Tick antigens recognized by serum from a guinea pig resistant to infestation with the tick Rhipicephalus appendiculatus

Immune resistance to infestation by an ixodid tick, Rhipicephalus appendiculatus, the vector of the cattle disease East Coast Fever, was induced in a guinea pig by repeated tick infestation. This resistance is expressed as the ability of the host to interfere with tick feeding. Resistance to ixodid tick feeding is an acquired response mediated by host antibody. We report the use of antibodies from a resistant host animal, in immunoblotting, to characterize the tick antigens recognized. The major tick antigens identified had molecular weights of 120,000, 94,000, 88,000, 77,000, 58,000, 46,000, 35,000, 31,000, 28,000, 25,000, 20,000 and 16,000. Most of these antigens were found in tick salivary glands. The presence and concentration of many tick salivary antigens appeared to vary with relation to the tick feeding cycle. Many of the antigens present in salivary glands were also detected in tick cement. Tick gut extract, although a poorer source of antigens, contained more of the 31,000 dalton antigen than salivary glands. Larval and nymphal tick extract lacked many of the antigens present in adult ticks. The data suggest that tick resistance is a complex phenomenon probably elicited by several different tick antigens.     

Dot-ELISA assessment of guinea pig antibody responses to repeated Dermacentor andersoni infestations

Acquired resistance to ixodid tick infestation is expressed by cattle and laboratory animals. Humoral factors appear to be involved in host acquired resistance to tick bite; however, specific immune responses have yet to be fully characterized. This study examined tick resistance expressed by Hartley guinea pigs upon repeated infestation with Dermacentor andersoni, and describes longitudinal development of antigen specific immunoglobulin over approximately 180 days. Guinea pigs were infested either 4 times with D. andersoni adults, or twice with nymphs. Both infestation groups, adults and nymphs, demonstrated a significant level of resistance to re-infestation, following initial exposure. Dot enzyme-linked immunosorbent assay (Dot-ELISA) was employed to detect antibody reactive with D. andersoni salivary gland antigens (SGA). Animals infested with adults had antibody that increased at a relatively constant rate until the fourth infestation, which was differentiated by a sharp increase in titer, that was maintained for approximately 2 wk. Guinea pigs that received nymph infestations had SGA-specific antibody; however, titers were lower than those in the adult infestation group. Antibody levels continued to increase approximately 80 days beyond the final (second) infestation for this group. A direct correlation between resistance and antibody titer was not evident, since resistance was relatively stable after the second infestation in both infestation groups, and tick-specific immunoglobulin levels continually increased.     

Serodiagnostic antibody responses to Psoroptes sp. infestations in bighorn sheep

The antibody responses of bighorn sheep (Ovis canadensis) infected with Psoroptes sp. mites were investigated by enzyme linked immunosorbent assay on western blots of P. cuniculi antigens. Serum from 20 Psoroptes sp.-infested bighorn sheep (O. canadensis mexicana, O. canadensis nelsoni, O. canadensis canadensis) from New Mexico, Nevada, California, and Idaho reacted strongly with mite antigens ranging from 12 to 34 kd. Serum from 35 Psoroptes sp.-free bighorn sheep of unknown tick infestation status and from three Psoroptes sp.-free bighorn sheep infested with Dermacentor hunteri ticks did not react with these antigens. Psoroptes sp.-specific antibody responses were present throughout a 16 mo period in one infected bighorn sheep, but were not detectable 8 mo following successful treatment. These results demonstrate that specific serodiagnosis of Psoroptes sp. infestation is feasible in bighorn sheep and suggest that antibody responses are indicative of current or recent infestation.     

Female tick Hyalomma marginatum marginatum salivary glands: preliminary study on protein changes during feeding process and antigens recognized by repeatedly infested cattle

Proteins extracted from salivary glands of unfed, three days and five days fed adult Hyalomma marginatum marginatum were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). We have noticed changes during the three feeding steps. Some proteins disappeared during feeding process (23, 38, 39, 40 to 50, 95 and 112 kDa), they might be proteins which were converted in other substances and are secreted. Other antigens (13 to 14, 20, 25, 29, 165 and 210 kDa) were synthesized as a result of tick attachment and feeding. They may be related to growth and development or are the ciment which fixed the adult. Also, three Holstein calves were infested five times with 100 pairs of adult ticks of the same species. The five infestations were performed two weeks from the previous infestation. The sera before infestations and after each infestation were used in western-blot analyses to identify antigens from five days salivary gland extracts of the primary infestation of ticks. Three antigens (18.7, 50 and 80 kDa) were revealed weakly after the first and the second infestations by sera samples but not at infestation onward. Others (13.5, 17 to 18.5, 25, 30, 70, 133, 176 and 193 kDa) were revealed only by sera taken after manifestation of resistance (third infestation). A 13.5 kDa antigen was particularly revealed when resistance had appeared and became more evident after the fourth and fifth infestations. The late antigens recognized might be associated with establishment of calves resistance against ticks.     

Offspring quality and tick infestation load in brood rearing great tits Parus major

Although the impact of nest-dwelling ectoparasites on nestlings in altricial birds is relatively well documented, little information is available on the fitness consequences of bird–ectoparasite systems with limited ectoparasite transmission between parents and their offspring. In this particular context we tested the hypothesis that parental infestation by a haematophageous field ectoparasite, the sheep tick Ixodes ricinus , in a hole-breeding passerine bird, the great tit Parus major , ultimately leads to the reduction of nestling quality. Observational data of a long-term study show that 15 days after hatching, nestlings' body condition (body mass corrected for body size) was negatively associated with the tick load of the mother, while no association was found with tick load of the father. A field experiment was conducted to test if adult infestation by ticks leads to a reduction in offspring quality. The field experiment consisted of a two by two factorial design: neither, one or both parents were infested with a high tick load (12 nymphs) when nestlings were eight days old. Four days after manipulation of the parents, the effect of the treatment on nestlings' health was assessed by measuring haematological and biochemical parameters. Body condition of nestlings was monitored from three days after hatching, until seven days after tick manipulation of the parents. Although parental tick manipulation resulted in successful tick feeding, nestlings' health parameters were not affected. We therefore suggest that the negative association between nestling condition and parental tick load does not reflect a causal effect of parasites, but either reflects a common environmental factor affecting parental infestation levels and offspring condition, or reflects parental quality. We propose different explanations why this association is expressed in female parents only.     

Competition,facilitation or mediation via host? Patterns of infestation of small European mammals by two taxa of haematophagous arthropods

1. We studied the effect of flea infestation on the pattern of tick (Ixodes ricinus and Ixodes trianguliceps) infestation on small mammals. 2. We asked (1) whether the probability of an individual host being infested by ticks was affected by its infestation of fleas (number of individuals and species) and (2) whether the abundance and prevalence of ticks in a host population was affected by the abundance, prevalence, level of aggregation, and species richness of fleas. 3. The probability of a host individual being infested by ticks was affected negatively by flea infestation. At the level of host populations, flea abundance and prevalence had a predominantly positive effect on tick infestation, whereas flea species richness had a negative effect on tick infestation. 4. The effect of flea infestation on tick infestation was generally greater in I. ricinus than in I. trianguliceps, but varied among host species. 5. It can be concluded that the effect of fleas on tick infestation of small mammals may be either negative or positive depending on the level of consideration and parameters involved. The results did not provide support for direct interactions between the two ectoparasite taxa, but suggested population and community dynamics and the defence system of the hosts as possible factors.     

Vaccination with 'concealed' antigens for tick control

Ticks are responsible for substantial economic losses to the livestock industry, necessitating intensive use of chemical acoricides in many parts of the world. Problems of chemical residues, cost of acoricides, and development of resistance by ticks, have long been recognized and have helped to stimulate interest in tick control by immunological means (see Box 1). One approach has been to seek ways to enhance the natural immunity often acquired by animals in response to tick infestation. An alternative, discussed here by Peter Willadsen and David Kemp, is to vaccinate with 'concealed' tick antigens not normally encountered by the host, and so stimulate a different immune effector mechanism.     

Characterization of two cDNAs encoding serine proteinases from the hard tick Haemaphysalis longicornis

Host vaccination against tick infestation is at present the most practical and sustainable alternative tick control method to the current acaricide use which has serious limitations. However the success of this approach to control ticks depends upon the identification of target vaccine antigens. Members of the serine proteinase gene family may represent an interesting group of proteins to target as candidate antigens because of their involvement in regulation of many physiological functions and development processes in a wide range of organisms. We used RT-PCR with the 3' and 5' RACE to clone two cDNAs encoding full-length serine proteinases from the hard tick, Haemaphysalis longicornis. RT-PCR degenerate primers were designed from amino acid sequences surrounding active sites, His(57) and Ser(195) conserved among most known serine proteinase. Gene specific primers designed from nucleotide sequences of the RT-PCR products were used to prime the 3' and 5' RACE. Southern blotting analysis showed that both HLSG-1 and -2 are single copy. The 2 cDNAs, HLSG-1 and -2 are 1.2 and 1.0 kb long in size with open reading frames encoding polypeptides with 37.7 and 31.2 kDa predicted molecular mass respectively. Northern blotting analysis of total RNA from unfed and partially fed whole ticks showed that the expression of mRNAs for both HLSG-1 and -2 was induced by blood feeding. Expression analysis by RT-PCR showed that both HLSG-1 and -2 are expressed in other tick organs in addition to salivary glands and midguts. The 6 serine proteinase consensus cyteine residues are well conserved in both HLSG-1 and -2. We have discussed our findings with respect to tick vaccine development research.     

Host community structure and infestation by ixodid ticks: repeatability,dilution effect and ecological specialization

Abundance of a species in a location results from the interplay between the intrinsic properties of that species and the extrinsic properties, both biotic and abiotic, of the local habitat. Intrinsic factors promote among-population stability in abundance, whereas extrinsic factors generate variation among populations of a species. We studied (a) repeatability and (b) the effect of abundance and species richness of small mammals on the level of their infestation by larvae and nymphs of Ixodes ricinus (ecological generalist) and Ixodes trianguliceps (ecological specialist). We asked if tick infestation parameters are characteristic (=repeatable) for a particular host species or a particular stage of a particular tick species. We also asked how abundance and diversity of hosts affect the level of tick infestation on a particular host species. We predicted that the dilution effect (decrease in tick infestation levels with an increase of host abundance and/or species richness) will be better expressed in an ecological generalist, I. ricinus, than in an ecological specialist, I. trianguliceps. We found that (a) tick abundance, prevalence and aggregation were generally repeatable within tick species/stage; (b) tick abundance and prevalence, but not the aggregation level, were repeatable within host species; (c) the proportion of variance among samples explained by the differences between tick species and stages (as opposed to within-tick species and stage) was higher than that explained by the differences between host species (as opposed to within host species); and (d) the relationship between tick infestation parameters and host abundance and diversity revealed the dilution effect for I. ricinus but not for I. trianguliceps.     

镰形扇头蜱两个组织蛋白酶L-样半胱氨酸蛋白酶新基因的克隆与序列分析

蜱是动物常见的外寄生虫,并且传播多种人和动物的疾病,严重危害畜牧业发展和人类健康。为了寻找基因工程疫苗候选抗原基因,根据半胱氨酸蛋白酶的保守性氨基酸序列及镰形扇头蜱氨基酸密码子偏好设计引物,PCR扩增、测序并分析得到2个镰形扇头蜱的半胱氨酸蛋白酶基因片段cysA和cysB,再通过RACE的方法得到全长基因序列。cysA全长168bp,编码332个氨基酸;cysB全长1153bp,编码335个氨基酸。经过分析, CysA和CysB均与其他蜱种或物种的组织蛋白酶L样半胱氨酸蛋白酶有高度同源性,两者均含有半胱氨酸蛋白酶活性位点处的保守性氨基酸序列, 因此cysA, cysB均为镰形扇头蜱两个新的组织蛋白酶L-样半胱氨酸蛋白酶基因。RT-PCR分析表明,CysA和CysB在镰形扇头蜱的不同发育阶段表达情况不一。     

A synthetic bmti n-terminal fragment as antigen in bovine immunoprotection against the tick Boophilus microplus in a pen trial

Cattle tick control remains a serious problem in cattle farms worldwide, due to the limited success achieved with chemicals. Although the use of vaccines for tick control may open possibilities for an integrated approach, the search for other protective antigens is still necessary to improve control. Boophilus microplus is a rich source of BmTIs, serine proteinase inhibitors that are present throughout the tick's lifecycle. The present paper reports a pen trial conducted to evaluate the performance of a synthetic BmTI N-terminal fragment as antigen against the tick in bovines. The trial was conducted with two groups of eight crossbred cattle under controlled infestation: one group was vaccinated with BmTI peptide using saponin as adjuvant and the other was kept as control. Challenge was performed with 15,000 larvae. The specific IgG response was measured by ELISA, with successful outcomes for vaccinated animals. Vaccination resulted in an 18.4% level of efficacy when compared with the control group.     

Ticks Ixodes uriae and the breeding performance of a colonial seabird, king penguin Aptenodytes patagonicus

High densities of penguins in their colonies and the continuous use of these sites over consecutive reproductive periods increase the risk of development of tick populations. We have studied the effects of tick parasitism by Ixodes uriae in a colony of king penguins Aptenodytes patagonicus at Possession Island during three breeding seasons. We investigated the prevalence and periods of tick infestation during the one-year breeding cycle of penguins. The effects of tick parasitism on penguin breeding performance were assessed from photographs of the colony and with an automatic penguin identification system. We compared two groups of penguins carrying individual subcutaneous electronic tags, one group breeding in an infested area and the other in a non-infested area. Tick feeding activity was coincident with the periods when adult penguins stayed ashore for six days or more, i.e. during the incubating period. This duration corresponds to the duration of a tick meal on the host. The level of infestation varied between years. Penguins showed a lower incubating success in infested areas during a year of high infestation. In an infested area, individuals seen with ticks had a lower breeding success in rearing a one-year old chick than those seen without ticks.     

Rejection of ticks from guinea pigs by anti-hapten-antibody-mediated degranulation of basophils at cutaneous basophil hypersensitivity sites: role of mediators other than histamine

Previous studies have established that recruitment of basophils to sites of tick feeding in guinea pigs is required to effect immune resistance. In the current study, actively sensitized guinea pigs treated three times daily with H-1 (mepyramine) and H-2 (cimetidine) histamine receptor antagonists, during the challenge tick infestation period, expressed normal resistance to Amblyomma americanum larvae. Similarly, naive guinea pigs treated with anti-histamines four times daily, beginning 7 days before transfer of immune serum and tick challenge and continuing through the tick infestation period, also expressed normal antibody-mediated resistance to A. americanum. These results indicated that histamine was not an important basophil mediator of the resistance response. Ticks allowed to feed on tissue rich in basophils that were induced by sensitization and subsequent local challenge with non-tick protein antigen, keyhole limpet hemocyanin (KLH), expressed normal yield. Ticks that fed on similar tissue rich in basophils induced by sensitization and challenge with KLH, in which the basophils expressed anti-picryl specificity due to systemic passive transfer of anti-picryl antibodies, were rejected when basophils were induced to degranulate by i.v. challenge with picryl antigen at 6 hr (29% rejection), 12 hr (18% rejection), 24 hr (22% rejection), and 48 hr (37% rejection) post-tick attachment. However, basophil degranulation at 18, 72 and 96 hr post-tick attachment had no adverse effect on tick feeding. These hosts were protected from systemic anaphylaxis by treatment with the anti-histamine mepyramine. Release of histamine occurred at tick feeding sites, but vasoactive effects were blocked by mepyramine treatment as evidenced by a lack of increased vascular permeability (bluing) at these sites compared with non-tick-infested tissues, or to cutaneous basophil hypersensitivity (CBH) sites of animals not protected with mepyramine. These results indicate that local recruitment and subsequent degranulation of basophils via immune mechanisms dependent on non-tick antigens can lead to tick rejection, and that basophil-derived mediators other than histamine are involved in this immune resistance response to A. americanum ticks. The identity of the crucial basophil mediator(s) is not known. The significant susceptibility of ticks to basophil-mediator release at 6 to 12 hr and 24 to 48 hr post-attachment coincides with the tick attaching and fast-feeding phases, respectively, suggesting that these phases of tick parasitism are particularly susceptible to the effect of basophil mediators other than histamine.     

Acquired resistance to ixodid ticks induced by tick cement antigen

Antisera from guinea pigs made resistant to infestation with an ixodid tick of east and central Africa,Rhipicephalus appendiculatus, were used to identify the tick antigens they recognized by immunoblotting. Most of the antigens were found in tick salivary glands and in tick attachment cement. Antisera fromR. appendiculatus-resistant guinea pigs also recognized some salivarygland antigens in ticks of other species (R. pulchellus, R. evertsi, Amblyomma variegatum andA. gemma). Antibodies against the most strongly recognizedR. appendiculatus antigen, a 20-kDa molecule, were only poorly reactive with similar-sized molecules in the other ticks. A 94-kDa antigen, which appeared to have broader cross-reactivity, was purified fromR. appendiculatus attachment cement, and a monospecific rabbit serum was raised against it. This antiserum clearly recognized a molecule of similar molecular weight inR. pulchellus andR. evertsi. Intravenous inoculation of rabbits with the purified molecule elicited delayed-type hypersensitivity to the antigen. The hypersensitive rabbits demonstrated resistance to feeding ofR. appendiculatus ticks but slight enhanced feeding ofR. pulchellus ticks. These results are discussed with respect to their relevance for artificial induction of tick-feeding resistance.     

Acquired resistance in dogs to repeated infestation with Ixodes scapularis (Acari: Ixodidae) reduces tick viability and reproductive success

The purpose of this study was to determine whether dogs develop acquired resistance to adult Ixodes scapularis infestation in an experimental model. Five dogs were each infested with ten mating pairs of ticks every week for 7 consecutive weeks, another five dogs were each infested with ten mating pairs once every 2 weeks for 10 weeks and four dogs served as controls not exposed to ticks. All ticks were allowed to feed to repletion and were collected only after dropping from the host. Several variables were measured to determine the extent of blood feeding success. Regression analysis indicated that the engorgement success, survival and mean tick engorgement weight declined with repeated infestation in both groups of dogs (p<0.05). Tick oviposition as well as the F1 viability declined with each successive infestation in both groups. These results suggest that repeated infestation with I. scapularis elicits a protective immune response against tick feeding and could serve as a limiting factor in the spread and transmission of Borrelia burgdorferi.     

A case of histologically diagnosed tick infestation on the scalp of a Korean child

A scalp mass surgically excised from a 4-year-old Korean boy was identified as a tick through histological observations. In sections of the mass, characteristic features of a tick, including its gross contour, cuticular structures, well developed musculature and salivary glands, and the capitulum, were discovered. In particular, the capitulum is anteriorly protruded, which strongly suggests that the specimen be a hard tick of family Ixodidae. However, the present histological features were not enough to determine the genus and species of the tick, because information on sectional morphologies of different tick species is unavailable. This is a rare case of tick infestation on the scalp diagnosed in histological sections.     

Vaccination againstBoophilus microplus: Localization of antigens on tick gut cells and their interaction with the host immune system

Cattle have been vaccinated againstBoophilus microplus with antigens derived from partially fed female ticks. The immune response of the host lyses the gut cells of adult ticks, causing a reduction in the number, weight and reproductive capacity of engorging ticks. This response is different from the immunity that cattle acquire after repeated tick infestation. Evidence is presented that the antigens used in vaccination are located on the plasma membrane of the gut cells and it is unlikely that these antigens are secreted into the host during feeding. Vaccination using such concealed antigens may not encounter the mechanisms of immune evasion that parasites usually demonstrate.In-vitro assays suggest that vaccination immunity is not dependent on the need to stimulate cell-mediated responses. Immunoglobulin G alone, or with the aid of complement, is enough to damage tick gut.The normal function of the one protein antigen isolated so far is unknown but we speculate that it serves some vital function on the cell plasma membrane.     

Ixodid ticks on wild donkeys in a Mediterranean nature reserve (Asinara National Park): diversity and risk factors

The Sardinian coloured donkey Equus asinus (Perissodactyla: Equidae) and its albino colour morph represent the wildlife species most typical of the island of Asinara. This Mediterranean island represents a favourable context for ticks and tick‐borne diseases; however, knowledge of the tick fauna on Asinara is scarce. A total of 106 Sardinian donkeys were inspected for tick infestation from June to November 2015. All ticks found were collected, classified by stage and sex, and identified to species level. The level of infestation of each donkey was determined; both the overall tick infestation and infestations of each detected species were classified on a scale of 1–3 to give an infestation score (IS). Overall, 256 hard ticks were collected from 60 of 106 donkeys (56.6%). Rhipicephalus bursa, Haemaphysalis punctata and Hyalomma marginatum (all: Ixodida: Ixodidae) infested 26.4%, 28.3% and 6.6% of donkeys, respectively. Different variables affected the IS. With reference to overall tick infestation, a higher IS was observed in donkeys grazing on grassland and Mediterranean shrubland and in albino donkeys compared with coloured donkeys. The collected ticks included species involved in the transmission of pathogens to humans, which highlights the risks for public health in a tourist destination such as Asinara National Park.     

Populations of the ticks Ixodes (Pholeoixodes) hexagonus and Ixodes (Pholeoixodes) canisuga infesting suburban foxes, Vulpes vulpes

Only two species of tick (Ixodes hexagonus and Ixodes canisuga) were found to infest suburban foxes. The populations of these two ticks were examined, their distributions within the host population described, and infestation levels of I hexagonus discussed in relation to the sex, age and behaviour of the host. The most important factor regulating the level of tick infestation is probably the degree of den usage by the host. The tick infestations were found to have minimal effect on the host, and even an abnormally high level of infestation found on one fox was not considered to be lethal.