Beauveria bassiana (Ascomycota: Hypocreales) as a management agent for free-living Amblyomma americanum (Acari: Ixodidae) in Ohio.

Unfed adult Amblyomma americanum were exposed to the entomopathogenic fungus Beauveria bassiana and placed in the field. Ticks exposed to the fungus experienced higher mortality than those in the control group (P = 0.001). It is recommended that for inclusion in a tick management program, the fungus be applied in the late evening during the months March through June as this is when ticks are most active and conditions suitable for fungal activity occur.

Neem-tree (Azadirachta indica Juss.) extract as a feed additive against the American dog tick (Dermacentor variabilis) in sheep (Ovis aries).

Acaricides can be conveyed to ticks via the blood of their hosts. As fruit and kernel extracts from the Meliaceae family, and, in particular the tetranortriterpenoid azadirachtin (AZA) inhibits tick egg production and embryogenesis in the Ixodidae ticks, we investigated the effects of Neem Azal, an extract containing 43% AZA, given as a feed additive to lambs artificially infested with engorging adult Dermacentor vairiabilis ticks. After tick attachment, the lambs were allotted to three dietary treatments: AZA0 (control, n=10), AZA0.3 (n=5), and AZA0.6 (n=5), with feed containing 0%, 0.3%, and 0.6% AZA on DM basis, respectively. In half of the AZA0 lambs, ticks were sprayed on day 4 after attachment with an ethanol:water:soap emulsion containing 0.6% AZA (AZA0S). In spite of its very pungent odor, the neem extract was well accepted by all but one lamb. No differences were found between treatment groups in liver enzymes in blood, and there was no indication of toxicity. The plasma AZA concentrations after 7 and 14 days of feeding AZA were (4.81 and 4.35 microg/mL) for the AZA0.6 and (3.32 and 1.88 microg/mL) for the AZA0.3 treatments, respectively (P<0.0001). Treatments were not lethal to ticks, but tick weights at detachment were 0.64, 0.56, 0.48, and 0.37 g for ticks from the AZA0, AZA0.3, AZA0S, and AZA0.6 treatments (P<0.04), respectively, suggesting that blood AZA impaired blood-feeding. The highest mortality rate after detachment was for AZA0.6 (P<0.09). As AZA affects embryo development and ticks at the molting stages, we expect that following treatments of hosts for longer periods, one-host ticks will be more affected than the three-host tick D. variabilis.

Bovine hypodermosis: prevalence and economic significance in southern Punjab, Pakistan.

Prevalence and economic losses of warble fly infestation (WFI) in cattle and buffaloes were recorded in Dera Ghazi Khan and Rajan Pur districts of southern Punjab (Pakistan). A total of 18,000 cattle and 9000 buffaloes were examined in the field and slaughter house. Only one species of Hypoderma, i.e. Hypoderma lineatum was recorded from this area. The prevalence of WFI was higher in slaughter house versus field, cattle versus buffaloes, males versus females, and young versus old animals in both the districts. The highest month-wise prevalence was recorded in December and the lowest in July. Organ-wise prevalence of WFI was highest in oesophagus followed in order by rumen and spleen. The economic losses based on the value of warbled and warble-free hides were estimated as Rs. 22.8 million per annum (currency exchange rate is Rs. 57.5=US 1 dollar).

Ixodid ticks feeding on humans in South Africa: with notes on preferred hosts, geographic distribution, seasonal occurrence and transmission of pathogens.

This paper records the identities of 558 ixodid ticks feeding on 194 humans in South Africa. These ticks belonged to 20 species in six genera and those most frequently encountered were AmblYomma hebraeum, Haemaphysalis leachi, Hyalomma marginatum rufipes, Hyalomma truncatum, Rhipicephalus appendiculatus, Rhipicephalus gertrudae and Rhipicephalus simus. With the exception of the larvae of R. appendiculatus, the incidents of these ticks feeding on humans correlated well with their seasonal occurrences on preferred hosts. Ticks were also collected at monthly intervals, for 14 consecutive months, from the clothing of a game-guard providing protection for field-workers engaged in the collection, by means of flannel strips, of free-living ticks from the vegetation of four localities in the southern region of the Kruger National Park. In addition, with the exception of 3 months when the particular worker was absent, ticks that fed on one of the field-workers were collected over the same 14-month period. A total of 54,429 free-living ticks belonging to 14 species and six genera were collected from the vegetation at the four localities during this time and 3751 ticks belonging to 11 species and six genera from the clothing of the game-guard. The larvae of A. hebraeum and Boophilus decoloratus were the most numerous of the immature ticks, and H. leachi and R. simus of the adults on both the vegetation and the guard's clothing. Ticks fed on the field-worker on six occasions and 14 were collected, all of which were A. hebraeum larvae.

Ixodes rubicundus nymphs are short-day diapause-induced ticks with thermolabile sensitivity and desiccation resistance.

The Karoo Paralysis tick, Ixodes rubicundus Neumann (Acari: Ixodidae), is a semi-voltine ixodid that survives stressful environmental conditions using morphogenetic diapause (eggs and engorged nymphs) and desiccation resistance. Both photoperiod and temperature influence diapause induction in the engorged nymph. Ixodes rubicundus nymphs are typical long-day photoperiodic organisms. The critical photoperiod is approximately 13.5 h light, 10.5 h dark, and they display a thermolabile response. The period between detachment and apolysis in engorged nymphs is modified by photoperiod; however, apolysis to ecdysis is not affected by photoperiod. Thus, initiation of development, but not the actual process is controlled by photoperiod. Most engorged nymphs delayed metamorphosis when exposed to short-day regimen (LD 12 : 12 h) after feeding. Nymphs exposed to pre- and post-feeding long-day regimen (LD 14 : 10 h) developed. Times for 50% of nymphs to apolyse when exposed to photoperiods of LD 14 : 10 h, 13.5 : 10.5 h, 13 : 11 h and 12 : 12 h were 28, 36, 40 and 58 days, respectively. Times for 50% of engorged nymphs to ecdyse ranged from 38 to 40 days after apolysis. Nymphs were sensitive to photoperiodic exposures before, during and after feeding. Nymphs exposed to long day (LD 14 :10 h) before and during feeding, moulted at 20 degrees C; however, most exposed to 10 degrees C followed by 20 degrees C (post-feeding) went into diapause. Both short- (10 : 14 h) and long- (14 : 10 h) day exposed engorged nymphs survived 45 days at 0% r.h. (n = 73), but diapause-destined ticks kept at 13 degrees C lost the least mass (29.5+/-9.5%, SD), while nondiapause ticks at 23 degrees C lost the most (48.7+/-8.2%, SD). Termination of diapause and transition to development probably coincides with a definite increase of water vapour uptake by engorged nymphs. Comparatively, I. rubicundus engorged nymphs are more desiccation tolerant than a North-American counterpart, Amblyomma cajennense (Fabricius) (Acari: Ixodidae), which is also semi-arid- to xeric-adapted. Diapause conveys important survival attributes that enable engorged I. rubicundus nymphs to inhabit a semi-arid environment with great temperature extremes, and to synchronize their activity periods with seasons and host utilization patterns.

Palp-splaying behavior and a specific mouthpart site associated with active water vapor uptake in Amblyomma americanum (Acari: Ixodidae).

Ticks actively absorb water vapor from unsaturated air via their mouthparts. A wax technique was used to cover select areas of the mouthparts of the lone star tick, Amblyomma americanum (L.), to examine whether a specific site is involved in uptake. Covering dorsal, ventral, and distal regions failed to inhibit uptake, whereas wax applied to the proximal region, specifically the hypostome-cheliceral junction, blocked water absorption. A novel occurrence of a palp-splaying behavior was observed during hydrating conditions (93% RH), while ticks were motionless, compared with those held at desiccating conditions (63% RH), which were more active and the palps were not splayed.

Antibody to a cDNA-derived calreticulin protein from Amblyomma americanum as a biomarker of tick exposure in humans.

The antibody responses of human and animal hosts were studied to determine the utility of antibody against recombinant tick calreticulin (rTC), a cDNA-derived protein isolated from salivary glands of Amblyomma americanum L., as a biologic marker of tick exposure. Rabbits fed upon by either A. americanum or Dermacentor variabilis Say developed significant anti-rTC antibody responses, as measured by both ELISA and immunoblot assay. In contrast, gerbils exposed to Aedes aegypti did not develop anti-rTC antibodies, as measured by ELISA or immunoblot assay. The utility of the assay was next evaluated in humans at high risk for tick exposure. During April through September 1990, 192 military personnel who originated from either Fort Chaffee, Arkansas or Fort Wainwright, Alaska were studied during maneuvers in tick infested areas at Fort Chaffee. Study subjects completed a questionnaire and had pre- and post-maneuvers serum specimens analyzed for antibodies to rTC. In adjusted analysis (controlling for age, fort of origin, attached tick during maneuvers, and bed netting use), the use of bed netting and home station were associated with post-maneuvers anti-rTC antibody seropositivity by ELISA. Subjects from Fort Wainwright were more likely to be seropositive for anti-rTC antibody (adjusted odds ratio = 5.3, 95% confidence interval [CI] = 1.1-25.6). Personnel who did not report the use of bed netting were more likely to be anti-rTC seropositive (adjusted odds ratio = 6.8, 95% CI = 1.4-32.4). Immunoblot assays showed that humans had specific anti-rTC antibody responses. The animal experiments demonstrate that hosts exposed to naturally feeding ticks develop anti-rTC antibodies. The data also indicate that hosts exposed to Ae. aegypti saliva may not develop antibodies against rTC. Observations in tick-exposed humans support the hypothesis that anti-rTC antibody seropositivity is a biologic marker of tick exposure.

Evaluation of three commercial tick removal tools.

OBJECTIVE: To evaluate three commercially available tick removal tools against medium-tipped nontissue tweezers. METHODS: We evaluated three commercially available tick removal tools against medium-tipped tweezers. Three inexperienced users randomly removed attached American dog ticks (Dermacentor variabilis Say) and lone star ticks (Amblyomma americanum L.) from laboratory rabbits in a university animal facility using all tools during one removal session. RESULTS: Tick damage occurring from removal and quantity of attachment cement were compared. No tool removed nymphs without damage and all tools removed adults of both species successfully. American dog ticks proved easier to remove than lone star ticks, whose mouthparts often remained in the skin. CONCLUSIONS: Nymphal ticks were consistently removed more successfully with commercial tools when compared with tweezers but with more difficulty than adults were removed. The commercial tick removal tools tested are functional for removal of nymphs and adults and should be considered as viable alternatives to medium-tipped tweezers.

Tick saliva: recent advances and implications for vector competence.

Secretions of the tick salivary glands are essential to the successful completion of the prolonged feeding of these ectoparasites as well as the conduit by which most tick-borne pathogens are transmitted to the host. In ixodid ticks the salivary glands are the organs of osmoregulation, and excess water from the bloodmeal is returned via saliva into the host. Host blood must continue to flow into the feeding lesion as well as remain fluid in the tick mouthparts and gut. The host's haemostatic mechanisms are thwarted by various anti-platelet aggregatory, anticoagulatory and anti-vasoconstrictory factors in tick saliva. Saliva components suppress the immune and inflammatory response of the host permitting the ticks to remain on the host for an extended period of time and, adventitiously, enhancing the transmission and establishment of tick-borne pathogens. Over the years much work has been done on the numerous enzyme and pharmacological activities found in the tick saliva. The present article reviews the most recent work on salivary gland secretions with special emphasis on how they favour pathogen transmission.

Impact of feeding, molting and relative humidity on cuticular wax deposition and water loss in the lone star tick, Amblyomma americanum.

To determine whether there is a correlation between the amount of lipids on the surface of ticks and their permeability to water, we quantified cuticular surface wax and measured water-loss rates in the ixodid tick Amblyomma americanum from nonfed nymph to egg-laying female. Ticks deposited no extra cuticular lipids during feeding, permitting maximum transpiratory water loss that presumably helps to concentrate the bloodmeal; and ticks deposited additional cuticular wax after apolysis that reduced integumental water loss, which likely prepares ticks for off-host existence. A remarkable three-fold boost in surface wax deposition and extreme water retention were noted after host drop-off following feeding. This wax is likely host-derived. Fed nymphs could discriminate between low and high relative humidity, enabling pharate adults to conserve lipid that would otherwise be lost with the exuvia and feces. This conservation strategy likely adds to the lipid pool needed by the tick to survive in a dry environment and complements the tick's behavioral abilities for seeking out optimum conditions for water conservation and host location.

Cold-hardiness of a laboratory colony of lone star ticks (Acari: Ixodidae).

The cold-hardiness of a lone star tick, Amblyomma americanum (L.), laboratory colony was characterized. Fed and unfed larvae, fed and unfed nymphs, and unfed adults did not survive exposure to -17 degrees C for 7 d. After an 8-d exposure to -10 degrees C, adults tolerated cold better than immatures and unfed specimens fared better than fed ticks. Exposing unfed 6-wk-old (postmolt) adult males and females to -15 degrees C for increasing intervals up to 2 h suggests that males were more tolerant to cold than were females. Half of all adults were alive 3 d after the 2-h low-temperature treatment. Males may have survived because of a significantly higher hemolymph osmotic pressure, although the solute concentration increased for both sexes after a 2-h exposure to 0 degree C. Acclimation to 5 degrees C for 7 d had no influence on supercooling points for unfed males and females, engorged nymphs and larvae, and eggs. None of the life stages survived supercooling, which strongly suggests that this species is freeze intolerant. Intolerance of immature stages to chilling may be a limiting factor in the northern distribution of lone star ticks in North America.

Survival and water-balance characteristics of unfed adult Amblyomma cajennense (Acari: Ixodidae).

Off-host survival, water balance, and cold tolerance of unfed adult, Cayenne ticks, Amblyomma cajennense (F.), were examined to evaluate species characteristics important to zoogeography and off-host ecology. Survivorship decreased when males and females were subjected to progressively drier constant environmental conditions. Average maximum survival was 641.2 and 682.5 d at 85% RH and 23 degrees C (2.98 mm Hg) for males and females, respectively. Mean survival in both sexes was progressively less variable in drier conditions. Slopes of log-linear models of survival days based on saturation deficit (mm Hg) were significantly different between males and females at 50%, but not at 25 or 0%. Whole-body water loss rates for 4-wk-old adults were measured at 0% RH and 23 degrees C until ticks became nonambulatory. The mean whole-body water loss rate of females, 0.06128% h-1, was 11.3% less than for males, 0.06914% h-1. Although nonambulatory ticks appeared dead, >1/2 of the individuals from each sex regained ambulatory status after they were removed from 0% RH and exposed to 96% RH for 24 h. Among these, male ticks averaged 0.44 more recuperative (ambulatory) cycles than females, although, the duration encompassing all recuperative cycles was generally longer for females and on average, females gained 8.16% more weight than males upon each rehydration. Estimates of the mean critical equilibrium activity for males and females were 0.74 av and 0.79 av, respectively. A. cajennense adults were found to be less tolerant to -12.5 degrees C than adult lone star ticks, Amblyomma americanum (L.), whose distribution encompasses more temperate regions. Although A. cajennense exhibit little host preference and are capable of extended off-host survival, the establishment of populations beyond this species zoogeographic distribution may be constrained by an intolerance to cold.

Fluid secretion by isolated tick salivary glands dependent on an intact cytoskeleton.

Isolated salivary glands from female Dermacentor variabilis (Say) were pre-treated with either cytochalasin D or nocodazol, followed by stimulation with dopamine. Glands pre-treated with 1 microM-cytochalasin D did not secrete fluid; pre-treatment with 1 nM-cytochalasin D did not significantly reduce fluid secretion. Glands pre-treated with 1 microM-nocodazol had a significant reduction in fluid secretion (P = 0.003); 1 nM-nocodazol did not significantly affect fluid secretion. Ligation of the main salivary duct and pre-treatment with 1 microM-cytochalasin D significantly increased gland weight compared to the dopamine stimulated controls (P = 0.0412). Cytochalasin D-treated type III acini had a significantly larger mean diameter compared to the dopamine control (P = 0.0047). Glands treated with 1 mM-verapamil plus 10 microM-dopamine exhibited a significant decrease in fluid secretion (P = 0.017), and when ligated, had a significantly decreased weight compared to the controls (P = 0.0028).

Tick (Acari: Ixodidae) attachment cement and salivary gland cells contain similar immunoreactive polypeptides.

A specific antiserum (12C) raised to a 90-kDa immunogenic component of salivary glands of the tick Rhipicephalus appendiculatus recognized similar 90-kDa polypeptides from salivary glands of the American dog tick, Dermacentor variabilis, and the lone star tick, Amblyomma americanum, as well as 70-kDa polypeptides in the cement of D. variabilis, A. americanum, and R. sanguineus (brown dog tick). The reduction in size of the polypeptide for these ticks suggests that it is modified in some way during or after secretion. Immunostaining of salivary glands of unfed- and partially-fed female D. variabilis localized an immunoreactive protein in the d- and e-cells of the type III acini. The quantity of label in granules of glands from unfed ticks was visibly greater than in the granules of glands from partially fed ticks, suggesting that this component is secreted within the first 2 d of feeding. Collectively, these data support the conclusion that a 90-kDa polypeptide of saliva is conserved among ixodid tick genera and is a component of the attachment cement.

Mediation of host selection by cuticular hydrocarbons in the honeybee tracheal MiteAcarapis woodi (Rennie).

Using a simple two-choice bioassay and video analysis of individual locomotory tracks, it was determined that a preference for young-bee hosts over old-bee hosts in female honeybee tracheal mites,Acarapis woodi (Rennie), is chemically mediated. When presented with a choice of cuticular extracts from 5-day-old and <1-day-old adult bees, mites showed a significant preference for the young-bee extract in three of four bee colony sources. This discrimination was due apparently to a greater positive response elicited by the young-bee extract rather than a deterrent effect of old-bee extract, as the extract of old bees evoked a significantly higher response than a hexane control. A bioassay of silica-gel fractions of young-bee extract indicated that cuticular hydrocarbons alone were responsible for tracheal mite response. Further fractionation of a hydrocarbon fraction by argentation chromatography demonstrated that both saturated and unsaturated hydrocarbons were involved in the mite response, but when presented in opposition, mites showed a stronger response to the saturated than to the unsaturated components. Mites placed in zones treated with cuticular extract of young bees exhibited higher angular velocities than those placed on hexane, causing them to remain in the extract-treated zones for extended periods. These results point to a possible control strategy whereby migration of tracheal mites to young-bee hosts could be disrupted by artificially altering the chemical profile of the honeybee cuticle.

Influence of feeding and starvation on the persistence and transmission of Quaranfil virus by Argas (Persicargas) arboreus (Acari: Argasidae).

Viral antigen was detected in the cytoplasm and in associated membranes of salivary gland acinus cells by indirect immunofluorescence and immunoperoxidase staining. Viral ribonucleoproteins (indicated histochemically by presence of pyroninophilic granules) which had accumulated in the cytoplasm of salivary gland type B (granular) acini of unfed Argas (Persicargas) arboreus Kaiser, Hoogstraal & Kohls were no longer visible 24 h after feeding. Virus in tick salivary glands increased from 300 to 500 plaque-forming units during the brief feeding interval (approximately 1 h), but virus was not detectable by 72 h. Overall salivary gland, ovarian, and synganglion tissue levels of Quaranfil virus decreased in the 96 h after feeding, except for synganglion samples in which virus titers increased during 24 h after feeding. Starvation for 105 d resulted in a sevenfold increase in salivary gland viral content compared with those starved 45 d, whereas synganglion tissue titers for Quaranfil virus became undetectable, and ovarian tissue values were similar to those starved for 45 d. Feeding had a greater effect on viral persistence in tissues for ticks starved 60 additional d (comparing 45 with 105 d) in that no Quaranfil virus was detected in any tissue after 48 h (compared with 72 h). Feeding infected ticks (with short extrinsic incubation) on chicks resulted in a peak of host mortality on days 7 and 8, whereas long extrinsic incubation resulted in sporadic mortality over 20 d of monitoring.

Amblyomma americanum: identification of tick salivary gland antigens from unfed and early feeding females with comparisons to Ixodes dammini and Dermacentor variabilis.

Salivary gland antigens involved in host resistance to tick feeding by Amblyomma americanum (lone star tick) have been identified. Gland extracts from unfed and partially fed 12-, 48-, 72-, 96-, and 120-hr females and their corresponding midgut tissues were analyzed by immunoblotting with sera from naturally immune and hyperimmune sheep and rabbits. Polypeptides at 90, 75, 58, 45, 33, and 23 kDa from the salivary glands of A. americanum females were consistently observed with antibodies from both sheep and rabbits. No antigens unique to tick midgut tissue were detected with immune sera. Female Dermacentor variabilis and Ixodes dammini shared 90- and 45-kDa salivary gland antigens with A. americanum, and these may represent conserved polypeptides. We speculate that some of the salivary gland antigens represent components of tick cement, while others are playing some other yet undetermined role in tick feeding.

Characterization of ixodid tick salivary-gland gene products, using recombinant DNA technology.

Ticks secrete an array of lesion-maintenance factors into the host via the salivary glands while feeding, some of which elicit an immune response by the host that adversely affects the ability of the tick to feed and reproduce. Our approach to characterizing these factors has been to make expression libraries from mRNA of salivary glands (from unfed and 3-day-feeding Amblyomma americanum females) which will serve as sources of the genes (clones) that code for them. Thus far, we have detected 10 positive clones in primary screens using polyspecific antiserum from rabbits hyperimmunized to 3-day-feeding tick salivary glands. We also report making a cDNA library from whole unfed females, and a genomic library from whole unfed ticks, which will serve as additional sources of genetic information for characterizing salivary-gland secretory products. Immunoblots of salivary glands from A. americanum females feeding for various intervals (unfed, and 12, 48, 72, and 96 h) revealed the presence of several prominent polypeptides (90 & 45 kDa) when probed with the same rabbit antiserum that was used to screen the expression library. Ixodes dammini had several immunogens in common with A. americanum at 96 h (90, 45, 43 and 23 kDa). We plan to use monospecific antiserum raised to antigens detected in our immunoblots (e.g. 90 kDa) to further screen the expression libraries, in addition to using the polyspecific antiserum already in hand. We discuss the future use of the salivary-gland genes for characterizing secretory products which facilitate attachment to the host (cement) and maintain the lesion during the lengthy feeding interval.