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  • Publication
    AccĆØs libre
    A standardised in vivo and in vitro test method for evaluating tick repellents
    (2013)
    Krƶber, Thomas
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    The threat of transmission of Lyme borelliosis and tick-borne encephalitis by ixodid ticks has resulted in an increasing number of tick repellents coming onto the market. To allow proper evaluation of the efficacy of different types of compounds and their formulations, there is a need for standardised methods for testing ticks repellents. Ticks show a marked negative geotactic response following contact with a potential host, i.e., they climb up in order to locate attachment and feeding sites, whereas exposing ticks to repellents induces positive geotaxis, i.e., ticks walk downwards or drop off the treated host or substrate. We describe here complementary tests that employ these geotactic responses to evaluate repellents: one in vitro on a warm glass plate and the other on the lower human leg (shin). The compounds tested were DEET, EBAAP, icaridin, capric acid, lauric acid, geraniol, citriodiol, citronella essential oil and lavender essential oil, all non-proprietary ingredients of widely distributed tick repellent formulations.

    In controls on both the warm glass plate and the human leg, the majority of Ixodes ricinus nymphs walk upwards. By contrast, in both the in vitro and in vivo tests, effective doses of repellents cause ticks to either walk downwards or fall off the substrates, termed here ā€œaffected ticksā€. The ED75 values for affected ticks on the human leg indicate that the test products can be divided into three groups: (1) icaridin, EBAAP, DEET and capric acid with values between 0.013 and 0.020 mg/cm2, (2) citriodiol and lauric acid with values between 0.035 and 0.058 mg/cm2, and (3) geraniol, citronella oil and lavender essential oil with values between 0.131 and 1.58 mg/cm2. The latter three products can be considered as less effective repellents. The tests on the warm glass plate resulted in very similar efficacy rankings for the products tested in vivo, and the ticksā€™ behavioural responses also corresponded closely to those observed on the treated human leg. The ED75 values on the glass plate ranged from half to one sixth needed on the leg. The warm glass plate test thus provides a reliable alternative to human subjects for an initial evaluation of new repellents, and is particularly appropriate for testing products with still to be determined human toxicity and dermatological effects.
  • Publication
    AccĆØs libre
    Antiectoparasitic activity of the gum resin, gum haggar, from the East African plant, Commiphora holtziana
    (2008)
    Birkett, Michael A.
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    Al Abassi, Sate
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    Chamberlain, Keith
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    Hooper, Antony M.
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    Pettersson, Jan
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    Pickett, John A.
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    Slade, Robin
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    Wadhams, Lester J.
    The mechanism of ixodid tick (Acari: Ixodidae) repellency by gum haggar, a resin produced by Commiphora holtziana (Burseraceae), was investigated by evaluating activity against the cattle tick, Boophilus microplus. In an arena bioassay, a hexane extract of the resin of C. holtziana exhibited a repellent effect lasting up to 5 h. The hydrocarbon fraction of the resin extract was shown to account for the repellent activity, and was analysed by coupled gas chromatographyā€“mass spectrometry (GCā€“MS). Major sesquiterpene hydrocarbons were tentatively identified as germacrene-D, Ī“-elemene and Ī²-bourbonene. The identity and stereochemistry of the former compound was confirmed as the (+)-isomer by peak enhancement using enantioselective GC, whereas the latter 2 compounds, which are most likely degradation products of germacrene-type precursors, were identified through isolation by preparative gas chromatography followed by microprobe-NMR spectroscopy. GC comparison of gum haggar with another resin, C. myrrha, which was inactive in the tick bioassay, showed that the latter contained much lower levels of these hydrocarbons. To assess the suitability of the gum haggar resin as a general acarine repellent, further tests were made on a major acarine pest of European and US animal husbandry systems, the red poultry mite, Dermanyssus gallinae (Acari: Dermanyssidae). Gum haggar extract, and the isolated hydrocarbon fraction, showed strong repellent effects in an olfactometer assay, and again gum myrrh showed no effect. These findings provide a scientific basis for the observed anti-tick properties of gum haggar, and demonstrate the potential for its development as a general acarine repellent for use in animal husbandry systems.
  • Publication
    AccĆØs libre
    Behavioural and chemoreceptor cell responses of the tick, Ixodes ricinus, to its own faeces and faecal constituents
    (Springer, 2001)
    Grenacher, Stoyan
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    Krƶber, Thomas
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    ;
    Ticks are ectoparasites of vertebrates and utilize a variety ofinfochemicals for host finding and acceptance as well as for intraspecific aggregation and mating responses. Individual male and female Ixodes ricinus, the vector of Lyme disease in Europe, readily arrest onfilter paper strips contaminated with their own faeces. I. ricinus also responds, but to a lesser degree, tofaeces-contaminated papers enclosed in metal mesh envelopes, i.e. without directly contacting the faeces, suggesting a role for volatiles in the arrestment response. The faecal constituents guanine, xanthine, uric acid and 8-azaguanine (a bacterial breakdown product of guanine) also caused arrestment of individual I.ricinus males and females. However, mixtures of these products induced arrestment of I. ricinus at doses one hundred fold lower than the lowest active dose of any of them tested singly. Saline extracts of faeces activated receptor cells in terminal pore sensilla on the first legtarsi of I. ricinus. One cell in these sensilla responded in a similar dose dependent manner to guanine and 8-azaguanine, whereas a second cell was more sensitive to lower doses of 8-azaguanine. The response threshold approached 100 fM for both cells. These findings suggest that faeces and faecal breakdown products are implicated in aggregation responses of I. ricinus. This may account for the clumped distribution of this ectoparasite on the ground and contribute to the high proportion of mated individuals recorded prior to host colonization.
  • Publication
    AccĆØs libre
    Responses of the tropical bont tick, Amblyomma variegatum (Fabricius), to its aggregation-attachment pheromone presented in an air stream on a servosphere
    (2000)
    McMahon, Conor
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    Male Amblyomma variegatum ticks feeding on a host release a mixture of o -nitrophenol and methyl salicylate which serves to attract conspecifics. The behavioural responses of A. variegatum on a servosphere to these volatiles presented in an air stream are detailed here. In still air, ticks walked on all eight legs, but with long halts. In contrast, the air stream caused continuous walking and induced a reaching response where the forelegs actively sampled the air. Such reaching increased the angular velocity and reduced walking speed, effects that were amplified in the presence of vapours from o -nitrophenol and methyl salicylate in the air flowing over the ticks. Vapour from a 1:1 mixture of o -nitrophenol and methyl salicylate was attractive over a 104-fold concentration range providing an increase in upwind displacement of 20ā€“40%, significantly higher than the natural ratio where o -nitrophenol vapour predominates. Although the responses to o -nitrophenol vapour were variable when presented alone, this chemical was consistently attractive when delivered with steer hair odour ā€“ unattractive on its own. Moreover, the upwind walk to this combination did not cause a change in speed or angular velocity. This supports the hypothesis that the response to the pheromone is enhanced by host odour.
  • Publication
    AccĆØs libre
    Identification of vertebrate volatiles stimulating olfactory receptors on tarsus I of the tick Amblyomma variegatum Fabricius (Ixodidae): I. Receptors within the Hallerā€™s organ capsule
    (1994)
    Steullet, Pascal
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    Gas chromatography-coupled electrophysiological recordings (GC-EL) from olfactory sensilla within the capsule of Haller's organ of the tick Amblyomma variegatum indicate the presence of a number of stimulants in rabbit and bovine odours, and in steer skin wash. Some of these stimulants were fully identified by gas chromatography-mass spectrometry analysis and by matching electrophysiological activity of synthetic analogues as: 1) hexanal, 2-heptenal, nonanal, furfural, benzaldehyde, and 2-hydroxybenzaldehyde (in all extracts); 2) heptanal, 2-, 3-, and 4-methylbenzaldehyde, and Ī³-Valerolactone (only in bovine and rabbit odour). Careful examination of the electrophysiological responses permit characterization of 6 receptor types: 1) a benzaldehyde receptor, 2) a 2-hydroxybenzaldehyde receptor, 3) three types of receptors responding differently to aliphatic aldehydes, and 4) a lactone receptor.
  • Publication
    AccĆØs libre
    Isolation of 2,6-dichlorophenol from the cattle tick Boophilus microplus: Receptor cell responses but no evidence for a behavioural response
    (1994)
    de Bruyne, Marien
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    2,6-Dichlorophenol, a compound known as a sex pheromone for several metastriate tick species, was isolated from different life-stages of the cattle tick Boophilus microplus. Receptor cells in two wall-pore single-walled sensilla on the tarsus I of male ticks responded to this compound in a dose-dependant manner. Using these receptors as specific detectors for compounds in the effluent of a gas chromatograph, we detected 2,6-dichlorophenol in extracts of females, males, engorged nymphs and larvae of this one-host tick, but not in an extract of eggs. No other components of the extracts elicited responses from these olfactory sensilla. However, male B. microplus were not arrested on a glass bead treated with 2,6-dichlorophenol and placed on a membrane in a host-simulating arena, whereas a bead treated with a female extract did evoke a strong arrestment response. In addition, no odour-conditioned anemotaxis, change in angular velocity or speed of males walking on a locomotion compensator was observed in response to this compound in a conditioned air-stream. We could therefore not establish a role for 2,6-dichlorophenol on its own as a semiochemical in males of this species.
  • Publication
    AccĆØs libre
    Identification of vertebrate volatiles stimulating olfactory receptors on tarsus I of the tick Amblyomma variegatum Fabricius (Ixodidae): II. Receptors outside the Hallerā€™s organ capsule
    (1994)
    Steullet, Pascal
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    Bovine odour excites olfactory receptor(s) in a wall-pore olfactory sensillum on the anterior pit of Haller's organ in Amblyomma variegatum. Gas chromatography-coupled electrophysiology recordings from this sensillum reveal the presence of 4 active compounds in bovine odour. The two strongest stimulants were identified as 2-nitrophenol and 4-methyl-2-nitrophenol by gas chromatography-coupled mass spectrometry, and by matching electrophysiological activity of synthetic analogues. Synthetic analogues of known vertebrate-associated volatiles also stimulate other olfactory receptors in sensilla on the surface of tarsus I: a lactone receptor responding to Ī³-valerolactone and 6-caprolactone; different fatty acid receptor types responding best to either pentanoic acid, 2-methylpropanoic acid or to butanoic acid; three receptors responding to NH3; and one receptor responding to 3-pentanone. Gas chromatography-coupled mass spectrometry analysis of vertebrate volatiles revealed presence of a number of these olfactory stimulants in concentrates of rabbit and steer odour, i.e. 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, pentanoic acid, and Ī³-valerolactone.
  • Publication
    AccĆØs libre
    Perception of breath components by the tropical bont tick, Amblyomma variegatum Fabricius (Ixodidae): I. CO2-excited and CO2-inhibited receptors
    (1992)
    Steullet, Pascal
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    Wall-pore olfactory sensilla located in the capsule of Haller's organ on the tarsus of Amblyomma variegatum ticks bear cells responding to vertebrate breath: one of these sensilla contains a CO2-excited receptor and a second sensillum has a CO2- inhibited receptor. Each of these antagonistic CO2-receptors, which display typical phasic-tonic responses, monitors a different CO2-concentration range. The CO2-inhibited receptor is very sensitive to small concentration changes between 0 and ca. 0.2%, but variations of 0.01% around ambient (ca. 0.04%) induce the strongest frequency modulation of this receptor. An increase of just 0.001ā€“0.002% (10ā€“20 ppm) above a zero CO2-level already inhibits this receptor. By contrast, the CO2-excited receptor is not so sensitive to small CO2 shifts around ambient, but best monitors changes in CO2 concentrations above 0.1%. This receptor is characterized by a steep dose-response curve and a fast inactivation even at high CO2-concentrations (>2%). In a wind-tunnel, Amblyomma variegatum is activated from the resting state and attracted by CO2 concentrations of 0.04 to ca. 1%, which corresponds to the sensitivity range of its CO2-receptors. The task of perceiving the whole concentration range to which this tick is attracted would thus appear to be divided between two receptors, one sensitive to small changes around ambient and the other sensitive to the higher concentrations normally encountered when approaching a vertebrate host.