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Biochemical investigation into the toxic salivary secretion of the tick, Ornithodoros savignyi Audouin (1827)
Get PDFThesis (DSc (Agric))--University of Pretoria, 1976.Ticks, including the Sand tampan Ornithodoros savignyi, pre= sent important economic problems over a large area in the Republic of South Africa (S 1), In all known diseases of which ticks are the vectors, the causal agents (protozoa, rickettsias, Ana plasms, viruses, spirochetes and bacteria) have been identified with the exception of tick toxicoses in which pathogenic agents are evidently not involved (S 2) - Thus, the causal agents associated with tick toxicoses caused by the Sand tampan have not been identified. These causal agents appear to develop only in the invertebrate host (the ticks) and their timely removal from the vertebrate hosts, is followed by a subsidence of the symptoms. Recovered animals develop a durable immunity (S 3) The observations regarding tick paralysis and sweating sickness are most likely also applicable to tick toxicosis caused by the Sand tampan. Claims of toxins present in the salivary secretions of ticks have been made (S 1). Evidence supporting these claims arc presented in this thesis. Information is provided concerning the chemical composition of the salivary secretion and ~n isolation procedure by means of which a toxic component present in the secretion may be obtained in a pure form. In addition, some chemical components and activities in the oral secretion of the bontbosluis, l\. nblyomrna hebraeurn, were investigated and the results reported. Sand tampan ticks used for the investigations were collected in the North West Cape Kalahari with traps baited with solidified carbon dioxide (S 5), Oral secretions of the ticks were obtained by parasympathetic stimulation as described by Howell (S 6), Some properties of the secretion are shown in Table S l, the enzymatic activities undoubtedly enhance the entry of the toxin into the tissues of the host. Isolation of the toxic activity was achieved by means of gel and ion exchange chromatography and isoelectric focusing. The toxin was found to be homogeneous according to ultra-centrifugal and gel-electrophoretic techniques and isoelectric focusing. Some characteristics of the toxic component are presented in Table S 2, Removal of the carbohydrate moiety or s-carboxymethylation of the toxin results in a complete loss of toxic activity. A non-toxic glycopeptide which showed most of the above characteristics was found in the salivary secretion. It differed slightly from the toxin with respect to the isoelectric point (0,3 units lower) and amino acid sequence (Table S 3). A detailed structural comparison of these two glycopeptides could aid in the elucidation of the structural requirements essential for toxicity. The syndromes produced by the salivary secretion (S 25) indicate that the toxin may exert its effect via increased capillary permeability or fragility. Since a method has been developed for the purification of the toxin and its chemical nature has been partially described, the biochemical lesion(s} can now be investigated. The total free amino acids present in the salivary secretion of the bontbosluis, Amblyomma hebraeum was determined. A total of 1553 μg of amino acids per ml secretion was found. This value is ·approximately 3 times higher than the value for the Sand tampan and approximately ·20 times higher than for human saliva. The salivary secretion of this tick was fractionated by gel chromatography. One of the fractions showed weak hyaluronidase activity and was found to be homogeneous according to sedimentation equilibrium centrifugation. The molecular mass, determined by the latter method was found to be 9680. The amino acid composition of the fraction revealed, with a few exceptions, similarity with that of hyaluronidase isolated from bovine testicular tissue.BiochemistryDSc (Agric)Unrestricte
The detection and isolation of a paralysis toxin present in Argas (Persicargas) walkerae
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.One-day-old leghorn chickens were used in a laboratory assay to determine the toxicity of crude extracts of the tick Argas (Persicargas) walkerae and of fractions obtained during the isolation procedure. Extracts of unfed and engorged larvae, nymphae and females were tested using this in vivo test system. Only extracts of replete A. (P.) walkerae larvae produced paralysis.
A toxic fraction was isolated from replete larval extracts by gel-permeation and ion-exchange chromatography. This fraction with a pI of 4,5 showed 2 major bands corresponding to a Mᵣ of 32 kDa and 60 kDa after SDS-polyacrylamide gel electrophoresis.mn201
Investigation into the participation of male pheromones of Rhipicephalus evertsi evertsi during infestation
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.It was shown in this study that Rhipicephalus evertsi evertsi males emit pheromones in the preparasitic as well as in the parasitic phases. Conspecific unengorged male and female images exhibited a high response to water-soluble volatiles collected from unengorged males. A vigorous assembly of unfed female R. evertsi evertsi occurred on filter paper discs which had been in contact with either unfed or 5- and 9-day fed males. Progressively increasing percentages of conspecific unengorged males reacted to organic soluble volatiles obtained from 5- and 9-day fed males as well as to hexane washings of intact males which had fed for 9 days. It is concluded that R. evertsi evertsi males participate not only as pheromone receptive, but also as pheromone emitting, partners in the intraspecific communication of this tick species
Identification of the antigenic proteins of Cowdria ruminantium
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.Immunoblotting of Cowdria ruminantium proteins with sheep or bovine antiserum identified 2 antigenically conserved proteins, one being an immunodominant 31 kDa and the other a minor 27 kDa protein. These proteins are present in the electrophoretic profiles of the Welgevonden, Ball 3 and Kwanyanga stocks and are recognized by sheep antiserum to the Welgevonden, Ball 3, Kwanyanga, Mali, Comoro, Breed, Germishuys, Kümm and Mara stocks and by bovine antiserum to the Welgevonden stock of C. ruminantium. The stocks did not reveal identical or unique antigenic properties which could explain differences in pathogenicity and cross-immunity observed amongst the various stocks of C. ruminantium.mn201
The presence of Cowdria ruminantium antigen in various tissues of Amblyomma hebraeum imagines as detected by an enzyme-linked immunosorbent assay
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.Investigation into the presence of C. ruminantium antigen, using an enzyme-linked immunosorbent assay
(ELISA) in various tick tissues and haemolymph of adult Amblyomma hebraeum ticks revealed that the organism
invades a number of body parts and can be demonstrated in A. hebraeum. In females, the gut, salivary glands,
hypodermis and synganglion and in males, the salivary glands and gut showed the highest concentration.lmchunu2014mn201
An investigation into the emission sites of the volatile pheromone produced by females of Argas (Persicargas) walkerae
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.By leaving open in tum the genital orifice, coxal apertures and anus , the present study has shown that engorged female Argas walkerae release at least 1 volatile pheromone mainly, or maybe exclusively, from the genital aperture. Although this aggregation-inducing pheromone is perceived by all post-larval stages, engorged males of A. walkerae react most strongly. A possible, but only slight, emission through the anus and the coxal aperatures is also indicated. Most probably, this airborne biosubstance is the primary directing and orienting pheromone in the interrelations between members of this tick species
Sweating sickness : relative curative effect of hyperimmune serum and a precipitated immunoglobulin suspension and immunoblot identifiation of proposed immunodominant tick salivary gland proteins
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.Although of low morbidity, sweating sickness is readily induced in calves by infestation with positive Hyalomma truncatum adult ticks. This epitheliotrophic disease has no specific cure except by the administration of hyperimmune serum obtained from animals which have recovered and are subsequently immune to the disease. Treatment with hyperimmune serum, however, has associated problems of donor availability, possible serum contamination and i.v. administration of a relatively large volume. This paper compares the treatment and cure of sweating sickness using unrefined hyperimmune serum and that of an experimental suspension. The latter proved relatively inefficient probably due to a low concentration of effective immunoglobulins. Immunoblot analyses of the sera of affected animals, using tick salivary glands as antigen during the course of the trial revealed 4 tick salivary gland proteins with molecular masses of between 27-33 kDa. These are proposed as being associated with sweating sickness immunodominance.mn201
Detection of Cowdria ruminantium antigen and antibody during the course of heartwater disease in sheep by means of an enzyme-linked immunosorbent assay.
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.An enzyme-linked immunosorbent assay was used to detect Cowdria ruminantium antibodies during the course of heartwater disease. IgM antibodies reached a maximum on the 4th day after infection and disappeared on the 7th day. IgG antibodies first appeared on the 8th day and continued to increase during the remainder of the
observation period of 28 days. The presence of C. ruminantium in the blood fractions of diseased animals was demonstrated by an enzyme-linked immunosorbent assay. The earliest day of C. ruminantium antigen detection was in plasma and serum on the 4th day after inoculatton. Of all the blood fractions investigated, the red blood cells showed the highest concentration, and this reached a maximum on the 12th day after infection
Investigations into the function and chemical compositions of the porose areas secretion of Rhipicephalus evertsi evertsi during oviposition
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.Major differences were observed in hexane, ethanol and butanol extracts of eggs obtained from Rhipicephillus evertsi evertsi females in which the porose areas functioned normally (AP⁺ eggs) and from R. evertsi evertsi in which the porose areas were selectively destroyed by electrocautery (APˉeggs). Mass yields and UV spectra of the hexane extracts were similar for AP⁺ and APˉeggs. The UV spectra changed only slightly in the 294-320 nm range with respect to time and temperature. High performance liquid chromatography revealed 2 components which originate from the porose areas. Mass spectroscopy of these components indicated the presence of aliphatic and phenolic groups. The ethanol and butanol extracts showed quantitative but no qualitative differences with respect to AP⁺ and APˉeggs. Electrophoretic fractionation of the butanol extracts revealed the presence of proteins in the secretion of the porose areas. Apart from this information on the chemical composition of the secretion, no indication was obtained of their function during oviposition of R. evertsi evertsi
The detection of antibodies to Cowdria ruminantium in serum and C. ruminantium antigen in Amblyomma hebraeum by an enzyme-linked immunosorbent assay
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Adobe Acrobat XI Pro was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.A sensitive and reliable enzyme-linked immunosorbent assay for the detection of antibodies to Cowdria ruminantium in serum and C. ruminantium antigen in Amblyomma hebraeum nymphae is described. For the screening of antibodies, C. ruminantium from A. hebraeum nymphae, partially purified by wheat-germ lectin affinity chromatography, was used as antigen. To screen nymph populations, sera from either Ball 3 strain-infected sheep or Kumm-strain infected mice were used. By using appropriate controls the assays were rendered specific with respect to C. ruminantium.mn201
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