Description of Rhipicephalus hibericus sp. nov. (Ixodoidea: Ixodidae), a species of the Rhipicephalus sanguineus group in southwestern Europe.

We describe all the life stages of Rhipicephalus hibericus n. sp., provide the types, and present molecular support for a new species of the Rhipicephalus sanguineus sensu lato group, present in southwestern Europe, that has been historically confused with Rhipicephalus turanicus Pomerantzev, 1940. A new name is proposed for this taxon because it was impossible to ascribe to types of already described species in the group, deposited for more than 100 years in natural history institutions. The males have a dorsum showing deep and coarse punctations (absent in Rhipicephalus sanguineus sensu stricto) and adanal plates with large punctations (absent in R. sanguineus s.s.); the tail of the spiracular plate is as wide as the closest festoon (half the width in R. sanguineus s.s.). Females have large punctations in dorsal fields, a wide spiracular plate, and a "V" shaped genital opening; such a combination of characters cannot be found in other species of the group. Immatures are described from specimens collected on hosts (Rodentia and Eulipotyphla). Both larvae and nymphs are markedly smaller than R. sanguineus s.s. Nymphs display long, backward-projected auriculae; larvae are almost half the size of R. sanguineus s.s. The new species can hybridize with R. sanguineus s.s. in laboratory colonies producing an unfertile F2, laying brown and dry eggs that did not hatch. Phylogenetic analysis of partial coxI gene sequences placed R. hibericus in a well-supported clade with other sequences of R. sanguineus s.l. from Portugal, as a sister clade of R. sanguineus s.s. The new species does not belong to the R. turanicus group of species. Both 12S and 16S partial gene sequences were not as precise in the correct phylogenetic placement of R. hibericus, in part probably due to the existence of erroneously identified sequences in GenBank©. This description, together with the previous reinstatement of Rhipicephalus secundus and Rhipicephalus rutilus, and the description of the neotypes of R. sanguineus s.s. should help researchers to adequately identify their collections. Our findings demonstrate that R. turanicus is absent in southwestern Europe. Old collections should be re-examined to provide the actual range of the new species.

Morphological and molecular characterization of Rhipicephalus microplus and Rhipicephalus annulatus from selected states of southern India.

Ticks of the genus Rhipicephalus infesting cattle are the primary animal pests responsible for the annual economic loss of billions of dollars. Due to the morphological resemblance among the members of the Rhipicephalus (Boophilus) genus, species identification is very difficult. In this study, the adult R. annulatus and R. microplus ticks from two south Indian states viz., Kerala and Karnataka were subjected to morphological and molecular characterization. The R. microplus isolates from south India differed morphologically from true R. microplus clade A ticks. The ventral spur on the first pedipalp observed in male R. microplus was similar to that of R. australis. The phylogenetic analysis revealed that the R. microplus from these states clustered with R. microplus clade C. The mitochondrial cytochrome c oxidase I (COI) was identified as the preferred molecular marker compared to the internal transcribed spacer 2 (ITS2). The interspecific divergence between R. microplus and R. annulatus isolates from South India was 7.9 per cent based on COI. Moreover, based on COI, the R. microplus isolates revealed higher intraspecific divergence (2.9%) than R. annulatus (1%). The ITS2 sequences failed to differentiate R. microplus and R. annulatus.

Rhipicephalus annulatus, R. australis or R. microplus? Discordance between morphological and genetic data among three cattle tick species.

The taxonomy of ticks of the subgenus Boophilus has been extensively debated and is often complicated by the high intraspecific variation of morphological features between species. Notably, the cattle tick Rhipicephalus microplus is a species complex consisting of Rhipicephalus annulatus, Rhipicephalus australis and the three mitochondrial clades (A-C) of R. microplus. To gain insight into the taxonomic status of this species complex, we performed morphological and molecular analyses on these cattle ticks across four states in peninsular Malaysia. We morphologically identified 60 males and 104 females of R. microplus, 298 males and 374 females of R. australis, and one R. annulatus male in our field collection, of which the latter two species have never been recorded in Malaysia. However, all three morphologically identified species were molecularly assigned as R. microplus clade A based on the barcoding cytochrome oxidase subunit I (COI) analysis. The discrepancy between morphological and genetic data highlights an urgent need for further exploration and in-depth research into the taxonomic status of these sympatric tick species.

Five ixodid tick species including two morphotypes of Rhipicephalus turanicus on nestlings of Eurasian eagle owl (Bubo bubo) from south-eastern Bulgaria.

BACKGROUND: Birds are major hosts for many tick species (Acari: Ixodidae, Argasidae), and their role is especially important in transporting ticks over large distances along their seasonal migratory routes. Accordingly, most studies across Europe focus on the importance of avian hosts in tick dispersal, and less emphasis is laid on resident birds and their role in supporting tick life cycles. Eurasian eagle owls (Bubo bubo) exemplify the latter, but all the few studies on their tick infestation were carried out in Western Europe and even those did not involve a large sample size and did not assess infestation prevalence in natural habitats. METHODS: In this study, 320 ixodid ticks were collected from nestlings of Eurasian eagle owls during the period 2018-2020 in Bulgaria in south-eastern Europe. These ticks were analysed morphologically, and selected specimens molecularly based on cytochrome c oxidase subunit I (cox1) gene. The effects of environmental and habitat-related conditions and of the species of prey eaten by eagle owls on tick infestation were also evaluated. RESULTS: The majority of ticks were identified as adults of Rhipicephalus turanicus (n = 296). In addition, 15 Hyalomma marginatum (three males, 11 nymphs and a larva), one female of Haemaphysalis erinacei and of Ha. punctata, and a nymph of Ixodes ricinus were found. Among R. turanicus, two distinct morphotypes were observed, but they do not form a monophyletic clade in the phylogenetic tree based on the mitochondrial gene cox1. We found a positive correlation between the total number of ticks on nestlings from a particular nest and the number of medium-sized to large prey mammals brought to the nestling owls. Also, the most important predictor for tick abundance was the effect of the extent of arable land (negative), while forests and grasslands contributed less, with no effect observed in case of urbanized areas and watercourses. CONCLUSIONS: The intensity of tick infestation can be high on nestling Eurasian eagle owls (mean intensity 16.59 ticks/nestling). In this study, five different tick species were recorded, among which R. turanicus dominated. Two male morphotypes of this tick species were found, but their morphological differences were not reflected by genetic diversity or phylogenetic clustering. The most important factor determining tick abundance was the land-use structure.

Pictorial dissection guide and internal anatomy of the cattle tick, Rhipicephalus (Boophilus) microplus (Canestrini).

Ticks are pests and vectors of diseases that are of public health and veterinary importance. The cattle tick, Rhipicephalus microplus (Canestrini, 1888), is one of the most studied tick species because of its impact on livestock health and production in the tropical and subtropical parts of the world, costing the cattle industry billions annually. Control methods have evolved throughout the years but so has R. microplus. Reliance upon chemical control has created a consistent need to develop new technologies to overcome the pesticide resistance that occurs as the ticks adapt. In order to utilize the more advanced tools such as RNAi or Crispr/Cas9 systems, tick tissues need to be isolated and manipulated. Unfortunately, there are a limited number of dissection guides available providing a detailed view of tick internal anatomy. This manual includes photomicrographs to guide the dissection of R. microplus adults, male and female. Topography and anatomical differences between the internal organs of unfed and gravid adult females are described. We were able to locate the crucial tissues for cattle tick physiology and lay out spatial and temporal guidelines for their identification and dissection. Examples of how this information can be used at the nexus between organismal and molecular research to innovate tick control technologies is discussed.

Invasive cattle ticks in East Africa: morphological and molecular confirmation of the presence of Rhipicephalus microplus in south-eastern Uganda.

BACKGROUND: Rhipicephalus microplus, an invasive tick species of Asian origin and the main vector of Babesia species, is considered one of the most widespread ectoparasites of livestock. The tick has spread from its native habitats on translocated livestock to large parts of the tropical world, where it has replaced some of the local populations of Rhipicephalus decoloratus ticks. Although the tick was reported in Uganda 70 years ago, it has not been found in any subsequent surveys. This study was carried out to update the national tick species distribution on livestock in Uganda as a basis for tick and tick-borne disease control, with particular reference to R. microplus. METHODS: The study was carried out in Kadungulu, Serere district, south-eastern Uganda, which is dominated by small scale livestock producers. All the ticks collected from 240 cattle from six villages were identified microscopically. Five R. microplus specimens were further processed for phylogenetic analysis and species confirmation. RESULTS: The predominant tick species found on cattle was Rhipicephalus appendiculatus (86.9 %; n = 16,509). Other species found were Amblyomma variegatum (7.2 %; n = 1377), Rhipicephalus evertsi (2.3 %; n = 434) and R. microplus (3.6 %; n = 687). Phylogenetic analysis of the 12S rRNA, 16S rRNA and ITS2 gene sequences of R. microplus confirmed the morphological identification. CONCLUSIONS: It is concluded that R. microplus has replaced R. decoloratus in the sampled villages in Kadungulu sub-county, since the latter was not any longer found in this area. There is currently no livestock movement policy in force in Uganda, which could possibly limit the further spread of R. microplus ticks. Future surveys, but also retrospective surveys of museum specimens, will reveal the extent of distribution of R. microplus in Uganda and also for how long this tick has been present on livestock without being noticed.

Morphometric similarity among populations of Rhipicephalus microplus (Acari: Ixodidae) in the north-western region of Colombia.

Rhipicephalus microplus is one of the most widely distributed and economically important ticks able to transmit pathogens to vertebrate hosts which cause major constraints to public and livestock health. A better understanding of their population structure is crucial for the development of more effective control strategies. Modern morphometrics has enabled the quantification of size and shape variations to investigate population differences in anatomical structures. We therefore applied landmark-based and outline-based geometric morphometric approaches to study 85 field-collected adult females of three R. microplus Colombian populations along a distance of 28-67 km using the scutum and the basis capituli. Although size differences were found between some populations, mean shape and metric disparity of the analyzed structures were very similar, resulting in low classification scores (<41%). Thus, both structures support a metapopulation of R. microplus at a microgeographic level. The variance of shape between structures does not co-vary, likely indicating morphological modularity. These data provide first clues to understand the metric variation of R. microplus among natural populations from north-western Colombia.

Mitochondrial and nuclear multilocus phylogeny of Rhipicephalus ticks from Kenya.

The morphological diversity of African ticks of the genus Rhipicephalus and subgenus Boophilus have been studied in detail. However, their taxonomy remains poorly resolved with limited molecular studies performed to improve inter-species discrimination. Herein, ribosomal cytochrome c oxidase I (COI), 12S ribosomal DNA (12S rDNA) and nuclear ribosomal DNA internal transcriber spacer 2 (ITS2) were analyzed in Rhipicephalus tick populations in Kenya. While the morphological and molecular criteria separated R. e. evertsi, R. pulchellus and R. appendiculatus from other members of the genus, except the morphologically similar sibling species R. zambeziensis, this was not the case for other tick populations. COI sequences of Rhipicephalus ticks from Ruma National Park (RNP) in Southwestern Kenya, that were morphologically similar to R. praetextatus/R. simus, a formed distinct clade and barcode gap group. 12S rDNA haplotypes of this population were 99% identical to a GenBank accession of R. muhsamae which is thought to be endemic in West and Central Africa. However, the ITS2 locus indicated that the RNP samples were genetically closest to ticks identified morphologically as R. praetextatus. The COI and 12S rDNA haplotype sequences of R. praetextatus clustered closely with R. simus reference sequences though the two species occurred in distinct barcode gap groups. Our results suggest that the R. simus/R. praetextatus/R. muhsamae comprise a closely related tick species complex found across sub-Saharan Africa and includes the yet to be described RNP population. More studies on the biology, ecology and genomics of all life stages of tick species in the complex may clarify their taxonomic status. A continent-wide study that combines morphology, DNA marker sequencing and emerging methods, such as mass spectrometry and whole-genome resequencing may reveal the diversity and distribution of taxa within the genus Rhipicephalus in sub-Saharan Africa.

Molecular, histological and ultrastructural characterization of cytotoxic effects of amitraz on the ovaries of engorged females of Rhipicephalus (Boophilus) annulatus.

In the present study, the cytotoxic effects of amitraz, an octopamine receptor agonist on the reproductive system of engorged adult females of Rhipicephalus (Boophilus) annulatus were assessed using histology, electron microscopy and octopamine beta (OCTß) receptor transcriptional expression analysis. Adult immersion test (AIT) was performed by immersing the fully engorged female ticks for 2 min in different concentrations of amitraz (200, 250, 300, 350 ppm). Amitraz at the dose of 300 ppm, caused an adult tick mortality of 16.66 ±â€¯6.80 per cent, inhibition of fecundity of 75.80 per cent and hatching of 50 per cent of ova laid by treated ticks. Histological changes in the ovaries of ticks collected after 24 h of treatment with amitraz (300 ppm), in comparison with controls (distilled water/methanol) were identified by microscopical examination of sections (4  µm) stained using haematoxylin and eosin. These changes included reduction in size and basophilia of stage I oocytes, presence of cytoplasmic vacuoles of various sizes around germinal vesicle of stage II oocytes, wavy basement membrane of stage III oocytes and reduction in size and number of mature stage IV and V oocytes. Electron microscopy was employed for understanding the structural changes in the ultrathin sections (60 nm) of ovaries. Ticks treated with amitraz showed major ultrastructural changes such as irregular nuclear membrane, crystolysis of mitochondria and detachment of external and internal layers of basal lamina of oocytes. The cDNA synthesized from the total RNA of whole ticks and ovaries of ticks treated with amitraz along with controls were used for relative quantification of Octopamine ß receptor (OCTß-R) expression based on the 2-ΔΔCT method by quantitative real time PCR (qRT PCR). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as endogenous control. Down regulation of expression of OCTß-R mRNA in the ovaries of amitraz treated ticks was observed compared to controls. Thus, the inhibition of fecundity observed in the ticks treated with amitraz can be attributed to the major structural changes and decreased expression of OCT ß receptor mRNA induced by it in the ovary.

General and Local Morphological Anomalies in Amblyomma lepidum (Acari: Ixodidae) and Rhipicephalus decoloratus Infesting Cattle in Uganda.

Morphological abnormalities in ticks seem to be rare phenomena in nature, and are underreported in Africa. In this article, we describe general and local anomalies in two Amblyomma lepidum females and one Rhipicephalus decoloratus female collected from cattle in Moroto and Kasese districts, Uganda. One A. lepidum specimen displayed metagynander gynandromorphism with the presence of both male and female features in the same organism. The second A. lepidum female showed slight asymmetry and lacked a genital aperture. The R. decoloratus displayed multiple anomalies that included asymmetry on the right side in association with ectromely, chitinous formations and constrictions on the left side. This article presents the first report of metagynander gynandromorphism, as well as genital aperture absence which is not linked to gynandromorphism, in A. lepidum collected from cattle.

Phylogenetic insights on Mediterranean and Afrotropical Rhipicephalus species (Acari: Ixodida) based on mitochondrial DNA.

A multigene phylogeny including 24 Rhipicephalus species from the Afrotropical and Mediterranean regions, based on mitochondrial DNA genes (COI, 12S and 16S), was constructed based on Bayesian inference and maximum likelihood estimations. The phylogenetic reconstruction revealed 31 Rhipicephalus clades, which include the first molecular records of Rhipicephalus duttoni (Neumann), and Rhipicephalus senegalensis (Koch). Our results support the R. pulchellus, R. evertsi and R. pravus complexes as more phylogenetically close to Rhipicephalus (Boophilus) than to the remaining Rhipicephalus clades, suggesting two main monophyletic groups within the genus. Additionally, the phenotypic resembling R. sanguineus s.l. and Rhipicephalus turanicus (Pomerantsev) are here represented by nine clades, of which none of the R. turanicus assemblages appeared as distributed in the Iberian Peninsula. These results not only indicate that both species include more cryptic diversity than the already reported, but also suggest that R. turanicus distribution is less extended than previously anticipated. This analysis allowed to improve species identification by exposing cryptic species and reinforced mtDNA markers suitability for intra/inter-species clarification analyses. Incorporating new species molecular records to improve phylogenetic clarification can significantly improve ticks' identification methods which will have epidemiologic implications on public health.

Morphological and phylogenetic analyses of Rhipicephalus microplus ticks from Bangladesh, Pakistan and Myanmar.

Ticks of the Boophilus subgenus, classified in the genus Rhipicephalus, are widespread in subtropical and tropical regions, but knowledge on their distribution in Asia is fragmentary. The most important representatives belong to the Rhipicephalus (Boophilus) microplus complex, which is composed of species that share many morphological similarities and are therefore difficult to distinguish. In this study, adult Rhipicephalus (Boophilus) ticks from Bangladesh, Myanmar and Pakistan were subjected to phylogenetic and Scanning Electron Microscopy (SEM) analyses. Phylogenetic analyses of cytochrome oxidase 1 (cox1), internal transcribed spacer 2 (ITS2) and 12S rRNA gene sequences confirmed that the R. microplus complex consists of at least five taxa: R. annulatus, R. australis, and R. microplus clades A-C. Ticks from Bangladesh, Myanmar and Pakistan were assigned to R. microplus clade C. SEM images revealed a wide range of variation in the morphology of the clade C adults, including morphological features previously identified as critical for distinguishing R. microplus from R. australis, which is illustrative for the complications in identifying species within the R. microplus complex using morphology only. Further morphological, genomic and crossbreeding studies are required to elucidate the species status of R. microplus clades A-C.

A new lipid carrier protein in the cattle tick Rhipicephalus microplus.

Tick infestation in cattle reflects the main cause of economic loss to cattle producers. This is due to several reasons but mainly to their ability to feed on blood and generate a huge amount of eggs. Lipid transport in arthropods is achieved by highly specialized hemolymphatic lipoproteins, which resemble those described in vertebrate blood. Such lipoproteins continuously deliver lipids through the blood to growing eggs. The injection of radioactive [3H] palmitic acid into tick hemocoel showed that the gut, ovary, fat body and Gene's organ were the main organs of incorporation of this labeled fatty acid. The rate of [3H] palmitic acid incorporation by the organs was high up to 30 min after injection. The [3H] palmitic acid incorporated by these organs was later found in phospholipids and neutral lipids. Here, we describe the purification and characterization of a key player of lipid dynamics in tick hemolymph. The Rhipicephalus microplus lipid-apolipoprotein complex (RmLCP) is a new high-density lipoprotein (1.18 g/mL), which accounts for over 90% of [3H] palmitic acid present in the hemolymph. It has a native molecular weight of 420 kDa and is composed of one subunit of 122 kDa. Protein identification analysis of RmLPC subunit showed two better hits: vitellogenin 2 (23% protein coverage) and vitellogenin 5 (29% protein coverage), respectively and similarities with hemolymphatic apolipoproteins of arachnids such as the tick Ixodes scapularis (80%), the mite Galendromus occidentalis (44%) and the spider Parasteatoda tepidariorum (43%) and also for the insects Locusta migratoria (45%), Drosophila melanogaster (42%) and Manduca sexta (47%) to vitellogenin 2 and tick Ixodes scapularis (83%), the crab Limulus polyphemus (55%) and the oyster Crassostrea gigas (55%) to vitellogenin 5. Furthermore, it shows a distinct lipid composition from most arthropod lipoproteins, being composed of 40% free cholesterol, 27% phospholipids, 20% triacylglycerol and 15% hydrocarbons. In addition to binding most hemolymphatic fatty acids, this lipoprotein also binds and transports free cholesterol. In conclusion, the present study provides insight into the macromolecules involved in arachnid metabolism, which have significant potential for future use for the biological control of ticks.

Morphological and molecular divergence of Rhipicephalus turanicus tick from Albania and China.

Rhipicephalus turanicus is an important tick species potentially carrying tick-borne pathogens. Several tick species have obvious subspecies divergence. However few studies aimed to examine the existence of divergence within R. turanicus. Therefore, a detailed morphological and molecular analysis was conducted for comparing R. turanicus from the Mediterranean Basin (represented by Albania) and Central Asia (Northwestern China). Altogether 315 adult ticks of R. turanicus (103 from Albania and 212 from China) were morphologically and molecularly analysed. DNA samples were used for mitochondrial 16S rRNA and cox1 gene sequences analysis. In addition, as potentially genetic markers, three fragments including partial nad1-16S rRNA, nad2-cox1, cox1-tRNA-Lys, were designed and then phylogenetically analyzed. Based on detailed morphological observations, only basis capituli length:width ratio (females), the length, the width and the length:width ratio of the scutum (males) had differences between R. turanicus from China and Albania. Gene divergences of 16S rRNA, cox1, partial nad1-16S rRNA, nad2-cox1 and cox1-tRNA-Lys from China and Albania ticks were 3.53-4.84, 3.57-4.92, 3.57-4.07, 3.57-4.39 and 3.18-4.69%, respectively. The evaluated five genetic markers revealed two phylogenetic branches in R. turanicus. Obvious differences exist within R. turanicus based on morphological and genetic analysis. Three newly designed genetic markers (partial nad1-16S rRNA, nad2-cox1 and cox1-tRNA-Lys) in this study may be suitable genetic tools for identification and analysis in R. turanicus. Subspecies analysis of R. turanicus from other regions of the world should be initiated in the future.

Morphological and molecular identification of Rhipicephalus (Boophilus) microplus in Nigeria, West Africa: a threat to livestock health.

The cattle tick Rhipicephalus (Boophilus) microplus was first reported in West Africa in Ivory Coast, in 2007. Since then it has made an aggressive eastward advancement having been reported in four other West African countries: Mali, Burkina Faso, Togo and Benin. We herein report the first molecular identification of this tick species in Nigeria, West Africa, and highlight the threat it poses to livestock health. A nation-wide tick survey was conducted in 12 out of 36 states across the various agro ecological zones of Nigeria over a 1 year period (April 2014-March 2015). In total 1498 ticks belonging to three genera collected from cattle were morphologically identified. Overall, Amblyomma species constituted the highest percentage of sampled ticks, 50.2% (752/1498), followed by Rhipicephalus (including the subgenus Boophilus) species, 29.4% (440/1498) and Hyalomma species, 20.4% (306/1498). The presence of Rh. (B.) microplus was identified morphologically from four out of the 12 states. This finding was confirmed for the first time in Nigeria using a molecular method targeting the ITS-2 region of the ticks in three of the 12 states. This study ascertained the presence of Rh. (B.) microplus in Nigeria in addition to a broad variety of cattle tick species, most of which are of veterinary importance. The implication of this finding is that there may be additional economic burden to livestock farmers due to increased cost of tick control occasioned by the acaricide resistance by this tick species widely reported from different climes. Additionally, there may be a potential upsurge in incidence of hemoparasitic infections in cattle leading to increased morbidity, cost of treatment and mortalities.

Invitro acaricidal activity of ethnoveterinary plants and green synthesis of zinc oxide nanoparticles against Rhipicephalus (Boophilus) microplus.

The present study was designed to investigate the invitro acaricidal effects of seven ethnoveterinary plants, zinc acetate and green synthesized zinc oxide nanoparticles against the Southern cattle tick, Rhipicephalus (Boophilus) microplus. The selected ethnoveterinary plants were extracted using ethanol and aqueous (water) solvents at 0.02mg/ml and 0.04mg/ml concentrations. Of these seven plants, Lobelia leschenaultiana showed the highest percentage of tick mortality. The ethanol extracts of L. leschenaultiana showed 93.33% mortality at 0.04mg/ml and its LC50 was 0.05mg/ml. However, zinc acetate exhibited 70% mortality at 0.04mg/ml (LC50: 0.0192mg/ml). Further, we synthesized ZnO nanoparticle using the leaf extracts of L. leschenaultiana and zinc acetate as the precursor material to control R. (B.) microplus. The structural characterization of the synthesized ZnO nanoparticles (Ll-ZnO NPs) was performed by UV-vis spectroscopy, X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, scanning electron microscopy (SEM) and transmission electron microsopy (TEM). UV-vis spectra showed the absorption band at 383nm. XRD analysis clearly showed the crystalline nature of Ll-ZnO NPs with various Bragg's reflection peaks at 100, 002, 101, 102, 110, 103, 200, 201 and 202 planes. FTIR analysis showed the possible functional groups of Ll-ZnO NPs with strong band at 3420.63 and 2922.48cm(-1). SEM and TEM analysis revealed that the Ll-ZnO NPs were spherical and hexagonal in shape with particle size ranging between 20 and 65nm. The mortality of R. (B.) microplus after treatment with Ll-ZnO NPs was 35, 57.5 and 82.5% at 0.001, 0.002 and 0.004mg/ml. On the otherhand, 100% mortality of R. (B.) microplus was observed at 0.008mg/ml (LC50: 0.0017mg/ml). The results indicated that the Ll-ZnO NPs have good acaricidal properties compared to L. leschenaultiana leaf extract and zinc acetate.

Histoarchitecture of the ovary of Rhipicephalus (Boophilus) annulatus during pre- and postengorgement period.

The present communication describes the detailed day wise study of histological changes of the ovary of Rhipicephalus (Boophilus) annulatus in the postengorgement period together with the systematic classification of their oocytes. The ovary of R. (B.) annulatus is panoistic type with an asynchronous development of oocytes. All the stages (II, III, IV, and V) of oocytes except stage I were similar to R. (B.) microplus. The stage I oocytes showed basophilia, which was not reported earlier in other species of ticks. Day wise changes were in the form of presence of oogonia in partially fed and day one engorged adults, considerable degeneration of oocytes on day two, emergence of new wave of oocytes on day three, presence of mature oocytes up to day eight, and complete degeneration of ovarian tissue from day eight onwards. The degenerative changes in the ovary appeared initially in the oocytes followed by germinal epithelium.

A list of the 70 species of Australian ticks; diagnostic guides to and species accounts of Ixodes holocyclus (paralysis tick), Ixodes cornuatus (southern paralysis tick) and Rhipicephalus australis (Australian cattle tick); and consideration of the place of Australia in the evolution of ticks with comments on four controversial ideas.

Seventy species of ticks are known from Australia: 14 soft ticks (family Argasidae) and 56 hard ticks (family Ixodidae). Sixteen of the 70 ticks in Australia may feed on humans and domestic animals (Barker and Walker 2014). The other 54 species of ticks in Australia feed only on wild mammals, reptiles and birds. At least 12 of the species of ticks in Australian also occur in Papua New Guinea. We use an image-matching system much like the image-matching systems of field guides to birds and flowers to identify Ixodes holocyclus (paralysis tick), Ixodes cornuatus (southern paralysis tick) and Rhipicephalus (Boophilus) australis (Australian cattle tick). Our species accounts have reviews of the literature on I. holocyclus (paralysis tick) from the first paper on the biology of an Australian tick by Bancroft (1884), on paralysis of dogs by I. holocyclus, to papers published recently, and of I. cornuatus (southern paralysis tick) and Rhipicephalus (Boophilus) australis (Australian cattle tick). We comment on four controversial questions in the evolutionary biology of ticks: (i) were labyrinthodont amphibians in Australia in the Devonian the first hosts of soft, hard and nuttalliellid ticks?; (ii) are the nuttalliellid ticks the sister-group to the hard ticks or the soft ticks?; (iii) is Nuttalliella namaqua the missing link between the soft and hard ticks?; and (iv) the evidence for a lineage of large bodied parasitiform mites (ticks plus the holothyrid mites plus the opiliocarid mites).

Ovary histology and quantification of hemolymph proteins of Rhipicephalus (Boophilus) microplus treated with Melia azedarach.

This study aimed to analyze ovary histology and quantify total protein in the hemolymph of Rhipicephalus (Boophilus) microplus females treated with hexane extracts from green fruits of Melia azedarach. Eight engorged females were immersed in the extract at 0.25% concentration, and eight in water containing 5% acetone (control). The females were dissected 72 hours after treatment, and the ovaries were weighed and subjected to standard histological techniques. The total protein concentration was measured in the hemolymph of 200 females, of which 100 were treated as described above and 100 served as a control. In the treated group, ovary weight reduction and predominance of immature oocytes were observed. In addition, there were decreases in the diameters of the cytoplasm and germ vesicle of the oocytes in the treated group, compared with the controls. The protein concentration in the hemolymph was higher in the treated group than in the controls. The morphological changes observed in the treated ovaries included: presence of vacuolization; alteration of oocyte morphology, which changed from rounded to elongated; deformation of the chorion; and disorganization of the yolk granules. These results demonstrate the action of M. azedarach fruit extracts on R. (B.) microplus oogenesis.

Ovary histology and quantification of hemolymph proteins of Rhipicephalus (Boophilus) microplus treated with Melia azedarach / Histologia do ovário e quantificação de proteínas na hemolinfa de Rhipicephalus (Boophilus) microplus tratado com Melia azedarach

This study aimed to analyze ovary histology and quantify total protein in the hemolymph of Rhipicephalus (Boophilus) microplus females treated with hexane extracts from green fruits of Melia azedarach. Eight engorged females were immersed in the extract at 0.25% concentration, and eight in water containing 5% acetone (control). The females were dissected 72 hours after treatment, and the ovaries were weighed and subjected to standard histological techniques. The total protein concentration was measured in the hemolymph of 200 females, of which 100 were treated as described above and 100 served as a control. In the treated group, ovary weight reduction and predominance of immature oocytes were observed. In addition, there were decreases in the diameters of the cytoplasm and germ vesicle of the oocytes in the treated group, compared with the controls. The protein concentration in the hemolymph was higher in the treated group than in the controls. The morphological changes observed in the treated ovaries included: presence of vacuolization; alteration of oocyte morphology, which changed from rounded to elongated; deformation of the chorion; and disorganization of the yolk granules. These results demonstrate the action of M. azedarach fruit extracts on R. (B.) microplus oogenesis.

Este estudo foi desenvolvido, visando analisar a histologia do ovário e quantificar as proteínas totais na hemolinfa de fêmeas de Rhipicephalus (Boophilus) microplus tratadas com extrato hexânico de frutos verdes de Melia azedarach. Oito fêmeas ingurgitadas foram tratadas por imersão com o extrato na concentração de 0,25%, e oito com água contendo 5% de acetona (controle). As fêmeas foram dissecadas 72 horas após o tratamento e os ovários foram pesados e submetidos a técnicas histológicas padrões. A concentração total de proteína foi mensurada na hemolinfa de 200 fêmeas, sendo 100 tratadas como descrito anteriormente e 100 como controle. Foi observada redução do peso dos ovários, predomínio de ovócitos imaturos e houve diminuição nos diâmetros do citoplasma e da vesícula germinal dos ovócitos do grupo tratado em comparação ao controle. A concentração de proteína na hemolinfa foi mais alta no grupo tratado que no controle. As alterações morfológicas observadas nos ovários tratados foram a presença de vacuolizações, alteração da morfologia dos ovócitos que mudaram de redondos para alongados, deformação do córion e desorganização dos grânulos de vitelo. Estes resultados demonstram a ação do extrato de M. azedarach na ovogênese de R. (B.) microplus.