, and Nevada. Sixty-three unique haplotypes were identified in the ticks tested, with 84% of the sequence variation attributable to among-population variation and 16% to within-population variation. A majority of the haplotypes were unique to their particular collection site, whereas only four collection sites shared haplotypes. Overall, very little evidence of gene flow among tick populations was detected, making it unlikely that widespread tick movement had introduced O. coriaceus and the EBA agent into new regions.

Abortion, Veterinary↗

PubMed2003

The major tick salivary gland proteins and toxins from the soft tick, Ornithodoros savignyi, are part of the tick Lipocalin family: implications for the origins of tick toxicoses.

The origins of tick toxicoses remain a subject of controversy because no molecular data are yet available to study the evolution of tick-derived toxins. In this study we describe the molecular structure of toxins from the soft tick, Ornithodoros savignyi. The tick salivary gland proteins (TSGPs) are four highly abundant proteins proposed to play a role in salivary gland granule biogenesis of the soft tick O. savignyi, of which the toxins TSGP2 and TSGP4 are a part. They were assigned to the lipocalin family based on sequence similarity to known tick lipocalins. Several other tick lipocalins were also identified using Smith-Waterman database searches, bringing the tick lipocalin family up to 20. Phylogenetic analysis showed that most tick lipocalins group within genus-specific clades, suggesting that gene duplication and divergence of tick lipocalin function occurred after tick speciation, most probably during the evolution of a hematophagous lifestyle. TSGP2 and TSGP3 show high sequence identity and group terminal to moubatin, an inhibitor of collagen-induced platelet aggregation from the tick, O. moubata. However, no platelet aggregation inhibitory activity is associated with the TSGPs using ADP or collagen as agonists, suggesting that TSGP2 and TSGP3 duplicated after divergence of O. savignyi and O. moubata. This timing is supported by the absence of TSGP2-4 in the salivary gland extracts of O. moubata. The absence of TSGP2 and TSGP4 in salivary gland extracts from O. moubata correlates with the nontoxicity of this tick species. The implications of this study are that the various forms of tick toxicoses do not have a common origin, but must have evolved independently in those tick species that cause pathogenesis.

Amino Acid Sequence↗

PubMed2004

Characterization of pathogenic and non-pathogenic African swine fever virus isolates from Ornithodoros erraticus inhabiting pig premises in Portugal.

Ten African swine fever virus isolates from the soft tick Ornithodoros erraticus collected on three farms in the province of Alentejo in Portugal were characterized by their ability to cause haemadsorption (HAD) of red blood cells to infected pig macrophages, using restriction enzyme site mapping of the virus genomes and by experimental infection of pigs. Six virus isolates induced haemadsorption and four were non-haemadsorbing (non-HAD) in pig macrophage cell cultures. The restriction enzyme site maps of two non-HAD viruses, when compared with a virulent HAD isolate, showed a deletion of 9.6 kbp in the fragment adjacent to the left terminal fragment and of 1.6 kbp in the right terminal fragment and an insertion of 0.2 kbp in the central region. The six HAD viruses isolated were pathogenic and produced typical acute African swine fever in pigs and the four non-HAD isolates were non-pathogenic. Pigs that were infected with non-HAD viruses were fully resistant or had a delay of up to 14 days in the onset of disease, after challenge with pathogenic Portuguese viruses. Non-HAD viruses could be transmitted by contact but with a lower efficiency (42-50 %) compared with HAD viruses (100 %). The clinical differences found between the virus isolates from the ticks could have implications for the long-term persistence of virus in the field because of the cross-protection produced by the non-pathogenic isolates. This may also explain the presence of seropositive pigs in herds in Alentejo where no clinical disease had been reported.

African Swine Fever Virus↗

PubMed2006

Kinetics of African swine fever virus infection in Ornithodoros erraticus ticks.

The kinetics of African swine fever virus (ASFV) infection in Ornithodoros erraticus ticks were investigated in specimens collected in the field at different times following an outbreak of the disease in Portugal in 1999 and in ticks infected experimentally with a virus isolated from a tick collected during this outbreak. In ticks collected from the field, initial screening for ASFV was carried out by PCR, followed by attempts to isolate the virus in macrophage cultures. Considering total numbers of ticks tested independently of developmental stages, ASFV DNA was detected in 42.3, 26.4 and 22.4% of specimens collected at weeks 0, 32 and 63 following the outbreak, respectively. Although virus was not isolated from most of these ticks, the proportion of isolations from large nymphs and adults increased between weeks 0 and 32 from 2 to 9 % and from 5 to 11.5%, respectively. These results, together with the higher virus titres at week 32, suggest that virus replication occurred. In contrast, virus isolations from small nymphs decreased over this period, from 5 to 1.3%. At week 63, infection rates decreased for all stages. Experimental infections showed the occurrence of virus replication within 4 weeks post-feeding and maintenance of high titres in almost 100% of ticks until 20 weeks post-infection. At weeks 41 and 61, a drop in virus titres and infection rates was observed. Relevant to the understanding of African swine fever epidemiology, our results show that ASFV replicates and persists in O. erraticus, but a viral clearance occurs at later times in both natural and experimental infections.

African Swine Fever↗

PubMed2004

New World relapsing fever Borrelia found in Ornithodoros porcinus ticks in central Tanzania.

Ticks were collected from 8 houses in Mvumi Mission village, near Dodoma, Tanzania. All ticks were examined for Borrelia infestation by flagellin gene-based nested polymerase chain reaction. All houses were highly infested with ticks, and all ticks collected were of the Ornithodoros porcinus species. Fifty-one out of 120 ticks were infected with spirochetes, and a flagellin gene sequence comparison showed that most of the spirochetes belonged to Borrelia duttonii, which is the causative agent of tick-borne relapsing fever in East Africa. The rest of the spirochetes were quite different from B. duttonii and instead resembled the New World tick-borne relapsing fever borreliae. Phylogenetic analysis using 16S ribosomal RNA gene sequences also supported the interpretation that the spirochete was a Borrelia species distinct from previously described members of the genus.

Animals↗

PubMed2004

African swine fever virus multigene family 360 genes affect virus replication and generalization of infection in Ornithodoros porcinus ticks.

Recently, we reported that African swine fever virus (ASFV) multigene family (MGF) 360 and 530 genes are significant swine macrophage host range determinants that function by promoting infected-cell survival. To examine the function of these genes in ASFV's arthropod host, Ornithodoros porcinus porcinus, an MGF360/530 gene deletion mutant (Pr4Delta35) was constructed from an ASFV isolate of tick origin, Pr4. Pr4Delta35 exhibited a significant growth defect in ticks. The deletion of six MGF360 and two MGF530 genes from Pr4 markedly reduced viral replication in infected ticks 100- to 1,000-fold. To define the minimal set of MGF360/530 genes required for tick host range, additional gene deletion mutants lacking individual or multiple MGF genes were constructed. The deletion mutant Pr4Delta3-C2, which lacked three MGF360 genes (3HL, 3Il, and 3LL), exhibited reduced viral growth in ticks. Pr4Delta3-C2 virus titers in ticks were significantly reduced 100- to 1,000-fold compared to control values at various times postinfection. In contrast to the parental virus, with which high levels of virus replication were observed in the tissues of infected adults, Pr4Delta3-C2 replication was not detected in the midgut, hemolymph, salivary gland, coxal gland, or reproductive organs at 15 weeks postinfection. These data indicate that ASFV MGF360 genes are significant tick host range determinants and that they are required for efficient virus replication and generalization of infection. The impaired virus replication of Pr4Delta3-C2 in the tick midgut likely accounts for the absence of the generalized infection that is necessary for the natural transmission of virus from ticks to pigs.

African Swine Fever↗

PubMed2001

Isolation and characterization of Borrelia parkeri in Ornithodoros parkeri (Ixodida: Argasidae) collected in Colorado.

This study describes the identification of Borrelia parkeri spirochetes in Colorado. Two isolates of B. parkeri (6230 and 6232) were recovered from Ornithodoros parkeri Cooley ticks collected at an inactive prairie dog town in Moffat County. Both isolates were partially characterized by sequencing and subsequent parsimony and neighbor-joining analyses of appropriate regions of the 16S ribosomal RNA, flagellin and P66 genes. Analyses of the 16S gene sequences from the Colorado isolates indicated that they were more closely related to B. parkeri and B. tucatae than to B. hermsii or the other species of Borrelia investigated in this study. Additional analyses of amino acid sequences for flagellin and P66, however, clearly demonstrated that isolates 6230 and 6232 were most closely related to B. parkeri. The possible significance of B. parkeri as an agent of human disease is discussed.

Animals↗

PubMed2003

Antibacterial hemoglobin fragments from the midgut of the soft tick, Ornithodoros moubata (Acari: Argasidae).

Midgut contents of Ornithodoros moubata showed strong antibacterial activity against Staphylococcus aureus. A combination of reversed-phase chromatography and mass spectrometric analysis was used to isolate two antibacterial peptides from the tick midgut lumen. Partial amino acid sequences by Edman degradation of these two peptides showed they are identical with the 1-11 and 3-19 portions of rabbit a hemoglobin. Host rabbit a hemoglobin appears to be cleaved between Met32 and Phe33 to produce these two antibacterial peptides. Isolation of a host bovine hemoglobin fragment with antimicrobial activity has been demonstrated in the Ixodid tick, Boophilus microplus (Fogaca et al. 1999). Similar immune mechanisms in the two major families of ticks, Ixodidae and Argasidae, appear to use the hemoglobin of the host as an antimicrobial agent in midgut defense.

Amino Acid Sequence↗