Hormonal control of salivary gland degeneration in the ixodid tick Amblyomma hebraeum

Under normal circumstances, salivary glands of female ixodid ticks begin degenerating within hours of completing the blood meal. We have monitored cytological, functional and biochemical changes in the tissue which are diagnostic of the degenerative process. Although ultimately degeneration also befalls salivary glands of partially fed ticks removed prematurely from the host, the process is considerably delayed. When we transplanted salivary glands from partially fed ticks into the haemocoels of replete specimens, autolysis was induced in the donor tissue, whereas such was not the case when similar glands were transplanted to the haemocoels of other partially fed ticks. We thus suggest that a humoral factor is involved in postprandial resorption of the salivary glands. Succinate dehydrogenase activity decreases, and acid phosphatase activity increases in the salivary glands as a function of time post-engorgement. However, these enzyme assays are not sensitive enough to detect the earliest stages of autolysis.     

“Male factors” in ticks: their role in feeding and egg development

Summary

Female ticks of the family Ixodidae utilize their salivary glands as the major organs for fluid balance, secreting back into the host a dilute saliva. Feeding is composed of three phases: a preparatory phase (1–2 days) during which the tick establishes the feeding lesion, a slow phase (~7 days) during which body weight increases 10-fold, and a rapid phase (~1 day) in which body weight increases a further 10-fold. Following engorgement, the salivary glands are resorbed by an autolytic process triggered by an ecdysteroid hormone. If a female is removed from the host prior to repletion, her subsequent behaviour depends mostly on two factors: the degree of engorgement achieved and whether or not she has mated. If removed during the preparatory or slow phase of engorgement, the salivary glands are not resorbed, the tick will lay virtually no eggs and she will reattach to a host if given the opportunity, all of this irrespective of whether she is virgin or mated. If removed during the rapid phase of engorgement, however, mated females will not reattach to a host even if given the opportunity. Instead, they will resorb the salivary glands within 4 days post-removal and lay a batch of eggs. Virgin females removed after exceeding 10-fold the unfed weight likewise refuse to resume feeding if given the opportunity, but salivary gland reabsorption is delayed (to 8 days post-removal); if any eggs are laid, they are infertile. A number of chemical “factors” entering the female during copulation influence her feeding behaviour and egg development. Here we discuss the complexities of these interactions and suggest how they might be adaptive to ticks in nature.     

Francisella tularensis prevalence and load in Dermacentor reticulatus ticks in an endemic area in Central Europe

A total of 7778 host‐seeking adult Dermacentor reticulatus (Ixodida: Ixodidae) ticks were examined for the prevalence of Francisella tularensis holarctica (Thiotrichales: Francisellaceae) in a natural focus of tularaemia in the floodplain forest–meadow ecosystem along the lower reaches of the Dyje (Thaya) river in South Moravia (Czech Republic) between 1995 and 2013. Ticks were pooled (10 specimens per pool) and their homogenates inoculated subcutaneously in 4‐week‐old specific pathogen‐free mice. Dead mice were sectioned, their spleens cultivated on thioglycollate–glucose–blood agar and impression smears from the spleen, liver and heart blood were Giemsa‐stained. Sixty‐four pools were positive for F. tularensis: the overall minimum infection rate (MIR) was 0.82%. Overall MIRs for the 4714 female and 3064 male D. reticulatus examined were 0.89 and 0.72%, respectively; MIRs fluctuated across years between 0.0 and 2.43%. The estimated bacterial load in infected ticks varied from 0.84 to 5.34 log₁₀ infectious F. tularensis cells per tick (i.e. from about seven to 220 000 cells). Ticks with low loads were more prevalent; more than 1000 infectious cells were detected in 24 ticks (0.3% of all ticks and 37.5% of infected ticks). Monitoring of D. reticulatus for the presence and cell numbers of F. tularensis may be a valuable tool in the surveillance of tularaemia.     

Monthly dynamics of ticks (Acari: Ixodida) infesting N'Dama cattle in the Republic of Guinea

The occurrence of ixodid ticks on N'Dama cattle was studied in the Republic of Guinea between June 1994 and May 1995. Monthly tick collections were performed on 80 animals from 14 villages located in Dabola, Kouroussa and Dinguiraye prefectures. A total of 19,804 ticks was collected and classified using standard taxonomic keys. The following tick species were identified: Amblyomma variegatum, Hyalomma marginatum rufipes, Hyalomma trunctum, Hyalomma nitidum, Rhipicephalus lunulatus, Rhipicephalus muhsamae, Rhipicephalus senegalensis, Rhipicephalus sulcatus, Rhipicephalus turanicus, Boophilus annulatus, Boophilus geigyi. Boophilus spp. were the most numerous adult ticks (57.1%), Am. variegatum adults constituted 27.4%, while 12.4% were Rhipicephalus spp. and 2.5% Hyalomma spp. Rhipicephalus turanicus and Hyalomma nitidum were recorded for the first time in the country. Am. variegatum and Boophilus spp. were present throughout the year, whereas Am. variegatum adults showed a peak during the rainy season between April and September. Immature stages collected belonged exclusively to the genera Amblyomma and Boophilus. Am. variegatum larvae and nymphs showed a peak during the dry season (October-March); no significant variation between seasons was observed for Boophilus immatures. A significantly higher infestation of cattle by Rhipicephalus spp. was found in Dabola and Kouroussa prefectures, located in the southern part of the study area, with similar climatic, vegetation and rainfall characteristics. Possible options for the control of ticks in the study area are briefly discussed.     

Morphofunctional changes of salivary glands of female ixodid ticks of subfamilies Ixodinae and Amblyomminae (Acari: Ixodidae) during feeding and their significance

Using light and electron microscopy, a dynamics of functioning of salivary glands of female ticks of the genus Ixodes was studied. Comparative analysis of morphofunctional changes and role of the salivary gland secretions of females of subfamilies Ixodinae and Amblyomminae during feeding was performed on the basis of literature data and our own studies.     

Molecular individuality: polymorphism of salivary gland proteins in three species of ixodid tick

Pooled tissue samples are frequently used in biochemical studies involving parasites in order to ensure that there is sufficient material for experimentation. A pooled sample is considered to represent the overall phenotypic characteristics of the investigated population. However, this will not be the case if there is a significant degree of molecular polymorphism among individuals in the sampled population. Here we demonstrate marked differences in the protein profile of salivary glands among individuals from three species of ixodid tick (Rhipicephalus appendiculatus, Amblyomma variegatum, Ixodes ricinus), and show that pooling the tissue of several individuals masks substantial qualitative differences among the individuals. Our observations indicate that much greater caution is needed in general when using pooled samples if the molecular diversity within the population is not clearly defined.     

Morphophysiological analysis of the salivary glands of Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) exposed to ozonated water: A control strategy

The tick Rhipicephalus sanguineus sensu lato has great medical and veterinary importance, mainly because the ability to transmit many diseases, causing harm to pets but also risks to public health. The blood spoliation and transmission of pathogens occur because of the immunosuppressive action of these ticks' saliva, a potent mixture of bioactive substances that is secreted by the salivary glands, one of the organs responsible for their biological success, and hence the target of studies for their control. Ozone has promise for use as an alternative acaricide, due to its proven efficiency in controlling agricultural and food pests, besides posing no risk of environmental contamination or to animal and human health. Therefore, this study evaluated the acaricidal potential of exposure of females of R. sanguineus s.l. to ozonated water at many concentrations and analysed the morphophysiological alterations of the salivary glands, employing histological and light microscopic techniques. The results demonstrated that the ozonated water at the concentrations investigated caused severe alterations in the salivary glands, bringing a new perspective for control of R. sanguineus s.l., through an ecologically correct method due to the absence of harm to non‐target organisms and the environment.     

Salivary gland degeneration and vitellogenesis in the ixodid tick Amblyomma hebraeum: Surpassing a critical weight is the prerequisite and detachment from the host is the trigger

The normal engorged body weight of female ixodid ticks (Acari: Ixodidae) is about 100× the unfed weight. Virgin female Amblyomma hebraeum normally do not feed beyond 10× the unfed weight. However, about 10–20% of a population of virgins will feed to perhaps 20× the unfed weight, but not much beyond that. In A. hebraeum, when females surpass about 10× the unfed weight, the following changes in physiology occur if they are removed from the host: (a) they will not reattach if given the opportunity, (b) their salivary glands (SGs) will undergo autolysis within 4 days if they are mated or 8 days if they are virgin, and (c) egg maturation and oviposition will occur in due course. Mated or virgin female ticks removed from the host below about 10× the unfed weight do not experience the latter changes (Kaufman, W.R., Lomas, L., 1996. ‘Male Factors’ in ticks: their role in feeding and egg development. Invertebrate Reproduction and Development 30, 191–198). In 1984 we named this transitional weight, the ‘critical weight’ (CW). Its absolute value is probably a species-specific characteristic (Kaufman, W.R., 2007. Gluttony and sex in female ixodid ticks: how do they compare to other blood-sucking arthropods? Journal of Insect Physiology 53, 264–273). Although mated females tend to engorge within a day of surpassing the CW, virgin females surpassing the CW can remain attached to the host for at least several weeks more. It is not known whether the physiological changes in the SGs and ovaries listed above occur in those large virgins that remain attached, although we suppose that this would be maladaptive. Instead, we hypothesize in this study that surpassing the CW is only a prerequisite for inducing these changes, and that detachment is the actual trigger. We support our hypothesis by demonstrating that large virgins, remaining attached to a host for 8 days, did not undergo SG degeneration nor complete egg maturation during the attachment period. Those changes occurred only within 8 days following detachment. So some type of sensory information associated with attachment to the host, and still undefined, inhibits expression of the physiological changes hitherto associated merely with surpassing the CW.     

Dynamics of Borrelia burgdorferi infection in nymphal Ixodes ricinus ticks during feeding

We report the sequential developmental events of Borrelia burgdorferi in histological sections of Ixodes ricinus nymphs before, during and after feeding. During the blood meal a decrease of approximately 50% in the number of infected ticks was recorded (eight out of 76, 11%) in comparison with the infection rate of unfed ticks (12 out of 56, 21%). Spirochetes were detected in tick salivary glands only after 2 days of attachment. From day 3 until drop-off, the number of infected ticks increased to 31% (15 out of 49). A quadratic logistic regression analysis showed that the variation in the number of infected ticks was significant, but only during the blood meal. The drop in the percentage of infected ticks during the first hours following attachment to the host is explained by our observation of spirochetes in the faeces of the ticks. The increase in the infection rate of replete ticks may be due to an uptake of spirochetes from the host skin at the feeding site.     

Ixodid tick salivary gland extracts inhibit production of lipopolysaccharide-induced mRNA of several different human cytokines

Extracts prepared from the salivary glands (SGE) of partially fed adult female Rhipicephalus appendiculatus ticks reduced the expression by human peripheral blood leukocytes (PBLs) of lipopolysaccharide (LPS)-stimulated cytokine mRNA. Treatment with SGE had no obvious effect on cytokine mRNA production when compared with untreated PBLs. LPS treatment induced or increased mRNA production for IFN, IFN, TNF-, IL-1, IL-1, IL-5, IL-6, IL-7 and IL-8. All the LPS-stimulated cytokine mRNAs were reduced when treated with a mixture of LPS and SGE. The results indicate the potential of ticks in modulating the cytokine network of their vertebrate hosts, possibly to facilitate blood feeding.     

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

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

Potential tick vectors for Theileria equi in Israel

Theileria equi Mehlhorn and Schein, 1998 (Piroplasmida: Babesiidae) is an important tick-borne pathogen of horses that is highly endemic in many parts of the world, including Israel. The present study evaluated the potential roles of five hard tick species [Hyalomma excavatum Koch, 1844; Hyalomma marginatum Koch, 1844; Rhipicephalus turanicus Pomerantsev 1936; Rhipicephalus annulatus Say, 1821; Haemaphysalis parva (Neumann, 1897) (all: Ixodida: Ixodidae)], previously found to infest horses in Israel, in acting as vectors for piroplasmosis. For this, DNA was extracted from whole ticks and, when possible, from the salivary glands in each species (n = 10–59). Polymerase chain reaction amplification and sequencing of the 18S rRNA gene were used to detect T. equi in 48 of the 127 ticks (37.8%) and in 21 of the 90 extracted salivary glands (23.3%) in all five species. All but two sequences were classified as T. equi genotype A; the remaining two were classified as genotype D. The findings of this study point to Ha. parva and R. annulatus as potential novel vectors of T. equi, and suggest that parasite genotype selection occurs within the tick vector.     

<Emphasis Type="Italic">In vitro</Emphasis> splenocyte proliferation responses of BALB/c mice to salivary gland extracts of three ixodid tick species (Acari: Ixodidae)

In vitro proliferation and cytokine production were investigated in BALB/c mice splenic cell cultures that were stimulated with concanavalin A (ConA) and lipopolysaccharide (LPS) and simultaneously exposed to salivary gland extracts (SGE) of unfed and partially fed adult ixodid ticks (Ixodes ricinus, Rhipicephalus appendiculatus, Amblyomma variegatum). Generally, tick SGE enhanced proliferation of unstimulated splenocytes and SGE of unfed ticks suppressed mitogen induced proliferation. Partially fed R. appendiculatus and A. variegatum suppressed ConA responses, while partially fed I. ricinus stimulated both ConA and LPS induced proliferation. A. variegatum and R. appendiculatus females slightly enhanced LPS responses 2 days after attachment but suppressed them at the end of the slow feeding phase. In 72 h ConA induced cell cultures, interferon gamma (IFN-γ) production was suppressed by SGE of all ticks, interleukin (IL)-10 production was enhanced by unfed I. ricinus and partially fed A. variegatum males and IL-5 production was enhanced by feeding R. appendiculatus females and A. variegatum males. The study revealed variability in the responsiveness of murine splenocytes to SGE of different ixodid tick species, whereby patterns of host immunomodulation within one tick species differed between sexes and changed during feeding.     

Relative importance of lizards and mammals as hosts for ixodid ticks in northern California

Lizards and mammals were trapped and examined for ticks from August 1992 to June 1993 in two habitat types, chaparral and woodland-grass, in northern California. Five tick species were collected from mammals (Dermacentor occidentalis, Haemaphysalis leporispalustris, Ixodes pacificus, I. spinipalpis, I. woodi), but only I. pacificus was found on lizards. Dermacentor occidentalis, I. pacificus, and I. woodi occurred in both habitats, whereas H. leporispalustris and I. spinipalpis were found only on animals trapped in chaparral. The tick species most commonly encountered on mammals was D. occidentalis in chaparral and I. pacificus in woodland-grass. Peak infestation of mammals occurred in spring for I. pacificus immatures and H. leporispalustris, summer for D. occidentalis immatures, fall through spring for I. woodi immatures, and fall through winter for I. spinipalpis. The primary aim of the study was to quantify the relative importance of the western fence lizard (Sceloporus occidentalis), which is reservoir-incompetent for Borrelia burgdorferi sensu lato (s.l.), and mammalian B.burgdorferi s.l.-reservoirs as hosts for the immature stages of I. pacificus in spring. The estimated relative utilization by I. pacificus of the western fence lizard versus mammals was 88% for larvae and 99% for nymphs in chaparral in May. When tick infestation data were corrected for a two-fold lower efficiency of field examinations for rodents than for lizards, the western fence lizard still accounted for 78% of larval and 98% of nymphal feedings. In woodland-grass, 46% of 100 I. pacificus larvae and 100% of 52 nymphs recovered from mammals or western fence lizards during May-June were collected from the lizards. However, this may represent an underestimate of the importance of the western fence lizard as a larval host in this habitat because inclement weather during the late May sampling period doubtless resulted in significantly decreased lizard activity. In conclusion, the western fence lizard was more heavily utilized by I. pacificus immatures, especially nymphs, than were rodents. This revised version was published online in July 2006 with corrections to the Cover Date.     

Tick attachment cement – reviewing the mysteries of a biological skin plug system

The majority of ticks in the family Ixodidae secrete a substance anchoring their mouthparts to the host skin. This substance is termed cement. It has adhesive properties and seals the lesion during feeding. The particular chemical composition and the curing process of the cement are unclear. This review summarizes the literature, starting with a historical overview, briefly introducing the different hypotheses on the origin of the adhesive and how the tick salivary glands have been identified as its source. Details on the sequence of cement deposition, the curing process and detachment are provided. Other possible functions of the cement, such as protection from the host immune system and antimicrobial properties, are presented. Histochemical and ultrastructural data of the intracellular granules in the salivary gland cells, as well as the secreted cement, suggest that proteins constitute the main material, with biochemical data revealing glycine to be the dominant amino acid. Applied methods and their restrictions are discussed. Tick cement is compared with adhesives of other animals such as barnacles, mussels and sea urchins. Finally, we address the potential of tick cement for the field of biomaterial research and in particular for medical applications in future.     

Quantification of vitellogenesis and its control by 20-hydroxyecdysone in the ixodid tick,Amblyomma hebraeum

Ovaries of the ixodid tick, Amblyomma hebraeum Koch, grew rapidly after engorgment as a result of yolk uptake. At 26 °C, oviposition began by day 10 post-engorgement, plateaued on days 16-18, and ended by day 38. Vitellin (Vt) was partially purified from ovaries of day 10 engorged ticks by gel filtration and ion exchange chromatography. This Vt comprises seven major and several minor polypeptides. Two polypeptides (211 and 148 kD) from haemolymph of engorged female ticks corresponded to minor polypeptides of similar molecular weight in the ovary. The haemolymph titre of the 211 and 148 kD polypeptides increased up to the onset of oviposition. These polypeptides were absent in males and non-vitellogenic females (day 0 engorged or day 10 partially-fed females), and were thus designated as vitellogenin (Vg). Antibodies raised against haemolymph Vg211 and 148 recognized these polypeptides in partially purified Vt, as well as six of the seven major polypeptides. Using these antibodies we developed an indirect, competitive ELISA to quantify Vg. Rise in haemolymph Vg-concentration lagged slightly behind the rise in haemolymph ecdysteroid (ES)-concentration, and Vg-synthesis was stimulated by injections of 20E into non-vitellogenic females. These observations indicate that an ES is the vitellogenic hormone in A. hebraeum.     

Dynamic Changes in Lyme Disease Spirochetes During Transmission by Nymphal Ticks

Ticks are not crawling needles, merely delivering infectious agents to vertebrate hosts. A sophisticated interplay takes place between ticks, pathogens, and vertebrate hosts. The relationship between Ixodes ticks and the Lyme disease spirochetes they transmit involves subtle changes in spirochete populations that maximize their chances of being transmitted. An understanding of this complex interplay will, hopefully, allow the development of new tools to block transmission of tick-borne agents. This revised version was published online in July 2006 with corrections to the Cover Date.     

Species specificity in photoperiodic control of nymphal development in four species of cricket from north-west China

Photoperiodic regulation of nymphal development was examined in four species of cricket collected in the Xinjiang‐Uygur Autonomy Region, China (approximately 43°N, 81–89°E). Fifty percent of individuals of Modicogryllus frontalis reared at 28°C reached adulthood in approximately 80 days in conditions of 11 h light : 13 h dark (LD 11:13) to 14:10, and in approximately 95 days under LD 15:9 to 16:8. Melanogryllus desertus started adult emergence earliest under LD 16:8 at 28°C, but some individuals required much longer to mature, and thus two peaks of adult emergence occurred at approximately 60 and 160 days after hatching. More individuals hatched during the late peak in LD 18:6 than in LD 16:8. The mean nymphal period was approximately 100 days in LD 11:13 to 14:10. Both species showed short‐day type photoperiodic responses, but Mo. frontalis developed faster than did Me. desertus. The latter occupied a wider range of habitat conditions and was more variable in life cycle than the former. Modicogryllus burdigalensis started adult emergence earliest in LD 16:8 at 28°C in the six photoperiods tested, most adults emerging within 60 days. The mean nymphal period was 80 days in LD 15:9, 135 days in LD 14:10 and 80–100 days in LD 11:13 to 13:11, showing an intermediate type of photoperiodic response. Acheta domesticus was a long‐day type species, and the proportion of delayed individuals increased with decreasing photoperiod. In the dry climate of Xinjiang‐Uygur, nymphal overwintering seems to be successful in all of the four different patterns of nymphal development.