An update on the ecological distribution of Ixodid ticks infesting cattle in Rwanda: countrywide cross-sectional survey in the wet and the dry season

As part of the epidemiological studies aimed at developing an East Coast fever (ECF) immunisation control strategy, which combines an infection and treatment method with strategic tick control, a countrywide tick survey was carried out in both the dry and the wet season to determine the abundance and the dynamics of the tick populations infesting cattle in Rwanda. Six Ixodid tick species where identified from a total of 12,814 tick specimens collected. Rhipicephalus appendiculatus, the main vector of ECF was the most abundant (91.8%) followed by Boophilus decoloratus (6.1%) and Ambyomma variegatum (1.2%). Few ticks from the three other less economically important Ixodid species (Rhipicephalus compositus, R. evertsi evertsi and Ixodes cavipalpus) were recovered. Both adult and immature stages of the most dominant tick species were found to be widespread with a year round presence. The numbers of ticks were high in low land and medium zones and declined markedly in the higher regions of Rwanda. The geographical distribution of various tick species throughout the country and their epidemiological implications are discussed.     

Survival of Rhipicephalus appendiculatus (Acarina: Ixodidae) and persistence of Theileria parva (Apicomplexa: Theileriidae) in the field

Newson R. M., Chiera J. W., Young A. S., Dolan T. T., Cunningham M. P. and Radley D. D. 1984: Survival of Rhipicephalus appendiculatus (Acarina: Ixodidae) and persistence of Theileria parva (Apicomplexa: Theileriidae) in the field. International Journal for Parasitology14; 483–489. Two paddocks with populations of the African brown ear tick (Rhipicephalus appendicuiatus), one of which carried virulent Theileria parva (the causative organism of East Coast fever [ECF] of cattle), were left unstocked for periods of 338 and 354 days. Groups of 1–3, ECF-susceptible cattle were then introduced eight times during the following year to assess the tick and disease challenge, The ticks were also monitored continuously on the ground. The test cattle developed fatal ECF from adult ticks which had fasted for up to 554 days. A non-pathogenic, antigenicaliy distinct, Theileria species was also detected which was still transmitted by adult ticks after 600 days. The mean survival time of unfed larvae was 175 days (max. 240 days); unfed nymphs 270 and 450 (max. 540 days); unfed adults 420 days (max. 730 days), with females in a clear majority after 270 days.     

Resistance in cattle againstRhipicephalus appendiculatus with an assessment of cross-resistance toR. pulchellus (Acari: Ixodidae)

A comparison of the yield of engorged nymphs ofRhipicephalus appendiculatus andR. pulchellus was carried out between cattle highly resistant toR. appendiculatus but susceptible toR. pulchellus, and cattle and rabbits susceptible to both species.Resistant cattle yielded fewer and smallerR. appendiculatus nymphs than either susceptible cattle or rabbits. More nymphs of both tick species dropped from susceptible cattle but no differences were detected in the mean weight of engorgedR. pulchellus nymphs fed onR. appendiculatus-resistant and susceptible cattle, although resistant animals yielded more nymphs ofR. pulchellus than ofR. appendiculatus.An acute inflammatory reaction, accompanied by intense pruritus and grooming, was observed in the ears of the resistant cattle infested withR. appendiculatus, but no reaction was detected in the opposite ears whereR. pulchellus was feeding.The low cross-resistance between these two closely related tick species suggests that different antigens may be involved in the development of resistance in cattle to different tick species.     

Cryopreservation of infective particles of Theileria parva

Cryopreservation of infective particles of Theileria parva. International Journal for Parasitology3: 583–587. Infective particles of Theileria parva, the causative organism of East Coast fever of cattle, were obtained from infected Rhipicephalus appendiculatus ticks, either by using an in vitro feeding technique or by grinding the ticks in a suitable medium. If foetal calf serum containing 15% glycerol (v/v) was added to the infective material and it was then distributed either to glass capillary tubes (in vitro tick feed) or glass tubes (ground tick supernate) it could be slowly frozen to either ?80°C or ?196°C without loss of viability. Stabilates, tested by rapid thawing and inoculation into ECF-susceptible cattle, remained viable for up to a year at these temperatures.     

Salivary gland of the tick vector (R. appendiculatus) of East Coast fever. I. Ultrastructure of the type III acinus

The brown ear tick Rhiplcephalus appendiculatus is the vector for East Coast fever, a disease that seriously limits livestock production in East Africa. The sporozoites of the infectious agent Theileria parva develop in the tick salivary gland. This paper describes the organization of the type III acinus of the gland and establishes unambiguous ultrastructural criteria for identification of the three secretory cell types: the d-cell, e-cell and f-cell. These observations are basic to exploration of possible cell-type specificity of the invading theileria and other aspects of host-parasite relations.     

Rhipicephalus appendiculatus and R. zambeziensis (Acari: Ixodidae) from Zambia: a molecular reassessment of their species status and identification

The closely related species Rhipicephalus appendiculatus and R. zambeziensis co-occur in a wide area in Zambia. In this area, specimens of both species have been collected on the same individual host at the same time. In addition, specimens that are morphologically intermediate between R. appendiculatus and R. zambeziensis have been found in this area. These observations cast some doubt on the species status of R. appendiculatus and R. zambeziensis. Because the two taxa have varied influences on the epidemiology of East Coast fever (ECF), a cattle disease for which they act as vectors, it is essential that their taxonomic status is clarified and their identification is accurate. Therefore a phylogenetic analysis of nucleotide sequences of the ITS2 and a fragment of the mitochondrial 12S rDNA was performed to reassess the specific status of both taxa. This revealed two well supported clades coinciding with R. appendiculatus and R. zambeziensis as two separate species. In order to facilitate species identification a PCR-RFLP diagnostic assay was developed based on BauI digestion of the ITS2 gene. This assay produced clear diagnostic banding patterns for the two species and is applicable over a wide range of both species’ distribution. Nucleotide sequence data reported in this paper are available in the GenBank™ under the accession numbers DQ849203–DQ849274.     

Electrospun micro- and nanofiber tubes for functional nervous regeneration in sciatic nerve transections

Background

Rhipicephalus (Boophilus) spp. ticks economically impact on cattle production in Africa and other tropical and subtropical regions of the world. Tick vaccines constitute a cost-effective and environmentally friendly alternative to tick control. The R. microplus Bm86 protective antigen has been produced by recombinant DNA technology and shown to protect cattle against tick infestations.

Results

In this study, the genes for Bm86 (R. microplus), Ba86 (R. annulatus) and Bd86 (R. decoloratus) were cloned and characterized from African or Asian tick strains and the recombinant proteins were secreted and purified from P. pastoris. The secretion of recombinant Bm86 ortholog proteins in P. pastoris allowed for a simple purification process rendering a final product with high recovery (35–42%) and purity (80–85%) and likely to result in a more reproducible conformation closely resembling the native protein. Rabbit immunization experiments with recombinant proteins showed immune cross-reactivity between Bm86 ortholog proteins.

Conclusion

These experiments support the development and testing of vaccines containing recombinant Bm86, Ba86 and Bd86 secreted in P. pastoris for the control of tick infestations in Africa.     

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.     

Cattle ticks of the genera <Emphasis Type="Italic">Rhipicephalus</Emphasis> and <Emphasis Type="Italic">Amblyomma</Emphasis> of economic importance in Tanzania: distribution assessed with GIS based on an extensive field survey

In order to implement a robust integrated tick and tick-borne disease control programme in Tanzania, based on ecological and epidemiological knowledge of ticks and their associated diseases, a national tick and sero-surveillance study was carried out in all 21 regions of the mainland, as well as on Mafia Island, between 1998 and 2001. The current distributions of Rhipicephalus appendiculatus, R. pravus, Amblyomma variegatum, A. gemma, and A. lepidum are illustrated and discussed. Tick distribution maps were assessed using the Weights-of-Evidence method (WofE), and employing temperature, humidity, NDVI, rainfall, and land-cover predictive data. Ground-truthing was done to check correspondence both of the data employed in prediction with land-cover characteristics discerned in the field as well as of the surveyed and predicted tick distributions. Statistical methods were used to analyse associations of the tick species with their environment, cattle density, and other ticks. Except for R. appendiculatus, no appreciable changes were demonstrated in the predicted and observed tick distributions compared to the existing maps that originated in the 1950–1960s. Cattle density influenced the distribution of A. variegatum and, to a certain extent, of A. lepidum, but had no appreciable influence on the distribution of any of the other ticks discussed in this paper, neither did livestock movement. Distinct differences for environmental requirements where observed between different tick species within the same genus. The predictive maps of R. appendiculatus and R. pravus suggest their mutually exclusive distribution in Tanzania, and simultaneous statistical analysis showed R. pravus as a greater specialist. Of the three Amblyomma species, A. variegatum is the most catholic tick species in Tanzania, while both A. gemma and A. lepidum belong to the more specialized species. Despite dissimilar habitat preferences, all three Amblyomma spp. co-exist in central Tanzania, where very heterogeneous habitats may simultaneously satisfy the environmental requirements of all three species. The current study, conducted about 4 decades after the last major survey activities, has shown that changing livestock policies, unrestricted livestock movement and a continuous change in climatic/environmental conditions in Tanzania have brought about only limited changes in the distribution patterns of R. appendiculatus, R. pravus and the three Amblyomma species investigated. Whether this observation indicates a relative indifference of these ticks to environmental and/or climate changes allows room for speculation.     

Further evidence for geographic differentiation in R. appendiculatus (Acari: Ixodidae) from Eastern and Southern provinces of Zambia

Studies in the biology, ecology and behaviour of R. appendiculatus in Zambia have shown considerable variation within and between populations often associated with their geographical origin. We studied variation in the mitochondrial COI (mtCOI) gene of adult R. appendiculatus ticks originating from the Eastern and Southern provinces of Zambia. Rhipicephalus appendiculatus ticks from the two provinces were placed into two groups on the mtCOI sequence data tree. One group comprised all haplotypes of specimens from the Eastern province plateau districts of Chipata and Petauke. The second group consisted of a single haplotype of specimens from the Southern province districts and Nyimba, an Eastern province district on the fringes of the valley. This variation provides additional evidence to the earlier observations in the 12S rDNA and ITS2 data for the geographic subdivision of R. appendiculatus from Southern province and Eastern province plateau. The geographic subdivision further corresponds with differences in body size and diapause between R. appendiculatus from these geographic areas. The possible implications of these findings on the epidemiology of East Coast fever (ECF) the disease for which R. appendiculatus is one of the vectors are discussed. Nucleotide sequence data reported in this paper are available in the GenBank^TM under the Accession Numbers DQ859259–DQ859266     

Monitoring of naturally acquired and artificially induced immunity toAmblyomma variegatum andRhipicephalus appendiculatus ticks under field and laboratory conditions

The ability of rabbits, goats and cattle to acquire immunity to the ixodid ticksAmblyomma variegatum andRhipicephalus appendiculatus was studied under laboratory and field conditions. Rabbits were successfully immunized with crude salivary gland extract (SGE) and midgut extract (ME) obtained from flat or partly fed femaleR. appendiculatus ticks. The lowest numbers of larvae were produced by females fed on rabbits immunized with unfed midgut extract. Similar reductions in larval production could be induced after three infestations of rabbits with adultR. appendiculatus. Also, successive feedings of nymphs ofR. appendiculatus on rabbits resulted in significantly reduced engorgement weights. Skin testing with SGE induced delayed-type hypersensitivity reactions, which could be correlated with immunity toR. appendiculatus in rabbits. Moreover, circulating antibodies were detected in rabbits with an ELISA using SGE ofR. appendiculatus.Immunity toA. variegatum nymphs could be induced in rabbits by repeated infestations, but this failed in goats. Immunization of goats with midgut extract from adultA. variegatum did not protect against subsequent nymphal challenge, but strong skin reactions were noticed when adults ticks fed on immunized goats. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of SGE and ME fromA. variegatum revealed the presence of 48 protein bands in SGE and 29 bands in midgut extract. Western blotting employing serum from a rabbit immune toR. appendiculatus recognized a number of bands in SGE fromR. appendiculatus, but also in SGE ofA. variegatum.Immunity acquired by cattle to ixodid tick infestations under field conditions was monitored by skin testing with SGE and western blot analysis. In general, cattle with the lowest tick numbers manifested the strongest delayed-type hypersensitivity responses. Finally, western blot analysis employing sera from tick-infested and tick-naive cattle could not be related to actual immune status.     

East Coast fever: the infectivity for cattle of infective particles of Theileria parva harvested in various substrates

Cunningham M.P., Brown C.G.D., Burridge M.J., Joyner L.P. and Purnell R.E. 1973. East Coast fever : the infectivity for cattle of infective particles of theileria parva harvested in various substrates. International Journal for Parasitology3: 335–338. Female Rhipicephalus appendiculatus ticks infected with Theileria parva were pre-fed on rabbits for 4 days before being removed and restrained on ‘plasticine’. They were then fed for 2-h periods on capillary tubes containing various substrates, the contents of which were subsequently inoculated into East Coast fever-susceptible cattle. Using this technique, precolostral calf serum and Eagle's Minimum Essential Medium, with the addition of Bovine Albumin Powder, were selected as substrates suitable for further laboratory investigation on East Coast fever as they were acceptable to the ticks and supported the viability of the parasites.     

Influence of the Biotope on the Tick Infestation of Cattle and on the Tick-Borne Pathogen Repertoire of Cattle Ticks in Ethiopia

Background

The majority of vector-borne infections occur in the tropics, including Africa, but molecular eco-epidemiological studies are seldom reported from these regions. In particular, most previously published data on ticks in Ethiopia focus on species distribution, and only a few molecular studies on the occurrence of tick-borne pathogens or on ecological factors influencing these. The present study was undertaken to evaluate, if ticks collected from cattle in different Ethiopian biotopes harbour (had access to) different pathogens.

Methods

In South-Western Ethiopia 1032 hard ticks were removed from cattle grazing in three kinds of tick biotopes. DNA was individually extracted from one specimen of both sexes of each tick species per cattle. These samples were molecularly analysed for the presence of tick-borne pathogens.

Results

Amblyomma variegatum was significantly more abundant on mid highland, than on moist highland. Rhipicephalus decoloratus was absent from savannah lowland, where virtually only A. cohaerens was found. In the ticks Coxiella burnetii had the highest prevalence on savannah lowland. PCR positivity to Theileria spp. did not appear to depend on the biotope, but some genotypes were unique to certain tick species. Significantly more A. variegatum specimens were rickettsia-positive, than those of other tick species. The presence of rickettsiae (R. africae) appeared to be associated with mid highland in case of A. variegatum and A. cohaerens. The low level of haemoplasma positivity seemed to be equally distributed among the tick species, but was restricted to one biotope type.

Conclusions

The tick biotope, in which cattle are grazed, will influence not only the tick burden of these hosts, but also the spectrum of pathogens in their ticks. Thus, the presence of pathogens with alternative (non-tick-borne) transmission routes, with transstadial or with transovarial transmission by ticks appeared to be associated with the biotope type, with the tick species, or both, respectively.     

Ecology and phenology of ticks in Zambia: Seasonal dynamics on cattle

A study of the seasonality and infestation rates of ticks was carried out in 11 cattle herds in different ecological habitats in Zambia between 1980 and 1982. Wherever possible supplementary data were obtained from opportunistic collections from cattle and other hosts.Analysis of over 1000 tick collections from cattle indicated that infestation rates of the most important species,Amblyomma variegatum andRhipicephalus appendiculatus vary in different ecological habitats: (i) In Western Province, infestations are much lower than elsewhere; (ii) in Central and Southern Provinces, moderate to high infestations occur; and (iii) in Eastern Province,R. appendiculatus numbers are generally low andA. variegatum numbers are moderate.These two species, however, have similar life cycles throughout their range with one generation per year. Larvae occur mainly from March to May, nymphae from May to September, and adults ofA. variegatum from October to December and ofR. appendiculatus from December to April. Boophilus decoloratus appears to have two to four generations per year but is uncommon during the rainy season. In some areas in central ZambiaRhipicephalus compositus adults are seasonally common in September–October whereasRhipicephalus evertsi is more or less ubiquitous. Low to moderate infestations ofHyalomma truncatum andHyalomma rufipes occur in most areas.At least 14 other less common or rare species ofRhipicephalus, Amblyomma, Haemaphysalis andIxodes were taken infrequently from cattle. These and other host-specific species were also collected from dogs, sheep, various wildlife hosts and the environment.Infestation rates, seasonality and host-relationship of tick species are discussed in relation to their ecology. Relevant biosystematic and disease relationships are reviewed briefly. The baseline data derived from this study are adequate for integrated analysis with those from other ecological and economic investigations to formulate tick control strategies.     

Effect of different East Coast Fever control strategies on fertility, milk production and weight gain of Sanga cattle in the Central Province of Zambia

Five different East Coast Fever (ECF) (Theileria parva infection) control strategies, based on tick control and/or ECF immunization, were tested in groups of traditionally managed Sanga cattle in the Central Province of Zambia over a period of 2.5 years. Two groups were kept under intensive tick control (sprayed weekly), one group immunized and one non-immunized. Two further groups were under no tick control, one group immunized and one non-immunized, while a fifth group was immunized against ECF and maintained under strategic tick control (18 sprays per year). Tick control increased milk production and weight gain but not fertility. Immunization had neither marked detrimental nor beneficial effects on the cattle productivity. The combination of strategic tick control and immunization resulted in the highest level of production and at the same time reduced the potential risk from other tick-borne diseases.     

Repellent and acaricidal properties of Ocimum suave against Rhipicephalus appendiculatus ticks

An oil extracted from the leaves of a tropical shrub Ocimum suave was found to repel as well as kill all stages of the tick Rhipicephalus appendiculatus. In an in vitro bioassay for the larvae, the LC₅₀ of the oil in liquid paraffin was 0.024%. A 10% solution was found to kill all immatures and more than 70% of adults feeding on rabbits. Rabbits were protected for 5 days against attaching larvae using a 10% solution. Preliminary experiments undertaken with cattle kept in the field suggest that the oil may have potential in tick control, and a role in integrated tick management.     

Construction of a genetic map for Theileria parva: identification of hotspots of recombination

The tick-borne protozoan parasite Theileria parva is the causal agent of East Coast Fever (ECF), a severe lymphoproliferative disease of cattle in eastern, central and southern Africa. The life cycle of T. parva is predominantly haploid, with a brief diploid stage occurring in the tick vector that involves meiotic recombination. Resolved genetic studies of T. parva are currently constrained by the lack of a genome-wide high-definition genetic map of the parasite. We undertook a genetic cross of two cloned isolates of T. parva to construct such a map from 35 recombinant progeny, using a genome-wide panel of 79 variable number of tandem repeat markers. Progeny were established by in vitro cloning of cattle lymphocytes after infection with sporozoites prepared from Rhipicephalus appendiculatus ticks fed on a calf undergoing a dual infection with the two clonal parental stocks. The genetic map was determined by assigning individual markers to the four chromosome genome, whose physical length is approximately 8309 kilobasepairs (Kb). Segregation analysis of the markers among the progeny revealed a total genetic size of 1683.8 centiMorgans (cM), covering a physical distance of 7737.62 Kb (∼93% of the genome). The average genome-wide recombination rate observed for T. parva was relatively high, at 0.22 cM Kb⁻¹ per meiotic generation. Recombination hot-spots and cold-spots were identified for each of the chromosomes. A panel of 27 loci encoding determinants previously identified as immunorelevant or likely to be under selection were positioned on the linkage map. We believe this to be the first genetic linkage map for T. parva. This resource, with the availability of the genome sequence of T. parva, will promote improved understanding of the pathogen by facilitating the use of genetic analysis for identification of loci responsible for variable phenotypic traits exhibited by individual parasite stocks.     

Detection of lumpy skin disease virus in saliva of ticks fed on lumpy skin disease virus-infected cattle

Lumpy skin disease is an economically important disease of cattle that is caused by the lumpy skin disease virus (LSDV), which belongs to the genus Capripoxvirus. It is endemic in Africa and outbreaks have also been reported in the Middle-East. Transmission has mostly been associated with blood-feeding insects but recently, the authors have demonstrated mechanical transmission by Rhipicephalus appendiculatus as well as mechanical/intrastadial and transstadial transmission by Amblyomma hebraeum. Saliva is the medium of transmission of pathogens transmitted by biting arthropods and, simultaneously, it potentiates infection in the vertebrate host. This study aimed to detect LSDV in saliva of A. hebraeum and R. appendiculatus adult ticks fed, as nymphs or as adults, on LSDV-infected animals, thereby also demonstrating transstadial or mechanical/intrastadial passage of the virus in these ticks. Saliva samples were tested for LSDV by real-time PCR and virus isolation. Supernatants obtained from virus isolation were further tested by real-time PCR to confirm that the cytopathic effects observed were due to LSDV. Lumpy skin disease virus was detected, for the first time, in saliva samples of both A. hebraeum and R. appendiculatus ticks. At the same time, mechanical/intrastadial and transstadial passage of the virus was demonstrated and confirmed in R. appendiculatus and A. hebraeum.     

Toxicity of Khaya senegalensis (Meliaceae) Seed Oil Against the Larvae of Boophilus decoloratus: A One-Host Tick in Cattle

The in vitro toxicity of seed oil of Khaya senegalensis (family: Meliaceae) was tested against the larvae of a one host tick, Boophilus decoloratus (family: Ixodidae or hard tick) known as ‘blue tick’ parasitic mainly to cattle commonly found in savannas of tropical equatorial Africa. The 20, 40, 60, 80, and 100 % concentrations of seed oil were found to kill all (100 % mortality) the larvae after 28, 28, 24, 20 and 20 h respectively.     

Allopatric speciation in ticks: genetic and reproductive divergence between geographic strains of Rhipicephalus (Boophilus) microplus

Background  

The cattle tick, Rhipicephalus (Boophilus) microplus, economically impact cattle industry in tropical and subtropical regions of the world. The morphological and genetic differences among R. microplus strains have been documented in the literature, suggesting that biogeographical and ecological separation may have resulted in boophilid ticks from America/Africa and those from Australia being different species. To test the hypothesis of the presence of different boophilid species, herein we performed a series of experiments to characterize the reproductive performance of crosses between R. microplus from Australia, Africa and America and the genetic diversity of strains from Australia, Asia, Africa and America.