David H. Kemp

Immunization of cattle against Boophilus microplus using extracts derived from adult female ticks: effects of induced immunity on tick populations.

Abstract Immunization of cattle against Boophilus microplus using extracts derived from adult female ticks: effects of induced immunity on tick populations. International Journal for Parasitology16: 27–34. Injection of extracts derived from adult female ticks induced partial immunity to B. microplus in both Bos tawus and Bos taurus × Bos indicus breeds. Cattle showed a variable response to vaccination with the complete extract; 7/15 showed good resistance to ticks, 5/15 were intermediate and the remaining three animals showed poor resistance. The immunity induced was still evident after 14 weeks of daily challenge with 1000 larvae and tick populations on vaccinated cattle were reduced by 70% compared to matched controls during this period. In another experiment, cattle were challenged twice with 20,000 larvae and tick populations on the vaccinated group were reduced by over 90% compared to a matched unvaccinated group. Following vaccination, serum antibodies to soluble extracts of adult ticks were detected by gel diffusion and radio-immunoassay but antibody levels in individual cattle were not correlated with their immunity to ticks. Protective antigens were found both in the supernatant and the pellet derived from tick extracts. Insoluble complexes formed by reacting serum from successfully vaccinated cattle with soluble crude tick extracts were not protective.

Immunization of cattle against Boophilus microplus using extracts derived from adult female ticks: Feeding and survival of the parasite on vaccinated cattle

Abstract Kemp D. H. , Agbede R. I. S. , Johnston L. A. Y. and Gough J. M. 1986. Immunization of cattle against Boophilus microplus using extracts derived from adult female ticks: feeding and survival of the parasite on vaccinated cattle. International Journal for Parasitology 16 : 115–120. Boophilus microplus adults and larvae were observed in chambers on three European breed cattle which had been vaccinated against the tick and on three unvaccinated controls. The moulting of larve on vaccinated cattle was delayed by up to 12 h but otherwise they were unaffected. On two of the three vaccinated cattle there was progressive death of tick females throughout feeding and up to 60% of females had a damaged gut. These females either failed to engorge, or if they did, many died before egg laying. Males also suffered gut damage. In contrast, the females which survived the first day on control cattle usually completed engorgement and neither females nor males showed damaged gut. No hypertensivity reaction or serious exudation was seen at the site of tick attachment on any animals. This was in contrast to resistance acquired following repeated tick infestations where hypersensitivity reactions caused rejection of attaching ticks, especially larvae, and where no gut damage occurred. Vaccination therefore produced effects which were additional to those that follow repeated infestations. Damage to tick gut was also demonstrated in vitro and a role for complement in this effect was suggested.

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.

Tests to determine LC50 and discriminating doses for macrocyclic lactones against the cattle tick, Boophilus microplus.

Laboratory tests were carried out on larvae and adults of the cattle tick Boophilus microplus to determine the toxicity of macrocyclic lactone acaricides (MLs). Technical and commercial MLs were used in larval packet test (LPT), larval immersion test (LIT) and adult immersion test (AIT). In LIT and AIT the toxicity of MLs was much higher than for LPT. In the AIT, diluting the injectable formulation of MLs in water was as effective as dilution in ethanol+Triton X-100. LC50, LC99.9 and 95% confidence limits were determined so that a discriminating dose (DD) could be set for larval and adult tests in order to diagnose potential resistance to MLs in field samples of the tick. These DDs are for Australian strains of B. microplus and may not be suitable for other strains until further work is carried out. The value of these diagnostic tests can only be verified if or when resistance to MLs emerges in ticks.

Effects of cattle tick (Boophilus microplus) infestation on the bovine immune system.

Abstract The immunosuppressive effect of experimental Boophilus microplus infestation on bovine peripheral blood lymphocytes (PBL) and on host antibody production to a protein antigen (ovalbumin) was examined. Boophilus microplus infestation caused a marginal decrease in the percentage of T lymphocytes in PBL, which was observed in both lightly (5000 larvae) and heavily (40 000 larvae) infested cattle, and began at the second infestation and continued until the end of the fourth infestation. The percentage of B lymphocytes in heavily tick-infested cattle was less than that in non-infested control cattle after the fourth infestation. The response of PBL from tick-infested cattle to phytohemagglutinin (PHA) was always less than that of tick-free cattle after the second infestation. No noteworthy differences were detected between the three stages of tick infestation, that is, 1 week before the peak of adult engorgement, the middle of the peak and 1 week after all ticks had dropped. Boophilus microplus saliva (100 μl ml−1) suppressed 47% of the response of bovine PBL to PHA in vitro. This suppressive effect of saliva may contribute to the lowerresponsiveness of PBL from tick-infested cattle. Antibody production by tick-infested cattle was examined during the third and fourth heavy tick infestation. Tick-infested cattle showed a diminished response against ovalbumin after the second immunization. The immunosuppressive effects of tick infestation may play an important role in tick survival or in the transmission of tick-borne diseases in the field.

Digestion in the cattle-tick Boophilus microplus: Light microscope study of the gut cells in nymphs and females

Abstract The gut caeca of B. microplus were studied by light microscopy using paraffin and methacrylate embedded material. It has been shown that during feeding of nymphs and adults, the midgut consists of five cell types, stem cell, digest cell, secretory cells (s1) and (s2) and basophilic cell. The stem cell differentiates into any of the other cell types. The digest cell matures through a series of stages and has up to three generations during feeding on the host. The final generation has two distinct cell types, the first type is thought to be capable of both phagocytosis and pinocytosis. Cells of the second type are predominant at the end of feeding, and may be specialized to ingest and digest haemoglobin. The final stage of the digest series is the spent digest cell which discharges its content into the gut lumen or is excreted whole. The basophilic cell has structures which suggest that one of its functions is to transport digested materials, water and ions across the gut. Secretory cell (s1) secretes a glycoprotein which may be a haemolysin and secretory cell (s2) secretes the gut “colloid” mass, an acid mucopolysaccharide, which may function as an anticoagulant. Intracellular digestion leads to the breakdown of host blood and storage of lipid and glycogen in the digest cells.

Localization of a low abundance membrane protein (Bm86) on the gut cells of the cattle tick Boophilus microplus by immunogold labeling

A preembedding immunogold technique was used to locate Bm86, an antigen from the gut digest cells of the cattle tick Boophilus microplus. Gut from partially engorged female ticks was everted to expose the cells, lightly fixed in 4% paraformaldehyde, and then incubated in rabbit antisera against a recombinant form of Bm86. Following incubation in a secondary antibody conjugated to 1-nm colloidal gold, Bm86 antigenic sites were visualized for both light and electron microscopy using silver enhancement. Bm86 was shown to be located predominantly on the microvilli of digest cells. Antiserum against a nonglycosylated Escherichia coli recombinant form of Bm86 was used to avoid cross-reactivity with carbohydrate epitopes of other digest cell proteins.

Prostaglandin E2 production by the cattle tick (Boophilus microplus) into feeding sites and its effect on the response of bovine mononuclear cells to mitogen

Prostaglandin E2 (PGE2) secretion by the cattle tick Boophilus microplus into feeding sites was quantified. It was detected by the in vitro tube feeding experiment and it was determined that a semi-engorged female tick could produce and transmit 1.8 ng PGE2 into the feeding site. Using the in vitro membrane feeding system, newly molted adult ticks were also shown to secrete 0.04-0.15 ng PGE2 into the feeding site; however, female ticks produced more PGE2 than male ticks. The immune suppressive effect of PGE2 in the saliva of B. microplus on the bovine mononuclear cells (MNC) was also examined. PGE2 in the saliva was suspected of being a major component that inhibited the blastogenic response of MNC to a T-cell mitogen phytohemagglutinin. As bovine MNC are sensitive to low level concentration of PGE2, the PGE2 transmitted into feeding sites was suspected to be sufficient to produce physiological effects on the bovine host.

Feeding electrograms and fluid uptake measurements of cattle tick Boophilus microplus attached on artificial membranes

Abstract Waladde S. M., Kemp D. H. and Rice M. J. 1979. Feeding electrograms and fluid uptake measurements of cattle tick Boophilus microplus attached on artificial membranes. International Journal for Parasitology 9 : 89–95. Newly moulted adult females of the cattle tick Boophilus microplus readily attach on a modified Baudruche membrane. Apparatus design permits the media offered below the membrane to be kept at 37°C and to be changed easily. Patterns of feeding activity were recorded within 2 h of placing the ticks on the membrane by monitoring the changes in electrical resistance between a tick and the medium with a high input impedance electric circuit. Differences in patterns of sucking and salivation were related to the chemical composition of media presented below the membrane. These observations suggest that the newly discovered cheliceral taste receptors of B. microplus are able to mediate changes in feeding patterns in response to stimulation by different chemical solutions in the feeding lesion. Incorporation of phosphorus-32 into the medium allows the volume ingested by ticks to be measured. The techniques described here facilitate the study of host factors that influence attachment, engorgement and detachment of the cattle tick.

Generic approaches to obtaining efficacious antigens from vector arthropods

The development of vaccines to control ectoparasites is dependent upon the identification of key parasite antigens. While a rational, pragmatic approach to antigen identification has yielded a successful vaccine candidate from ticks, there may be problems with such an approach when dealing with other ectoparasites. As an alternative approach, the search for vaccine candidates may be facilitated by cloning and expressing parasite genes encoding proteins involved in key physiological roles. A number of criteria may be applied to short-list candidate vaccines, these being; (a) host antibodies should be able to gain access to the parasite antigen; (b) sufficient antibody must gain access to the antigen target; (c) the formation of antibody-antigen complex should disrupt the normal function of the parasite antigen (d) the antigen should share conserved structural/sequence motifs with related, characterised, proteins, thus allowing the use of recombinant DNA methods to clone and express the candidate antigen. We propose three major groups of parasite antigens which may fulfill these criteria; serine proteases, chemoreceptors/ion channels and neuropeptides.