Spatial distribution of the Karoo paralysis tickIxodes rubicundus (Acari: Ixodidae) within a false upper Karoo veld type

Spatial distribution of the Karoo paralysis tick (Ixodes rubicundus) has been studied in a paralysis enzootic area of the southwestern Orange Free state. Significantly more females than males were collected from the vegetation. Most ticks (99%) used grass as questing sites at a height of approximately 45 cm (range 10–97 cm), which correlates with the size of host animals. No significant differences between the questing height of male and fermale ticks were evident. The ticks were found on most of the grass species present and, except for the genusEragrostis no specific preference was evident. When questing, the capitulum ofI. rubicundus is usually (90%) directed towards the ground. Seventy-eight percent of the ticks occurred singly on a specific grassblade or flowerhead. In those cases where two ticks occurred together, no sexual aggregation was evident. Most ticks occurred under or close ( =62 cm) to the corwn cover of specific shrub or tree species. These plant species were characterized by a dense crown cover with a mat of decaying leaf litter underneath. The preference of the tick for wild olive trees (Olea europaea fricana) suggests that except for the physical characteristics of this tree species, host factors may also be important. Significantly more ticks were distributed on the cooler, more protected southern sides of vegetation in comparison to the northern sides.     

A comparison of the attraction of nymphs and adults of the ticksAmblyomma hebraeum andA. variegatum to carbon dioxide and the male-produced aggregation-attachment pheromone

The attraction of nymphs of the ticksAmblyomma hebraeum andA. variegatum to CO₂ alone and CO₂ together with the male-produced aggregation-attachment pheromone (AAP) was investigated. Matching experiments on the attraction of the adults of the two species were run for comparison. Nymphs ofA. hebraeum were strongly attracted to sources of CO₂ and CO₂/AAP at distances of 10 and 15 m. At distances of 20 and 25 m significantly more nymphs were attracted to CO₂/AAP than to CO₂ alone. Adults ofA. hebraeum were not attracted to CO₂ alone at any distance; these ticks were attracted to CO₂ and AAP together for distances of up to 25 m. The patterns of attraction of nymphs and adults ofA. variegatum at 10 m were similar to those recorded inA. hebraeum.     

Hyalomma truncatum (Acari; Ixodidae): investigations into the scototaxis of unfed adult ticks

Under controlled test conditions, unfed male and female Hyalomma truncatum ticks exhibited a positive scototaxis to stationary, two-dimensional targets. Upright-positioned rectangles were the most attractive targets. The attractiveness of these targets increased with their size. Significantly more ticks responded scototactically positively to the targets under a luminance contrast ratio of 5:1, as compared with other luminance contrast ratios. Targets with an elevation angle of 13° were occupied more frequently than objects with higher elevation angles. Scototaxis was the same towards a stationary and a sinusoid oscillating target. When an upright-positioned rectangle was combined with a CO₂ gradient, the number of ticks that migrated into the CO₂ gradient and contacted the target did not increase significantly. The interval between exposure and first locomotion of the ticks, however, was significantly shorter under the influence of a CO₂ gradient than in all other experiments without a CO₂ gradient. A temperature gradient simulating a natural host (cattle) did not alter the scototaxis. The results of these investigations suggest that the positive scototaxis exhibited by adult H. truncatum ticks is not likely to be part of their appetence behaviour but rather searching behaviour to find adequate protection from harsh climatic conditions.     

Responses of the ticksAmblyomma hebraeum andA. variegatum to known or potential components of the aggregation-attachment pheromone. III. Aggregation

Ten known or potential components of the aggregation-attachment pheromone (AAP) of the ticksAmblyomma hebraeum andA. variegatum, as well as mixtures of these components, extracts of prefed males and live prefed males, were tested as aggregation stimulants. In field assays, laboratory-reared unfed male and female ticks were released 20 cm downwind of CO₂/pheromone release sites; the numbers of ticks that aggregated at the release sites were recorded after 30 min. InA. variegatum, aggregation was induced by methyl salicylate,o-nitrophenol, 2,6-dichlorophenol, phenylacetaldehyde and some mixtures containing these compounds; a strong aggregation response was induced by an extract of five prefed malesA. variegatum and a weak response was induced by an extract of 50 prefed males ofA. hebraeum. InA. hebraeum, aggregation was induced by phenylacetaldehyde, mixtures of compounds that included phenylacetaldehyde, extracts of 50 prefed males ofA. hebraeum orA. variegatum and 50 live prefed males ofA. hebraeum. InA. variegatum, aggregation was inhibited if compounds that do not occur naturally in the AAP of the species were included in mixtures. InA. hebraeum, phenylacetaldehyde appeared to act as an arrestant for ticks that had been attracted to release sites by other compounds.     

Odor-mediated flight behavior ofAnopheles gambiae gilesSensu Stricto andAn. stephensi liston in response to CO2, acetone, and 1-octen-3-ol (Diptera: Culicidae)

The flight behavior of Anopheles gambiae s.s. Giles and An. stephensi Patton exposed to different odor cues was studied in a wind tunnel. Odors consisted of CO₂, CO₂ + acetone (at two concentrations), and CO₂ + 1-octen-3-ol. Mosquitoes were released singly and their behavior was recorded on video. Parameters studied included flight velocity, percentage of time spent flying, percentage of time spent in plume, and number of turns toward the plume. Large differences in behavior toward the odors tested were observed. An. gambiaedid not respond well to CO ₂,whereas An. stephansiwas positively affected by this compound. In contrast, An. gambiaeresponded significantly to CO ₂ + acetone (at a low concentration), but the behavior of An. stephensiwas completely suppressed by this combination of odor stimuli. CO ₂ + a high concentration of acetone or CO ₂ + 1-octen-3-ol did not cause significant effects in An. gambiaecompared to no odor, while these treatments elicited strong behavioral responses in An. stephensi.The latter species responded particularly well to CO ₂ + 1-octen-3-ol. The results suggest that the observed differences may be inherent to the known differences in host preferences, where An. gambiaeis highly anthropophilic and An. stephensimore zoophilic. This would explain why the latter species responds well to CO ₂ and even better to CO ₂ + 1-octen-3-ol, a compound readily emitted by bovine ruminants.     

Improvement of field sampling methods for adult Karoo paralysis ticks,Ixodes rubicundus (Acari: Ixodidae), through addition of host odour

The responsiveness of Ixodes rubicundus ticks on questing substrates and the success of their attachment to non-living substrates were investigated. The purpose of this study was to relate responsiveness to conditions of temperature and humidity, to compare the efficacy of flagging and dragging methods to estimate the size of populations of adult I. rubicundus in the field, and to determine the possible influence of host odour on the efficacy of these methods. Responsiveness was tested under varying conditions of temperature and humidity under field conditions, and the same ticks were used to determine the duration of attachment to a flannel cloth, either impregnated with host odour from sheep wool or without it. Flagging and dragging methods were compared under laboratory conditions with cloths either treated with host odour or not. Within the range of ambient temperatures recorded during this study (7–25°C) most of the ticks (86%) were responsive. No obvious relationship between temperature, relative humidity and responsiveness of ticks was evident. Ticks remained on average 32.9 seconds (n=64) on cloth treated with host odour compared to 9.1 seconds (n=54) on untreated cloth. Flagging was 1.5–1.7 times as effective as dragging and treatment of the cloths with host odour increased the efficacy 2.4 (dragging) to 2.8 (flagging) times. The reactions of ticks to external stimuli should, therefore, be taken into account to maximize field sampling success.     

Attraction of Amblyomma variegatum (ticks) to the attraction-aggregation-attachment-pheromone with or without carbon dioxide

The responses of adult Amblyomma variegatum ticks released from various distances to different doses of the synthetic attraction-aggregation-attachment pheromone (AAAP) (made up of ortho-nitrophenol, methyl salicylate and nonanoic acid in paraffin oil), dispensed from the center of circular field plots, were studied in the presence or absence of elevated levels of CO₂. Up to 90% of the ticks released were attracted to the pheromone source in the presence of CO₂ within 3h. CO₂ alone was unattractive, similar to previous findings in Zimbabwe, but unlike results from a Caribbean A. variegatum population, which was significantly attracted to this signal. In the absence of CO₂, smaller but significant proportions of the released ticks were attracted to the pheromone, albeit more slowly, suggesting another variation in the responses of this bont tick to inter- and intra-specific signals. Our results are interpreted in the light of a study undertaken elsewhere demonstrating relatively high heterozygosity among tick populations. Possible directions of further research to explore the use of the pheromone in off-host control of the tick are also highlighted.     

Hyalomma truncatum and Rhipicephalus evertsi mimeticus (Ixodoidea,Ixodidae): reactions of adult ticks to vertically incident narrow- and wideband optical radiation with and without the influence of a CO2 gradient

Investigations concerning the reactions of unfed Hyalomma truncatum and Rhipicephalus evertsi mimeticus female and male ticks to vertically incident narrow- and wideband optical radiation at irradiances of 0.115 mWcm^-2 and 0.98 mWcm^-2, respectively, revealed that, independent of sex, adults of the two species are capable of perceiving a wide range of wavelengths. Considering the migration into the entry area of the optical radiation at the test chamber's ceiling as a positive phototaxis, H. truncatum always reacted with higher percentages than R. e. mimeticus. Compared to the controls, R. e. mimeticus ticks occupied the entry area of the optical radiation significantly more frequently only in the wavelength ranges of 415–474 nm, 529–628 nm, 611–707 nm and 190–2,600 nm, but always with low rates. High percentages of H. truncatum ticks, however, consistently reacted with a positive phototaxis in all the offered monochromatic sectors between 292–707 nm and also in the wide spectral range of 190–2,600 nm, particularly in the range of 470–520 nm. When ticks of both species were additionally confronted with a stable CO₂ gradient, continuously increasing from 0.18 vol% at the bottom to 0.90 vol% at a height of 40 cm in the test chamber, they moved less frequently to the entry area of the optical radiation, compared to ticks tested without CO₂, regardless of their exposure to darkness or to narrow- and wideband radiation. The percentages of ticks, however, that moved in a vertical direction, but did not reach the chamber's ceiling were always higher, with the exception of R. e. mimeticus at the wavelength range of 415–474 nm. A total of 30 unfed male and 30 unfed female adult ticks of both tick species were investigated at each combination of a narrow- and wideband spectral range, with and without the influence of a CO₂ gradient. Considering moving ticks only, the interval between exposure and first movement of ticks was shorter under the influence of an additional CO₂ gradient. The average delay in reaction with and without the stimulus of the CO₂ gradient was 156.2 s and 195.6 s, respectively, for H. truncatum and 126.6 and 226.3 s, respectively, for R. e. mimeticus.     

A shared epitope found in the major paralysis inducing tick species of Africa

Cross-reactivity between all the paralysis inducing tick species of veterinary relevance in Africa was demonstrated, by using a monoclonal antibody directed against the paralysis inducing toxin of Rhipicephalus evertsi evertsi. Western blot results, together with amino acid composition studies indicated that this monoclonal antibody recognizes protein bands of similar molecular mass and amino acid composition in R. evertsi evertsi and Ixodes rubicundus. This suggests that the Karoo paralysis toxin of I. rubicundus is possibly also a trimer with a high degree of homology to the spring lamb paralysis toxin of R. evertsi evertsi. The conclusive identity of these protein bands of 1. rubicundus could not be shown. Bio-assay studies performed on 1-day-old chickens suggested that the anti-spring lamb paralysis toxin monoclonal antibody also recognizes the paralysis toxin present in Argas (Persicargas) walkerae, by rendering some degree of protection against the effect of this toxin.     

Vertical migration of adult Ixodes rubicundus,the Karoo paralysis tick (Acari: Ixodidae)

Mortality due to Karoo paralysis amongst livestock is of considerable economic importance in South Africa. Indications are that the extent of vertical migration of adult ticks, to reach questing positions on grasses, is influenced by sudden climatic chages. Such influences as well as selection of questing substrates were investigated under controlled conditions. Vertical migration of adult Ixodes rubicundus on copper rods was determined in a square metal arena within a closed glass container in a room in which temperature and light could be controlled. Significantly more female and male ticks migrated vertically on 1.5 mm diameter rods than on 4.0 mm rods and on 450 mm long rods than on 100 mm rods. More ticks of both sexes migrated vertically at RH>80% than at RH<50% and at 12°C than at 21°C. More female than male and more older than younger ticks migrated vertically in all experiments. The results of this study contribute towards an understanding of the epidemiology of Karoo paralysis.     

Immunization of cattle with a semi-purified fraction of solubilized membrane-bound antigens extracted from the midgut of the tickRhipicephalus appendiculatus

Boran cattle were immunized with a semi-purified fraction (F₂) of solubilized membrane-bound antigens extracted from the midgut of the tickRhipicephalus appendiculatus. A subsequent challenge with all stages ofR. appendiculatus showed protection of the cattle to a significant degree. There was a decrease in engorgement weight of ticks, the number which fed above critical engorgement weight was considerably reduced, and about 38% of the larvae which fed to repletion did not moult. The major cause of death of adult female ticks was the increased grooming by cattle as a result of increased dermal irritation.     

Activation of the ambusher tick Rhipicephalus microplus (Acari: Ixodidae) exposed to different stimuli

The present study aimed to evaluate the behaviour of larvae of Rhipicephalus microplus exposed to different stimuli. A Y-olfactometer was positioned vertically and R. microplus larvae were exposed to environmental air, CO₂ alone, N,N-diethyl-3-methylbenzamide (DEET) alone, and CO₂ combined with the repellents DEET and (E)-2-octenal. Tests were also conducted with the olfactometer positioned horizontally; in this case, however, only CO₂ was tested. In all tests conducted with the Y-olfactometer positioned vertically, CO₂ activated R. microplus larvae even in the presence of DEET and (E)-2-octenal, although activation was lower when these repellents were used. In the absence of CO₂, larval behaviour against DEET was similar to that of the larvae in the control group. In the tests performed with the olfactometer positioned horizontally, the larvae had no significant response to the presence of CO₂. The larvae were not attracted to or repelled by any compound tested in either the vertical or horizontal position of the olfactometer. The lack of horizontal displacement, attraction or repellence may have been a result of the ambush behaviour of this tick species. However, when larvae were exposed to stimuli and the olfactometer was positioned vertically, the interference of attractant and repellent stimuli in larval behaviour was assessed.     

Use of carbon dioxide and natural or synthetic aggregation-attachment pheromone of the bont tick,Amblyomma hebraeum,to attract and trap unfed adults in the field

It was determined previously that unfed adults ofAmblyomma hebraeum become active in response to carbon dioxide but are not attracted by it. We have now shown in field experiments that males and females which are activated by carbon dioxide are attracted to the aggregation-attachment pheromone produced by males after a period of feeding on the host. Unfed adults were attracted too-nitrophenol, a component of the aggregation-attachment pheromone ofAmblyomma variegatum, pre-fed males ofA. hebraeum, and extracts of such males. Live males were more attractive than the extracts which, in turn, were more attractive thano-nitrophenol. The ticks were attracted to sources ofo-nitrophenol from as far away as 11 m in 90 min. Simple traps, baited with carbon dioxide and aggregation-attachment pheromone (from pre-fed males or extracts of pre-fed males), were used to capture unfed adults that had been released in the field. The response of unfed adults to carbon dioxide and the aggregation-attachment pheromone presents an obvious host-location mechanism and a means for the ticks to discriminate between suitable hosts (those with males already attached) and potentially unsuitable hosts (those with no males attached).     

Responses of Amblyomma americanum and Dermacentor variabilis to odorants that attract haematophagous insects

Carbon dioxide (CO₂), 1‐octen‐3‐ol, acetone, ammonium hydroxide, L‐lactic‐acid, dimethyl trisulphide and isobutyric acid were tested as attractants for two tick species, Amblyomma americanum and Dermacentor variabilis (Acari: Ixodidae), in dose–response bioassays using Y‐tube olfactometers. Only CO₂, acetone, 1‐octen‐3‐ol and ammonium hydroxide elicited significant preferences from adult A. americanum, and only CO₂ was attractive to adult D. variabilis. Acetone, 1‐octen‐3‐ol and ammonium hydroxide were separately evaluated at three doses against CO₂ (from dry ice) at a field site supporting a natural population of A. americanum nymphs and adults. Carbon dioxide consistently attracted the highest number of host‐seeking ticks. However, for the first time, acetone, 1‐octen‐3‐ol and ammonium hydroxide were shown to attract high numbers of A. americanum. Further research is needed to determine the utility of these semiochemicals as attractants in tick surveillance and area‐wide management programmes.     

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.     

Responses of the ticksAmblyomma hebraeum andA. variegatum to known or potential components of the aggregation-attachment pheromone. I. Long-range attraction

Nine known or potential components of the aggregation-attachment pheromone of the ticksAmblyomma hebraeum andA. variegatum were tested as long-range attractants. In field assays, laboratory-reared unfed male and female ticks were released 4 m downwind of treated filter-paper targets. Carbon dioxide was used to activate the ticks and the numbers attracted to the targets were recorded. Adults ofA. hebraeum were attracted to six compounds (benzaldehyde, 2,6-dichlorophenol, heptadecane, methyl salicylate, nonanoic acid and o-nitrophenol) and adults ofA. variegatum to four compounds (2,6-dichlorophenol, methyl salicylate, nonanoic acid and o-nitrophenol). The most important attractant forA. hebraeum was o-nitrophenol; methyl salicylate and o-nitrophenol were the most important attractants forA. variegatum. Nonanoic acid and o-nitrophenol attracted significantly more female than maleA. hebraeum. Both species were also attracted by a mixture of the nine compounds.     

Artificial-membrane feeding of the ixodid tick,Rhipicephalus appendiculatus,to repletion

An artificial feeding device was constructed with a Baudruche, membrane bearing olfactory and tactile stimuli, to induce voluntary attachment and feeding of the brown ear tick,Rhipicephalus appendiculatus. Preparation of the membrane and experimental conditions used are described. Unfed adults ofR. appendiculatus, 2–3 weeks after moulting, were confined on the membrane, which was treated with cattle ear-wash and other stimuli. They attached and commenced sucking blood within 12–48 h. Fully engorged ticks started detaching from the membrane on the 6th day. After a 5–6-day incubation period, they started laying eggs which hatched into normal larvae.     

Interspecific variations in attachment sites and density assessment in femaleIxodes rubicundus (Acari: Ixodidae) on domestic and natural hosts

The attachment sites of female Karoo paralysis ticks (Ixodes rubicundus) and equations to assess tick burdens on domestic (sheep and goats) and natural (mountain reedbuck) hosts were determined. No intraspecific differences in attachment sites for Merino sheep and Angora goats could be observed, but interspecific differences were evident.Ixodes rubicundus in general attaches on the ventral aspects of the host. More than 50% of these ticks attached to the fron quarters of the body of the domestic host, and 60% to the hindquarters of the natural host. The length of Merino sheep wool had no affect on either the attachment site or number of attached ticks. In general, more ticks were found attached to the woollen than non-woollen body areas. Equations are provided which can estimate tick burdens on hosts by counting the ticks on two or three selected body areas of the host. To facilitate sampling procedures, approximate sample sizes for the various hosts needed to estimate average tick burdens have been provided. Depending on the level of precision required (with bounds on the error of estimation equal to 1, 2 or 5 ticks), the number of animals in a flock of 200 Angora goats that must be examined varied from 26 to 85.     

Genotype-specific response of a lycaenid herbivore to elevated carbon dioxide and phosphorus availability in calcareous grassland

Effects of elevated CO₂ and P availability on plant growth of the legume Lotus corniculatus and consequences for the butterfly larvae of Polyommatus icarus feeding on L. corniculatus were investigated in screen-aided CO₂ control chambers under natural conditions on a calcareous grassland in the Swiss Jura mountains. Elevated CO₂ conditions and P fertilisation increased the biomass production of L. corniculatus plants and affected the plant chemical composition. CO₂ enrichment increased the C/N ratio and sugar concentration and decreased the N and P concentrations. C- and N-based allelochemicals (cyanoglycosides, total polyphenols and condensed tannins) were only marginally affected by CO₂ enrichment. P fertilisation increased the specific leaf area and concentrations of water, N, sugar and P, while the C/N ratio and the concentration of total polyphenols decreased. Furthermore, P availability marginally enhanced the effect of elevated CO₂ on the total dry mass and sugar concentration while the opposite occurred for the total polyphenol concentration. The changes in food-plant chemistry as a result of P fertilisation positively affected larval mass gain and accelerated the development time of P. icarus. Only a marginal negative effect on larval mass gain was found for CO₂ enrichment. However, we found genotype-specific responses in the development time of P. icarus to elevated CO₂ conditions. Larvae originating from different mothers developed better either under elevated CO₂ or under ambient CO₂ but some did not react to CO₂ elevation. As far as we know this is the first finding of a genotype-specific response of an insect herbivore to elevated CO₂ which suggests genetic shifts in insect life history traits in response to elevated CO₂.     

Laboratory studies on the olfactory behaviour of Anopheles quadriannulatus

The host preference of Anopheles quadriannulatus Theobald (Diptera: Culicidae), the zoophilic member of the malaria mosquito complex Anopheles gambiae Giles, was investigated in a dual‐choice olfactometer. Naïve female mosquitoes were exposed to CO₂, acetone, 1‐octen‐3‐ol, and skin emanations from cows and humans in various combinations. Their behavioural responses were recorded when they had entered one of either upwind traps from where the odours were being released. The mosquitoes did not respond to CO₂ when released at human or cattle equivalent concentrations. Too few mosquitoes responded to acetone to allow for a statistical analysis. The combination of CO₂+ 1‐octen‐3‐ol was repellent. Cow odour alone was slightly attractive, but there was a synergistic attractive effect of cow odour + CO₂. Surprisingly, the mosquitoes were attracted to human odour, and in a choice situation human odour was selected above cow odour + CO₂. Anthropophilic An. gambiae Giles s.s. was repelled by cow odour + CO₂ in contrast to An. quadriannulatus. In a choice situation, both mosquito species selected human odour above cow odour + CO₂. The implications of these results are discussed in the light of recent behavioural data from the field.