Alan R. Walker

Structural and histochemical changes in the salivary glands of Rhipicephalus appendiculatus during feeding

Abstract The salivary glands of the brown ear tick of cattle, R. appendiculatus , from both sexes and at all stages of feeding, were examined as whole glands and as sections for ultrastructural and histochemical changes. The type 1 acinus consists of a basal labyrinth formed by the interdigitations of a central cell and four peripheral cells. These cells form a specialized border with a central constrictor cell which surrounds the acinar duct. The plasma membrane of the central cell is exposed to the duct. The type 1 acini do not appear to secrete active saliva components involved in feeding. The type 2 acini undergo a great increase in synthetic and secretory activity during feeding in both sexes and secrete a lipoprotein probably to form part of the attachment cone and also glycoproteins and esterases of unknown functions. The type 3 acini of both sexes also secrete a lipoprotein probably to form part of the attachment cone. The f cells of these acini in the females transiently secrete a glycoprotein of unknown function and then transform to become part of a water excreting unit. In the males the secretory activity of the granular cells of the type 2 and 3 acini is maintained for further attachments. The type 4 acini of the males accumulate masses of proteinaceous granules. The system of interstitial cells and intercellular spaces in types 2, 3 and 4 acini is large and increasingly active during feeding.

Risk factors in habitats of the tick Ixodes ricinus influencing human exposure to Ehrlichia phagocytophila bacteria.

Ixodes ricinus L. (Acari: Ixodida) were sampled during 1996–99 in southern Scotland, on vegetation using cloth drags, on humans by removal from clothing and on roe deer (Capreolus capreolus L.) by searching legs of culled deer. Developmental microclimate was recorded by automatic recorders and questing microclimate by portable instruments during tick collections. Ticks and deer were examined for infection with Ehrlichia phagocytophila bacteria (Rickettsiales) using microscopy and polymerase chain reaction. This pathogen causes tick‐borne fever of sheep in Europe and human granulocytic ehrlichiosis in North America, but in Europe human clinical ehrlichiosis due to E. phagocytophila has not been recorded despite serological evidence of exposure.

Histology of digestion in nymphs of Rhipicephalus appendiculatus fed on rabbits and cattle naive and resistant to the ticks

Abstract Nymphs feeding on ears of four rabbits and four calves ( Bos taurus ) were examined during first and third or fourth infestations and also during the moulting period. The gut caecae were removed and examined by histochemistry and light and electron microscopy. Attachment sites of the nymphs were biopsied from all hosts and cell counts made by light microscopy. Resistance was expressed by reduction in numbers of ticks completing engorgement and reduced engorgement weights. The gut was comprised of a proliferative stem cell; a digestive cell that differentiated into a sessile type ingesting by pinocytosis and a motile type ingesting by phagocytosis; and a cell secreting a glycoprotein with acid phosphatase activity into the lumen. The gut grew during the early stages of feeding to accommodate the expansion during engorgement. On rabbits and cattle resistant to ticks the stem cells were damaged, with moribund nuclei and poorly differentiated cytoplasm. Thus there were fewer digestive and secretory cells and the gut did not expand to accommodate a full blood meal. The attachment sites were dominated by mononuclear cells and neutrophils in both host species at the first infestations but at the third or fourth infestations there was a considerable increase in proportions of eosinophils and basophils. Host granulocytes were traced to the lumen of the tick gut and to motile digestive cells which destroyed them by phagocytosis.

Age structure of a population of Ixodes ricinus (Acari: Ixodidae) in relation to its seasonal questing

Three sites in south west Scotland, each having one deciduous and one coniferous woodland with populations of roe deer, were sampled over three years for Ixodes ricinus Linnaeus ticks using cloth drags. Nymphs and adult ticks were age graded by dissection of gut and Malpighian tubules and staining for lipid with Sudan red. Temperature and relative humidity were recorded in developmental and questing microclimates. Two annual cohorts of both nymphs and adults were found. For both instars one cohort became apparent in early spring and the other in autumn. The timing of the cohorts of nymphs corresponded to peaks of numbers questing, but neither females nor males had any clear seasonal pattern of questing. The maximum life of questing nymphs and adults was three to four months. Nymphs and adults were found questing at all months of the year and at temperatures ranging from 3.5 degrees C to 29.3 degrees C. Questing of larvae in summer peaks of numbers was positively correlated with temperature and negatively correlated with relative humidity. These results are explained by a descriptive model which invokes behavioural and morphogenetic diapause. Tests of the model are discussed and it is proposed that the main adaptive advantage of the complex life cycle is to permit moulting and oviposition to occur at favourable summer temperatures in ticks that have engorged at any time of the year.

Natural prevalence of infection with Ehrlichia (Cytoecetes) phagocytophila of Ixodes ricinus ticks in Scotland.

Ixodes ricinus nymphs and adults were collected from vegetation and from sheep at four sites in Scotland typical of areas endemic for tick-borne fever in sheep caused by infection with Ehrlichia (Cytoecetes) phagocytophila (Rickettsiales). The great majority of ticks examined was from woodland sites adjacent to sheep farms where there was a high probability of them feeding on roe deer (Capreolus capreolus) in a non-domestic focus of infestation and infection. Ticks were examined for infection by five methods. Batches of ticks were examined either by feeding on susceptible sheep or by feeding on rabbits and then prepared as stabilate which was inoculated into susceptible sheep. The sheep were monitored for clinical signs of tick borne fever. Batches of ticks were examined by polymerase chain reaction for Ehrlichia phagocytophila. Salivary glands were dissected out and stained by the Feulgen method to detect Ehrlichia masses, and were examined by indirect fluorescent antibody test. Each of the methods detected infection in ticks and the prevalence of infection in nymphs with the various methods ranged from >0.25% to 2.0%. Small samples of adults examined by Feulgen staining of salivary glands indicated infection prevalences of 2.1% in males and 1.6% in females. It is considered that these low infection prevalences may be typical of natural foci of infection where deer could be a major host of ticks and E. phagocytophila.

The maintenance and survival of Theileria annulata in colonies of Hyalomma anatolicum anatolicum

Procedures are described for the colonization of Hyalomma anatolicum anatolicum ticks using gerbils, rabbits and cattle as hosts. On rabbits H. a. anatolicum undergoes a two-host cycle but methods are described for obtaining either unfed or engorged nymphs. Data are given on the life cycle timings and the numbers and timings of tick feeding regimes. The infection of H. a. anatolicum with two strains of Theileria annulata is described and methods given for monitoring the development and survival of T. annulata in H. a. anatolicum. Data are presented indicating the optimum maintenance conditions for T. annulata in H. a. anatolicum. Piroplasm parasitaemias in cattle greater than 2% gave high infection rates in adult ticks if engorged nymphs were moulted at 28 degrees C for 28 days. The Theileria will survive for ten months in such adults stored at 12 degrees C after moult, and when fed the ticks will produce maximum numbers of sporozoites on the third day of feeding, whatever their age. The moult of engorged nymphs is retarded at 18 degrees C but the Theileria in such ticks will develop normally when the ticks are moulted at 28 degrees C after four months storage at 18 degrees C.

Eradication and control of livestock ticks: biological, economic and social perspectives

Comparisons of successful and failed attempts to eradicate livestock ticks reveal that the social context of farming and management of the campaigns have greater influence than techniques of treatment. The biology of ticks is considered principally where it has contributed to control of ticks as practiced on farms. The timing of treatments by life cycle and season can be exploited to reduce numbers of treatments per year. Pastures can be managed to starve and desiccate vulnerable larvae questing on vegetation. Immunity to ticks acquired by hosts can be enhanced by livestock breeding. The aggregated distribution of ticks on hosts with poor immunity can be used to select animals for removal from the herd. Models of tick population dynamics required for predicting outcomes of control methods need better understanding of drivers of distribution, aggregation, stability, and density-dependent mortality. Changing social circumstances, especially of land-use, has an influence on exposure to tick-borne pathogens that can be exploited for disease control.

Histological study of the attachment sites of adult Rhipicephalus appendiculatus on rabbits and cattle.

Abstract Biopsies of tick attachment sites on ears of five rabbits and four calves ( Bos taurus ) were taken at first and third infestations and were examined by histochemistry, light microscopy and electron microscopy. Resistance was expressed by all hosts by reduction of tick engorgement weights. Attachment sites at first infestations on rabbits and cattle were similar and were characterized by an acute inflammatory abscess with a preponderance of neutrophils and macrophages infiltrating the site. Attachment sites at third infestations on rabbits and cattle were similar and were characterized by infiltration of neutrophils and macrophages and a 2- to 5-fold increase in the proportions of eosinophils and basophils. Lymphoblasts and plasmacytes were found in third infestation sites. In rabbits there was much necrosis and in cattle there were large intraepidermal pustules in third infestations. The cement attachment cone of the ticks was of lipoprotein and contained aminopeptidase. Salivary glycoproteins and esterases were detected in the attachment sites.

The salivary glands of Hyalomma anatolicum anatolicum: Nature of salivary gland components and their role in tick attachment and feeding

Abstract Gill H. S. and Walker A. R. 1988. The salivary glands of Hyalomma anatolicum anatolicum : nature of salivary gland components and their role in tick attachment and feeding. International Journal for Parasitology 18 : 83–93. A study was made of the histochemistry of the salivary glands and tick feeding sites of H. a. anatolicum during feeding. The attachment cement of H. a. anatolicum was lipoprotein in nature and appeared to have been derived from a cells of type II, and d and e cells of type III acini in the salivary glands. The secretory granules of these cells had similar histochemical properties to the cement. Deposits of glycoprotein materials were located at tick feeding sites in dermal tissues adjacent to the mouthparts and not in the attachment cement. This polysaccharide material is presumed to be secreted by b and c cells of type II acini. Strong aminopeptidase and moderate acid phosphatase activity was found localized in the cement cone and not in the tissues. Deposits of non-specific esterase material, probably derived from b , c 1 and c 3 cells of type II acini in the glands were demonstrated at tick feeding sites adjacent to the mouthparts. It is presumed that the esterase might increase vascular permeability in the dermis of the hosts by its direct action on mast-cells. A steady increase in ATPase and alkaline phosphatase activity in the interstitial cells and the lumenal border of type II and III acini during feeding suggests a possible role in osmoregulation. Despite the strong reactions for several oxidoreductases and other phosphatases in the salivary glands of H. a. anatolicum , no activity for these enzymes was detected in the cement or at feeding sites. The possible role of these salivary components in feeding is discussed.

Genotypic diversity, a survival strategy for the apicomplexan parasite Theileria parva

The tick-borne protozoan parasite Theileria parva causes East Coast fever (ECF), a severe lymphoproliferative disease of cattle that is a major constraint to the improvement of livestock in eastern, central and southern Africa. Studies in cattle experimentally infected with T. parva have shown that the protective cytotoxic T lymphocyte (CTL) response is tightly focused, with individual animals recognizing only one or two dominant antigens, the identity of which varies with MHC class I phenotype. It is well known that cross-protection between T. parva stocks is limited, but precise evaluation of genetic diversity in field populations of the parasite has been hampered by a lack of molecular markers spanning the genome. A recently described panel of satellite markers has provided evidence for substantial genotypic diversity and recombination but does not provide cover for large segments of the genome. To address this deficiency, we undertook to identify additional polymorphic markers covering these regions and we report herein 42 newly identified PCR-RFLP markers distributed across the 4 T. parva chromosomes, as well as 19 new satellite markers for chromosomes 1 and 2. This brings the total number of available polymorphic markers to 141 for the 8.5 Mb genome. We have used these markers to characterise existing parasite stabilates and have also shown that passage of the parasite through naïve cattle and ticks can lead to substantial changes of parasite populations in resulting stabilates. These markers have also been used to show that passage of mixed parasites through an immunised calf results in the removal of the immunising genotype from the parasite population produced by ticks fed on this animal.