Larval Ticks Research Articles

Bovine fecal chemistry changes with progression of Southern Cattle Tick, Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) infestation

Surveillance for cattle fever ticks is an essential activity in the U.S. Cattle Fever Tick Eradication Program which prevents reestablishment of these tick vectors of the pathogens causing bovine babesiosis. Other methods of detecting tick infested cattle could augment current physical inspection of restrained cattle by program inspectors. The objective of this study was to determine whether a single infestation of ∼5000 Rhipicephalus (Boophilus) microplus larvae induced changes in fecal chemistry that were detectable using near-infrared reflectance spectroscopy (NIRS). Fecal samples were collected daily from 6 tick-infested and 6 non-infested Bos taurus yearling heifers. Each infested animal received ticks from one of 6 different strains of laboratory colonies of R. microplus. Date of drop and daily sum of engorged female ticks were tabulated to characterize each infestation. Cluster, common factor, principal component and MANOVA analyses were used to define and assess fecal spectra changes associated with experimental stages of infestation. Cluster analyses found no significant differences in fecal samples from each of the 6 infested heifers. Two shifts in fecal chemistry of infested animals were identified by three clusters of NIRS fecal spectra. The first cluster was comprised of samples from pre-infestation to 9 days after infestation, a period inclusive of larval tick attachment and feeding. The second cluster was comprised of samples from day 10–22 corresponding to the period of nymphal feeding, adult feeding, and early drop of engorged females. A third cluster was comprised of samples from days 23−46 corresponding to the period of engorged female drop and declining tick numbers. A Tukey-Kramer multiple comparison procedure identified significant differences in fecal spectra between five experimental stages of R. microplus infestation for principal component 1 including pre-infestation to nymphal feeding, pre-infestation to adult feeding, larval feeding to adult feeding, nymphal feeding to adult feeding and nymphal feeding to engorged female drop; for principal component 2 including pre-infestation to nymphal feeding, pre-infestation to adult feeding, and pre-infestation to engorged female drop; and for principal component 3 including pre-infestation to drop, and adult feeding to drop. These significant pair-wise comparisons reflect developmental phases of tick attachment and blood-feeding that define periods of increasing, peak and declining stress identified in two fecal chemistry shifts defined by three fecal spectra clusters. Among non-infested animals, two shifts in fecal chemistry were also detected by three fecal-spectra clusters that occurred in synchrony with those of their tick-infested counterparts. There were no significant differences in principal components or MANOVA analyses between infested and non-infested animals and the pattern of significant pair-wise Tukey-Kramer multiple comparisons for non-infested animals were similar to those of infested animals. This unintended confounding effect is attributed to the manner in which all 12 animals were preconditioned as a group, then isolated in randomly assigned blind stalls in a common barn facility for the study, creating the basis for physiological stress resonance among non-infested animals.

Acaricide activity of Piper macedoi Yunck essential oil against Rhipicephalus sanguineus

This study evaluated the acaricide efficacy of Piper macedoi essential oil on larvae of ticks of the species Rhipicephalus sanguineus. The essential oil was extracted by hydrodistillation in a Clevenger-type apparatus. The test consisted of six treatments: from the group I to IV, samples corresponded to different concentrations of essential oil (500 μg.mL-1; 250 μg.mL-1; 100 μg.mL-1 and 50 μg.mL-1) diluted in Tween 80 at 2%. Groups V and VI corresponded to the negative controls (with distilled water and Tween 80 to 2%) and the positive control (with acaricide Amitraz at 12.5%), respectively. The essential oil was rich in apiole (39.81%) and dillapiole (26.47%). The essential oil of P. macedoi presented an activity against the larvae of R. sanguineus, with a better efficiency observed for concentrating 500 μg.mL-1, mortality of 80.67%, indicating a dose-dependent response.

Evaluation of deltamethrin-loaded Zn-Fe, Zn-Al-GA layered double hydroxide, and Fe-O nanoparticles against resistant Rhipicephalus annulatus ticks.

The over use of deltamethrin has resulted in the development of resistance. Thus, the search for a new method to improve its acaricidal effect is the target of the present study. Layered double hydroxide (LDH) compounds are now used as drug delivery systems. Deltamethrin (Butox®) was loaded on Zn-Fe LDH, Zn-Al-GA layered double hydroxide (LDH), and Fe-oxide nanoparticles NPs. Then its acaricidal efficacy was evaluated. The nanocomposites (NCs) were prepared by the co-precipitation method and characterized before and after deltamethrin loading. The deltamethrin-loaded NCs were applied against the phenotypically resistant Rhipicephalus annulatus tick (adult and larvae). The adult ticks treated by Butox® alone or Butox® loaded nanocomposites at different concentrations showed no mortality. A significant (P≤0.05) reduction in egg production index was observed at the recommended dose (X) (1 μl/ml distilled water) and its bi-folds (2X, 3X, and 4X) in ticks treated with deltamethrin/Zn-Fe LDH nanocomposites compared to deltamethrin alone. Moreover, no significant difference (P>0.05) was recorded in larval mortality between the treatments with deltamethrin alone and its loaded nanocomposites. Also, the nanomaterials alone without conjugation with deltamethrin revealed a low mortality rate. Deltamethrin-loaded nanocomposites even improve effect against adult tick but cannot overcome tick resistance to deltamethrin that needs more search. The acaricidal activity of deltamethrin was not improved after loading it with these nanocomposites.

Molecular investigation on transstadial/transovarial transmission of Hepatozoon canis in Rhipicephalus sanguineus sensu lato ticks

ABSTRACT To investigate the possibility of transovarial and transstadial transmission of Hepatozoon canis, a total of 953 Rhipicehlalus sanguineus sensu lato ticks at different stages were collected from 231 stray dogs housed at Blue cross, and 10 dogs presented at small animal clinics. The engorged nymphs collected from the cracks and crevices of Blue cross were allowed to moult in the laboratory. The ticks collected from the host and moulted ticks were pooled according to their sex and feeding status as fed, unfed males and females. Each tick was dissected individually and examined microscopically, then pooled as H. canis positive and negative. Microscopic examination of the 953 ticks revealed H. canis oocysts in 228 (89.76%) unfed adult males and females which moulted from the nymphs collected from cracks and crevices. Positivity of 74% was observed in unfed females while all unfed males (100%) were positive. Amongst the 213 fed males, 174 (81.69%) were positive, while only 210 (43.20%) of the 486 fed females were positive. No oocyst was detected from ticks collected from dogs presented to clinics. The occurrence of H. canis DNA in the tick was confirmed by PCR assay based on the amplification of 666 bp fragment of partial 18S rRNA gene. Amplification was observed in pooled samples collected from Blue cross, whereas no amplification was observed in ticks collected from dogs presented to clinics. The DNA samples from tick larvae did not show amplification, which indicated that H. canis is not transmitted transovarially.

Heritable variation in host quality as measured through an ectoparasite's performance

Obligate parasites need one or more hosts to complete their life cycle. However, hosts might show intraspecific variation in quality with respect to the parasites themselves, thus affecting on‐host and off‐host parasite performance. High heritability in host quality for the parasite may therefore exert long‐lasting selective pressures on the parasite and influence host–parasite coevolution. However, the amount of variation and heritability in host quality are unknown for most parasite species, especially in wild populations of hosts. Both measures were estimated in a wild‐caught birdParus majorthat was experimentally infested by two developmental stages (larva and nymph) of a ectoparasite (the tickIxodes arboricola). We examined variation in host quality through variation in tick performance, namely the on‐host performance (attachment success, feeding time, engorgement weight and feeding success) and the off‐host performance (moulting time, moulting success and overall survival). Herein we also investigated the influence on tick performance of host traits linked with the bird's life history and physiology such as body condition, sex, age and haematocrit. By correlating tick performance variables between larvae and nymphs feeding on the same bird at different times, we found a significant correlation in attachment success, suggesting consistent among‐host variation for this performance measure, but no significant larva‐nymph correlations for the other tick variables. Animal models relating tick performance variables to the host pedigree showed a strong heritable signal for host quality as measured through tick feeding time, and lower but substantial estimates in other performance variables. With regard to the host traits, feeding success and survival of tick larvae were lower on female birds, and nymphal survival was higher on older birds. Larval feeding time was negatively correlated with host haematocrit. This is one of the first studies showing consistent intraspecific variation and heritability of host quality for a multi‐stage ectoparasite.

First isolation of Borrelia lusitaniae DNA from green lizards (Lacerta viridis) and Ixodes ricinus ticks in the Czech Republic

This study aimed to reveal whether green lizards (Lacerta viridis), common hosts of tick larvae and nymphs, might be involved in the transmission cycle of Borrelia burgdorferi sensu lato in the Czech Republic. Green lizards were sampled in two areas at the Tiché Údolí Nature Reserve (site A: 50.1482 N, 14.3669E; site B: 50.1476 N, 14.3745 E), Central Bohemian Region, Czech Republic. The skin biopsy specimens and attached ticks (if any) were collected from 52 captured lizards. Also, questing ticks from both areas were collected by flagging. The touchdown polymerase chain reaction and gel electrophoresis revealed Borrelia lusitaniae in three lizard tissue samples. Most lizards (19/30, 63%) had at least one Borrelia positive tick. Borrelia lusitaniae formed 92% (34/37) and 59% (17/29) of all borreliae detected in larvae and nymphs, respectively. Borrelia lusitaniae (6/10, 60%) was also the major pathogen in questing nymphs from site B. At site A, 13% (2/16) of questing nymphs were positive for B. lusitaniae. Based on our data, it can be assumed that B. lusitaniae is a common pathogen at lizard sites in the Czech Republic, and further research to prove this hypothesis is therefore highly recommended. As lizards often inhabit urban areas, the data presented may also contribute to raising awareness of the possible spread and risk of Borrelia infection.

Acaricidal activity of Foeniculum vulgare against Rhipicephalus annulatus is mainly dependent on its constituent from trans-anethone.

Globally, the economic losses due to hard ticks infestation and the control of the associated diseases have been calculated at USD $13.9–18.7 billion per year. The economic impact is related to its direct damage to the skins, blood loss, anemia, severe immunological reactions and indirect losses that related to the effects of hemoparasites, cost of treatment for clinical cases and expenses incurred in the control of ticks. The current study evaluated the acaricidal activities of fennel Foeniculum vulgare essential oil and its main components; trans-anethole and fenchone; against R. annulatus. GC–MS analysis revealed that this oil contained 16 components representing 99.9% of the total identified compounds with E-anethole being the predominant component(64.29%), followed by fenchone (9.94%). The fennel oil and trans-anethole showed significant acaricidal activities. The LC50 of the fennel oil was attained at concentrations of 12.96% for adult ticks and 1.75% for tick larvae meanwhile the LC50 of trans-anethole was reached at concentrations of 2.36% for adult tick and 0.56% for tick larvae. On the contrary, fenchone showed no any significant adulticidal activities and its LC50 attained at a concentration of 9.11% for tick larvae. Regarding repellence activities, trans-anethole achieved 100% repellency at the concentration of 10% while fennel showed 86% repellency at the same concentration. Fenchone showed no repellency effect. Treatment of larvae with fennel, trans-anethole, and fenchone LC50 concentrations significantly inhibited the acetylcholinesterase activity. Meanwhile, glutathione s-transferase activity was significantly decreased in fennel treated larvae but no significant effect was found in the larvae of trans-anethole and fenchone groups. These results indicate that the acaricide effect of fennel oil may attributed to its high content of trans-anethole. This was supported by potent adulticidal, larvicidal, and repellency effects of trans-anethole against Rhipeciphalus annulatus tick and therefore it could be included in the list of acaricide of plant origin.

Winter Tick Burdens for Moose Are Positively Associated With Warmer Summers and Higher Predation Rates

Climate change is expected to modify host-parasite interactions which is concerning because parasites are involved in most food-web links, and parasites have important influences on the structure, productivity and stability of communities and ecosystems. However, the impact of climate change on host–parasite interactions and any cascading effects on other ecosystem processes has received relatively little empirical attention. We assessed host-parasite dynamics for moose (Alces alces) and winter ticks (Dermacentor albipictus) in Isle Royale National Park over a 19-year period. Specifically, we monitored annual tick burdens for moose (estimated from hair loss) and assessed how it covaried with several aspects of seasonal climate, and non-climatic factors, such as moose density, predation on hosts by wolves (Canis lupus) and wolf abundance. Summer temperatures explained half the interannual variance in tick burden with tick burden being greater following hotter summers, presumably because warmer temperatures accelerate the development of tick eggs and increase egg survival. That finding is consistent with the general expectation that warmer temperatures may promote higher parasite burdens. However, summer temperatures are warming less rapidly than other seasons across most regions of North America. Therefore, tick burdens seem to be primarily associated with an aspect of climate that is currently exhibiting a lower rate of change. Tick burdens were also positively correlated with predation rate, which could be due to moose exhibiting risk-sensitive habitat selection (in years when predation risk is high) in such a manner as to increases the encounter rate with questing tick larvae in autumn. However, that positive correlation could also arise if high parasite burdens make moose more vulnerable to predators or because of some other density-dependent process (given that predation rate and moose density are highly correlated). Overall, these results provide valuable insights about interrelationships among climate, parasites, host/prey, and predators.

Beech tree masting explains the inter-annual variation in the fall and spring peaks of Ixodes ricinus ticks with different time lags

BackgroundThe tick Ixodes ricinus is an important vector of tick-borne diseases including Lyme borreliosis. In continental Europe, the nymphal stage of I. ricinus often has a bimodal phenology with a large spring peak and a smaller fall peak. There is consensus about the origin of the spring nymphal peak, but there are two alternative hypotheses for the fall nymphal peak. In the direct development hypothesis, larvae quest as nymphs in the fall of the same year that they obtained their larval blood meal. In the developmental diapause hypothesis, larvae overwinter in the engorged state and quest as nymphs one year after they obtained their larval blood meal. These two hypotheses make different predictions about the time lags that separate the larval blood meal and the density of questing nymphs (DON) in the spring and fall.MethodsInter-annual variation in seed production (masting) by deciduous trees is a time-lagged index for the density of vertebrate hosts (e.g., rodents) which provide blood meals for larval ticks. We used a long-term data set on the masting of the European beech tree and a 15-year study on the DON at 4 different elevation sites in western Switzerland to differentiate between the two alternative hypotheses for the origin of the fall nymphal peak.ResultsQuesting I. ricinus nymphs had a bimodal phenology at the three lower elevation sites, but a unimodal phenology at the top elevation site. At the lower elevation sites, the DON in the fall was strongly correlated with the DON in the spring of the following year. The inter-annual variation in the densities of I. ricinus nymphs in the fall and spring was best explained by a 1-year versus a 2-year time lag with the beech tree masting index. Fall nymphs had higher fat content than spring nymphs indicating that they were younger. All these observations are consistent with the direct development hypothesis for the fall peak of I. ricinus nymphs at our study site. Our study provides new insight into the complex bimodal phenology of this important disease vector.ConclusionsPublic health officials in Europe should be aware that following a strong mast year, the DON will increase 1 year later in the fall and 2 years later in the spring. Studies of I. ricinus populations with a bimodal phenology should consider that the spring and fall peak in the same calendar year represent different generations of ticks.Graphical

New insight on some selected nanoparticles as an effective adsorbent toward diminishing the health risk of deltamethrin contaminated water.

Deltamethrin is a widely used insecticide that kills a wide variety of insects and ticks. Deltamethrin resistance develops as a result of intensive, repeated use, as well as increased environmental contamination and a negative impact on public health. Its negative impact on aquatic ecology and human health necessitated the development of a new technique for environmental remediation and wastewater treatment, such as the use of nanotechnology. The co-precipitation method was used to create Zn-Fe/LDH, Zn-AL-GA/LDH, and Fe-oxide nanoparticles (NPs), which were then characterized using XRD, FT-IR, FE-SEM, and HR-TEM. The kinetic study of adsorption test revealed that these NPs were effective at removing deltamethrin from wastewater. The larval packet test, which involved applying freshly adsorbed deltamethrin nanocomposites (48 hours after adsorption), and the comet assay test were used to confirm that deltamethrin had lost its acaricidal efficacy. The kinetics of the deltamethrin adsorption process was investigated using several kinetic models at pH 7, initial concentration of deltamethrin 40 ppm and temperature 25°C. Within the first 60 min, the results indicated efficient adsorption performance in deltamethrin removal, the maximum adsorption capacity was 27.56 mg/L, 17.60 mg/L, and 3.06 mg/L with the Zn-Al LDH/GA, Zn-Fe LDH, and Fe Oxide, respectively. On tick larvae, the results of the freshly adsorbed DNC bioassay revealed larval mortality. This suggests that deltamethrin's acaricidal activity is still active. However, applying DNCs to tick larvae 48 hours after adsorption had no lethal effect, indicating that deltamethrin had lost its acaricidal activity. The latter result corroborated the results of the adsorption test's kinetic study. Furthermore, the comet assay revealed that commercial deltamethrin caused 28.51% DNA damage in tick cells, which was significantly higher than any DNC. In conclusion, the NPs used play an important role in deltamethrin decontamination in water, resulting in reduced public health risk. As a result, these NPs could be used as a method of environmental remediation.

Investigation on haplotypes of ixodid ticks and retrospective finding of Borrelia miyamotoi in bank vole (Myodes glareolus) in Switzerland

The current status of tick species, important tick-borne bacteria and protozoan parasites is well-documented in Switzerland. However, reports on the genetic diversity and geographical relationships of tick species in this country appear to be in part lacking or outdated. Thus, the aim of this study was to collect ticks from various host species in southern Switzerland, to compare them in a geographical context and to screen in these samples rare tick-borne pathogens hitherto not reported or having low prevalence in Switzerland.In 2019–2020 altogether 177 ixodid ticks were collected from the vegetation, as well as from humans (n = 17), dogs (n = 23), cats (n = 41), red deer (n = 8), a European rabbit and a European hedgehog at 25 locations in three cantons of south Switzerland. Tick species were identified morphologically, followed by DNA extraction and comparison of mitochondrial haplotypes with molecular-phylogenetic methods. Tick DNA extracts, as well as sixty-two rodent liver or spleen tissue DNA extracts (representing six species) available from 2005 to 2006 were screened for trypanosomes, Occidentia massiliensis and Borrelia miyamotoi.Morphologically, three tick species were identified: Ixodes ricinus (n = 170), Rhipicephalus sanguineus sensu lato (n = 6) and I. hexagonus (n = 1). In contrast to companion animals (dogs, cats) immature ticks (larvae and nymphs) predominated on humans, which was a highly significant association (P < 0.0001). Molecular comparison of the cytochrome c oxidase subunit I (cox1) gene with GenBank data established the species as R. sanguineus sensu stricto and confirmed I. hexagonus, both showing 99.8–100% sequence identity to conspecific ticks from northern Italy. Seventy-nine specimens morphologically identified as I. ricinus revealed high 16S rRNA gene haplotype diversity and represented two phylogenetic groups. Two I. ricinus haplotypes from Switzerland belonged to the same haplogroup with I. inopinatus from Spain, Germany and Austria as well as with I. ricinus reported from a broad geographical range of Europe (including Italy, the Netherlands, Poland, Latvia and Sweden). All 141 tick DNA extracts (from five R. sanguineus s.l., 135 I. ricinus and one I. hexagonus) and 62 rodent tissue DNA extracts were negative for trypanosomes and O. massiliensis. However, B. miyamotoi was identified in a bank vole (Myodes glareolus) and three ticks by sequencing.From Switzerland, this is the first report of tick haplotypes that are phylogenetically closely related to I. inopinatus. However, based on their morphology, both specimens are considered as I. ricinus. These results highlight the importance that the identification of I. inopinatus should be based on coherent morphologic and molecular properties. This is also the first report of rodent-borne B. miyamotoi in Switzerland. Taking into account the year of collection (2005), in a chronological order this might be the first indication of B. miyamotoi in any rodent species in Europe.

Molecular Survey of Anaplasmataceae Agents and Coxiellaceae in Non-Hematophagous Bats and Associated Ectoparasites from Brazil

The Anaplasmataceae family (order Rickettsiales) encompasses obligately intracellular bacteria of the genera Anaplasma, Ehrlichia, and Neorickettsia. Together with Coxiella burnetii (Coxiellaceae family, order Legionellales), these bacteria represent important causative agents of diseases in humans and animals. The scarcity of studies that investigated the occurrence of these agents in bats and their associated ectoparasites, emphasizes the need to achieve a better understanding of the role of these animals in the maintenance of such bacteria. Herein, 418 samples (133 blood, 135 spleen, and 150 ectoparasites) are collected from 135 non-hematophagous bats belonging to 12 species in a periurban area of Campo Grande city, Mato Grosso do Sul state, midwestern Brazil. In the results, 1.65% (7/418), 12.04% (50/418), and 13.63% (57/418) of samples are positive in PCR assays for Anaplasma spp. (16S rRNA gene), Ehrlichia spp. (dsb gene), and Neorickettsia spp. (16S rRNA gene), respectively. Anaplasma spp. and Neorickettsia spp. are detected in one (5.26%) Ornithodoros hasei tick larva. Ehrlichia spp. is detected in 14% of bat flies (represented by Megistopoda aranea, Trichobius costalimai, and Strebla hertigi), 6% of tick larvae (O. hasei), 12% of Spinturnicidae mites (represented by Periglischrus sp., P. torrealbai, and P. acutisternus), and 38% of Macronyssidae mites (Steatonyssuss sp.). The obtained sequences are observed to be similar to Anaplasma phagocytophilum (97.42–97.6% identified), Ehrlichia minasensis (96.73–100% identified), Neorickettsia risticii (96.7–100% identified), and Neorickettsia findlayensis (95.07–100% identified) by BLASTn analyses, and closely related to Ehrlichia ruminantium by phylogenetic analyses based on the gltA gene. No bat samples (blood/spleen) are positive in the qPCR assay for C. burnetii based on the IS1111 gene. The present work shows, for the first time, the occurrence of Anaplasmataceae in bats and associated ectoparasites (ticks, mites, and bat flies) from Brazil.

Experimental evidence for opposing effects of high deer density on tick-borne pathogen prevalence and hazard

BackgroundIdentifying the mechanisms driving disease risk is challenging for multi-host pathogens, such as Borrelia burgdorferi sensu lato (s.l.), the tick-borne bacteria causing Lyme disease. Deer are tick reproduction hosts but do not transmit B. burgdorferi s.l., whereas rodents and birds are competent transmission hosts. Here, we use a long-term deer exclosure experiment to test three mechanisms for how high deer density might shape B. burgdorferi s.l. prevalence in ticks: increased prevalence due to higher larval tick densities facilitating high transmission on rodents (M1); alternatively, reduced B. burgdorferi s.l. prevalence because more larval ticks feed on deer rather than transmission-competent rodents (dilution effect) (M2), potentially due to ecological cascades, whereby higher deer grazing pressure shortens vegetation which decreases rodent abundance thus reducing transmission (M3).MethodsIn a large enclosure where red deer stags were kept at high density (35.5 deer km−2), we used an experimental design consisting of eight plots of 0.23 ha, four of which were fenced to simulate the absence of deer and four that were accessible to deer. In each plot we measured the density of questing nymphs and nymphal infection prevalence in spring, summer and autumn, and quantified vegetation height and density, and small mammal abundance.ResultsPrevalence tended to be lower, though not conclusively so, in high deer density plots compared to exclosures (predicted prevalence of 1.0% vs 2.2%), suggesting that the dilution and cascade mechanisms might outweigh the increased opportunities for transmission mechanism. Presence of deer at high density led to shorter vegetation and fewer rodents, consistent with an ecological cascade. However, Lyme disease hazard (density of infected I. ricinus nymphs) was five times higher in high deer density plots due to tick density being 18 times higher.ConclusionsHigh densities of tick reproduction hosts such as deer can drive up vector-borne disease hazard, despite the potential to simultaneously reduce pathogen prevalence. This has implications for environmental pathogen management and for deer management, although the impact of intermediate deer densities now needs testing.Graphical abstract

Cypermethrin and ivermectin resistance in field populations of Rhipicephalus sanguineus sensu lato (Latrielle, 1806) collected from dogs in south India.

The abuse of acaricides for control of Rhipicephalus sanguineus sensu lato ticks led to a strong selection pressure for acaricide resistance. Data on acaricide resistance in R. sanguineus s.l. populations from India is meager though veterinarians frequently report treatment failures. The present study was aimed to characterize the level of resistance in R. sanguineus s.l. against most commonly used drugs, cypermethrin and ivermectin in Andhra Pradesh, south India. Fourteen adult female tick populations were collected from naturally infested dogs at veterinary clinics, residence and stray dog in nine state municipal corporations/municipalities. Information on the history of dog treatment with acaricides was obtained by interviewing dog owners. The larval packet test (LPT) and larval immersion test (LIT) were used on the larvae of ticks to test for resistance to cypermethrin and ivermectin, respectively. Mortality data of each tick population was analyzed by probit analysis. Corresponding to the most susceptible population, thirteen (92.6%) and six (42.9%) tick populations were regarded as resistant to cypermethrin and ivermectin, respectively. The phenotypic level of cypermethrin (resistance ratios at LC50% varied from 1.55 to 13.29) and ivermectin (resistance ratios at LC50% ranged from 1.16 to 4.79) resistance varied distinctly between the populations. Additionally, all the populations were tested using PCR to demonstrate the frequency of the point mutation in sodium channel gene that corresponds with resistance of R. sanguineus s.l. to cypermethrin. A nucleotide substitution (T2134C) on domain III segment VI of the sodium channel gene was noticed in phenotypically resistant tick populations on DNA sequencing. Ivermectin resistance in the brown dog ticks is reported for the first time from India.

Efficacy of low-dose fipronil bait against blacklegged tick (Ixodes scapularis) larvae feeding on white-footed mice (Peromyscus leucopus) under simulated field conditions

BackgroundLyme disease, caused primarily by Borrelia burgdorferi sensu stricto, is the most prevalent vector-borne disease in the United States. Treatment of rodent pathogen reservoirs with an oral acaricide may suppress the production of infected host-seeking ticks posing a risk for human infection. A previous study showed that an oral fipronil bait effectively controlled larval Ixodes scapularis ticks on white-footed mice (Peromyscus leucopus) up to 15 days post-bait exposure. The present study expands upon this finding by exposing group-housed white-footed mice to fipronil bait under simulated field conditions prior to tick infestation.MethodsMice (n = 80) were housed in groups of 10 within large enclosures and offered a choice between fipronil bait within a commercial bait station and an alternative diet. The mice were assigned to two treatment groups and two control groups to undergo bait exposure durations of either 24 h (reduced) or 168 h (extended). Groups were further differentiated by the time point post-bait exposure when larval ticks were applied to mice within feeding capsules (reduced day 1, day 15; extended day 21, day 35). For 4 days post-tick introduction, attached larvae were observed by microscopy and replete larvae were recovered. Replete larvae were monitored for molting success. Plasma was collected from all treatment group mice to obtain fipronil plasma concentrations (CP).ResultsThe fipronil bait (0.005% fipronil) was palatable and controlled larval ticks on white-footed mice when presented under simulated field conditions. Efficacy in preventing attached larvae from feeding to repletion was 100% (day 1), 89.0% (day 15), 85.8% (day 21), and 75.2% (day 35). When also considering molting success, the fipronil bait prevented 100% (day 1), 91.1% (day 15), 91.7% (day 21), and 82.5% (day 35) of larvae attaching to mice from molting. The mean CP per mouse was 191.5 ng/ml (day 1), 29.4 ng/ml (day 15), 10.6 ng/ml (day 21), and 1.0 ng/ml (day 35).ConclusionsThe results suggest that fipronil bait will be consumed by white-footed mice in the presence of an alternative diet, and effectively control larval ticks on treated mice. A field trial is needed to confirm the results of this study. Low-dose fipronil bait may provide a cost-effective means of controlling blacklegged ticks to be integrated into tick management programs.Graphical

Integrated Tick Management in South Central Wisconsin: Impact of Invasive Vegetation Removal and Host-Targeted Acaricides on the Density of Questing Ixodes scapularis (Acari: Ixodidae) Nymphs.

As tick-borne disease incidence increases and pathogens expand into new areas, the need for effective tick management strategies is paramount. In this 5-yr study (2014-2018) conducted in south central Wisconsin, we assessed whether an integrated tick management approach, deployed during peak tick activity (May-August), was more effective at reducing black-legged ticks (Ixodes scapularis Say (Ixodida: Ixodidae)), than individual interventions. Using a factorial design, invasive vegetation removal (Amur honeysuckle, Lonicera maackii Ruprecht (Dipsacales: Caprifoliaceae) and common buckthorn, Rhamnus cathartica Linnaeus (Rosales: Rhamnaceae)) was coupled with deployments of permethrin-treated cotton nesting materials (tick tubes) that target the white-footed mouse (Peromyscus leucopus Rafinesque (Rodentia: Cricetidae)). Results show that the probability of encountering a larval tick by drag sampling was unaffected by treatments at the cumulative 5-yr level. However, vegetation removal significantly reduced larval encounters in 2014, 2015, and 2018, by 33%, 57%, and 61% respectively, and reduced the density of questing nymphal (DON) ticks by 45% in 2015 compared to controls. Despite the limited effect on DON, vegetation removal significantly reduced the cumulative 5-yr density of Borrelia burgdorferi sensu stricto infected nymphs (DIN) (70%) compared to controls as a result of decreased nymphal infection prevalence. Sites treated with tick tubes had lower DIN (66%) and DON (54%) across the study and nymphs were reduced every year following the initial year of deployment compared to controls. Combining treatments did not further reduce DIN or DONs. We conclude that long-term integration of tick tubes with invasive vegetation removal does not provide additional benefit over individual treatments alone.

Are Virginia opossums really ecological traps for ticks? Groundtruthing laboratory observations

Virginia opossums (Didelphis virginiana) are a common synanthrope in North America, and serve as host to many species of ectoparasites. Research on captive Virginia opossums estimated that opossums eat, on average, 5500 larval ticks (Acari: Ixodidae) per week. To investigate this apparent preference exhibited by opossums for ingesting ticks, we comprehensively analyzed stomach contents of 32 Virginia opossums from central Illinois. Using a dissecting microscope, we searched the contents exhaustively for ticks and tick body parts, without sieving or pre-rinsing the stomach contents. We did not locate any ticks or tick parts in the stomach contents of Virginia opossums. We also performed a vigorous literature search for corroborating evidence of tick ingestion. Our search revealed 23 manuscripts that describe diet analyses of Virginia opossums, 19 of which were conducted on stomach or digestive tract contents and four of which were scat-based analyses. None of the studies identified ticks in their analyses of diet items. We conclude that ticks are not a preferred diet item for Virginia opossums. Considering that wildlife unconditioned to laboratory conditions may exhibit non-typical behaviors, we recommend that lab-based studies of wildlife behavior be groundtruthed with studies based in natural conditions.

Molecular identification of Haemaphysalis sulcata (Acari: Ixodidae) larval stages collected using the Berlese funnel in Northern Iran

Molecular detection techniques based on conventional PCR and sequencing of the partial genome of living organisms are considered an important tool for identification. BLASTn and comparison of evolutionary relationships between taxa are the next analysis steps on sequences. The construction of a phylogenetic tree is done using genetic distance difference; nucleotide substitution models and comparison with the outgroup. In the present study, two samples of tick larvae were collected in Mirafzal forest area (36°07'39.0N 53°35'45.0E) located in Mazandaran province, northern Iran using the Berlese funnel. Tick samples were initially identified using a generic identification key (Bregetova et al. 1955) and in order to determine specific identification of species, PCR of partial internal transcribed spacer 2 (ITS2) gene and Sanger sequencing were performed. Morphological results showed that tick samples belonged to Haemaphysalis genus because they had a relatively short capitulum, no eyes with festoons. DNA extraction, PCR amplification and sequencing of a partial fragment (ca. 1400-bp) ITS2 gene of tick sample were done, successfully. The results of BLASTn showed that the larvae belonged to the species Haemaphysalis sulcata. A nucleotide DNA sequence of ITS2 was submitted to GenBank under the accession number MW929218. The molecular technique of this study is recommended to identify immature and dead tick samples that cannot be reared until the adult stage. The present study was the first report on the identification of H. sulcata larvae isolated from the Berlese funnel. The result of this study will help to better understand the biology of this tick and the presence of immature larval stages with questing behaviour in the soil environment.

Rearing of Hyalomma marginatum (Acarina: Ixodidae) under laboratory conditions in Morocco

Hyalomma marginatum Koch is one of the main tick vectors of human and animal tick-borne diseases. The objective of this study was to establish standard procedures for rearing H. marginatum under laboratory conditions. Such laboratory tick populations are required to study acaricide resistance of Hyalomma ticks. In our rearing program, larvae and nymphs were fed on New Zealand white rabbits, whereas adults were fed on sheep. Non-parasitic stages were held at 18 and 28 °C to study the effect of temperature on development and survival. In our experiments, H. marginatum ticks have maintained the characteristics of a two-host life cycle. The engorged larvae did not detach and moulted on the rabbit, after which the emerged nymphs continued to feed on the same animal. The life cycle duration of H. marginatum was influenced by temperature, with each non-parasitic stage—i.e., larva and nymph molting—developing faster at 28 than at 18 °C; preoviposition and oviposition periods were shorter at 28 than at 18 °C. At 18 °C, no eggs hatched. The whole cycle from the collection of an engorged field tick until the emergence of second-generation larvae took 189 days. One such tick on average results in 3500 eggs which over time, taking into account the losses at each developmental stage, develop into 1200 adult ticks. Rearing these ticks a second generation therefore could result in millions of larval ticks.

The deletion of chiMaD1, a horizontally acquired chitinase of Metarhizium anisopliae, led to higher virulence towards the cattle tick (Rhipicephalus microplus).

The first line of the Arthropods defense against infections is the hard-structured exoskeleton, a physical barrier, usually rich in insoluble chitin. For entomopathogenic fungi that actively penetrate the host body, an arsenal of hydrolytic enzymes (as chitinases and N-acetylglucosaminidases), that break down chitin, is essential. Notably, twenty-one putative chitinase genes have been identified in the genome of Metarhizium anisopliae, a generalist entomopathogenic fungus. As a multigenic family, with enzymes that, presumably, perform redundant functions, the main goal is to understand the singularity of each one of such genes and to discover their precise role in the fungal life cycle. Specially chitinases that can act as virulence determinants are of interest since these enzymes can lead to more efficient biocontrol agents. Here we explored a horizontally acquired chitinase from M. anisopliae, named chiMaD1. The deletion of this gene did not lead to phenotypic alterations or diminished supernatant's chitinolytic activity. Surprisingly, chiMaD1 deletion enhanced M. anisopliae virulence to the cattle tick (Rhipicephalus microplus) larvae and engorged females, while did not alter the virulence to the mealworm larvae (Tenebrio molitor). These results add up to recent reports of deleted genes that enhanced entomopathogenic virulence, showing the complexity of host-pathogen interactions.