Interfering with cholesterol metabolism impairs tick embryo development and turns eggs susceptible to bacterial colonization.

Cholesterol is a known precursor of arthropod molecules such as the hormone 20-hydroxyecdysone and the antimicrobial boophiline, a component of tick egg wax coat. Because the cholesterol biosynthetic pathway is absent in ticks, it is necessarily obtained from the blood meal, in a still poorly understood process. In contrast, dietary cholesterol absorption is better studied in insects, and many proteins are involved in its metabolism, including Niemann-Pick C (NPC) transporter and acyl-CoA:cholesterol acyltransferase (ACAT), as well as enzymes to convert between free cholesterol and esterified cholesterol. The present work addresses the hypothesis that tick viability can be impaired by interfering with cholesterol metabolism, proposing this route as a target for novel tick control methods. Two drugs, ezetimibe (NPC inhibitor) and avasimibe (ACAT inhibitor) were added to calf blood and used to artificially feed Rhipicephalus microplus females. Results show that, after ingesting avasimibe, tick reproductive ability and egg development are impaired. Also, eggs laid by females fed with avasimibe did not hatch and were susceptible to Pseudomonas aeruginosa adhesion and biofilm formation in their surfaces. The immunoprotective potential of ACAT against ticks was also accessed using two selected ACAT peptides. Antibodies against these peptides were used to artificially feed female ticks, but no deleterious effects were observed. Taken together, data presented here support the hypothesis that enzymes and other proteins involved in cholesterol metabolism are suitable as targets for tick control methods.

Innovative granular formulation of Metarhizium robertsii microsclerotia and blastospores for cattle tick control.

The tick Rhipicephalus microplus poses a serious threat to the cattle industry, resulting in economic losses aggravated by tick resistance to chemical acaricides. Strains of Metarhizium spp., a well-known group of entomopathogenic fungi, can contribute to managing this ectoparasite. We explored two novel granular, microsclerotia- or blastospores-based formulations of Metarhizium robertsii for R. microplus control under semi-field conditions. Fungal persistence in soil was also observed for 336 days. The experiment used pots of Urochloa decumbens cv. Basilisk grass, treated with 0.25 or 0.5 mg of granular formulation/cm2 (25 or 50 kg/ha) applied to the soil surface prior to transferring engorged tick females onto the treated soil. The fungal granules yielded more conidia with subsequent sporulation under controlled indoor conditions than in the outdoor environment, where the levels of fungus rapidly declined over time. Metarhizium-root colonization ranged from 25 to 66.7% depending on the propagule and rate. Fungal formulations significantly reduced the number of tick larvae during the humid season, reaching at least 64.8% relative efficacy. Microsclerotia or blastospores-granular formulations of M. robertsii can reduce the impact of R. microplus, and thus prove to be a promising tool in the control of ticks.

In vitro and in silico studies of the larvicidal and anticholinesterase activities of berberine and piperine alkaloids on Rhipicephalus microplus.

Rhipicephalus microplus is responsible for high economic losses in livestock and its control has become difficult due to the establishment of tick populations resistant to commercial acaricides. This study aimed to evaluate the in vitro larvicidal effect of the alkaloids berberine and piperine, and also to investigate their inhibitory mechanisms against the acetylcholinesterase enzyme. The effects of the alkaloids on larvae were observed through the immersion test at the following concentrations: 1.5; 3; 6; 12; 16 and 24 mM. Berberine and piperine presented larvicidal activity greater than 95 %, not differing from 100 % for the positive fipronil control (p > 0.05). Of the two alkaloids, piperine had a lower effective concentration (EC), with an EC50 of 6.04 mM. The acetylcholinesterase enzyme used in the study was obtained from R. microplus larvae (RmAChE) and the anticholinesterase activity was determined spectrophotometrically. The highest anticholinesterase activity, measured as inhibition concentration (IC), was observed for berberine (IC50 = 88.13 µM), while piperine showed lower activity (IC50 > 200 µM). Docking studies in RmAChE, followed by 10 ns molecular dynamics simulation, suggest that berberine stabilizes the RmAChE at lower Root-Mean-Square Deviation (RMSD) than Apo protein. Few hydrogen-bond interactions between berberine and RmAChE residues were balanced by hydrophobic and π-type interactions. Berberine fills preferentially the peripheral anionic site (PAS), which correlates with its non-competitive mechanism. These results suggest that berberine and piperine alkaloids have an in vitro acaricidal action on R. microplus larvae, and the likely mechanism of action of berberine is related to RmAChE inhibition when accessing the PAS residues. These findings could help the study of new natural products that could inhibit RmAChE and aid in the development of new acaricides.

An update on amitraz efficacy against Rhipicephalus microplus after 15 years of disuse.

Amitraz is an acaricide that is widely used in veterinary medicine to control the cattle tick Rhipicephalus microplus. However, controversy exists in the literature regarding the resistance of R. microplus to this product. The present work provides an update on the acaricidal efficacy of amitraz (Triatox®, 12.5 % amitraz) after 15 years without its use on a property. Two in vivo (bovines treated with amitraz and submitted to tick counts, n = 20 animals) and one in vitro (adult immersion test, n = 40 ticks) assays were performed to determine product efficacy. The efficacy of the commercial formulation tested in the first in vivo trial ranged from 14.1 to 47.0%, and in the second from 3.6 to 35.1%, for the 28 days of the experiments. Efficacy for the in vitro trial was 47.38%. The dose recommended by the manufacturer of the product did not cause mortality to most of the ticks of this strain, and efficacy/resistance was not reverted or modified after 15 years (estimated 60 tick generations).

A transcriptome and proteome of the tick Rhipicephalus microplus shaped by the genetic composition of its hosts and developmental stage.

The cattle tick, Rhipicephalus microplus, is a monoxenous tick that co-evolved with indicine cattle on the Indian subcontinent. It causes massive damage to livestock worldwide. Cattle breeds present heritable, contrasting phenotypes of tick loads, taurine breeds carrying higher loads of the parasite than indicine breeds. Thus, a useful model is available to analyze mechanisms that determine outcomes of parasitism. We sought to gain insights on these mechanisms and used RNA sequencing and Multidimensional Protein Identification Technology (MudPIT) to generate a transcriptome from whole larvae and salivary glands from nymphs, males and females feeding on genetically susceptible and resistant bovine hosts and their corresponding proteomes. 931,698 reads were annotated into 11,676 coding sequences (CDS), which were manually curated into 116 different protein families. Male ticks presented the most diverse armamentarium of mediators of parasitism. In addition, levels of expression of many genes encoding mediators of parasitism were significantly associated with the level and stage of host immunity and/or were temporally restricted to developmental stages of the tick. These insights should assist in developing novel, sustainable technologies for tick control.

Clinical safety of lauric acid for cattle and its in vitro and in vivo efficacy against Rhipicephalus microplus.

The aim of the present study was to test the in vitro acaricidal activity of saturated fatty acids (hexanoic, octanoic, decanoic, lauric, myristic, palmitic, octadecanoic, eicosanoic, docosanoic and tetracosanoic) against Rhipicephalus microplus and select a candidate compound for the subsequent determination of its clinical safety for mice and bovines as well as its in vivo efficacy (ethical clearance number 507/2013). None of the compounds exhibited in vitro larvicidal effectiveness, but acaricidal effectiveness was greater than 95 % in the adult immersion test at 40 mg/ml (hexanoic, octanoic, decanoic, lauric, myristic, palmitic and eicosanoic acids). After a second AIT evaluation of serial concentrations of the fatty acids, lauric and myristic acids were selected for the safety and in vivo efficacy assays. No adverse effect was found in the local lymph node assay in mice treated with lauric or myristic acid. Moreover, no clinical signs of systemic poisoning or dermatological, hematological or biochemical abnormalities were found in cattle after the topical application of 1 % lauric acid. In the dose determination test, the 1% solution of this compound exhibited 86% efficacy in cattle naturally infested by a field population of Rhipicephalus microplus susceptible to all chemical groups, except synthetic pyrethroids. The efficacy of 1 % lauric acid was 53.4 % in the dose confirmation test performed on another herd with a field R. microplus population resistant to all chemical groups of acaricides. In conclusion, fatty acids are potential bioactive compounds for the control of R. microplus. Topically applied lauric acid (C12) exhibits in vivo acaricide activity against adults, nymphs and larvae of R. (B) microplus and is safe for cattle.

1,8-cineole and castor oil in sodium lauryl ether sulphate disrupt reproduction and ovarian tissue of Rhipicephalus (Boophilus) microplus.

Essential and fixed oils have been researched as alternatives to chemical acaricides. The activity of volatile compounds from essential oils (1,8-cineole, citral and eugenol) at 1.0% (w/v) and fixed oil (castor oil) at 0.3% (w/v) dissolved in 2.0% (v/v) dimethyl sulfoxide (DMSO) + 0.2% (w/v) Tween 80® was assessed against Rhipicephalus microplus using immersion tests. 1,8-cineole (29.0%) and castor oil (30.2%) had the highest reproductive inhibition rate. A second experiment was performed to verify the effect of the 1,8-cineole (10.0% w/v) and, or castor oil (0.3% w/v) on tick reproduction using different solubilizing agents. The highest reproductive inhibition was observed for the combination of 1,8-cineole/castor oil (94.1%) and 1,8-cineole in 2.0% (w/v) sodium lauryl ether sulphate (SLES) (92.8%). A third experiment showed morphological changes in R. microplus oocytes at different stages of development, as well as in pedicel cells. The most intense effects were observed when ticks were immersed in the formulation containing 1,8-cineole (10.0% w/v) and castor oil (0.3% w/v) dissolved in 2% (w/v) SLES. These findings highlight the potential of this formulation as an alternative for managing cattle ticks as their cytotoxic effects can reduce R. microplus reproductive success.

Terpenes on Rhipicephalus (Boophilus) microplus: Acaricidal activity and acetylcholinesterase inhibition.

The Rhipicephalus (Boophilus) microplus tick is the main ectoparasite of cattle in tropical and subtropical regions worldwide. Resistance to chemical acaricides has become widespread affirming the need for new drugs to tick control. Terpenes have become a promising alternative for cattle tick control, however the mechanism of action of these compounds is still controversial. Inhibition of acetylcholinesterase (AChE) is a well established mechanism of action of organophosphate and carbamate acaricides, but the possible action of terpenes on tick AChEs has seldom been studied in resistant and sensitive strains of R. (B.) microplus. The aim of the present study was to evaluate terpene inhibition of AChE from resistant and sensitive strains of R. (B.) microplus in correlation with their acaricidal activity. Among the terpenes used in the present study, p-cymene, thymol, carvacrol, and citral displayed acaricidal activity with LC50 of 1.75, 1.54, 1.41, and 0.38 mg.mL-1 for the susceptible strain, and LC50 of 1.40, 1.81, 1.10, and 1.13 mg.mL-1 for the resistant strain. Thymol and carvacrol inhibited the AChE of the susceptible strain larvae with IC50 of 0.93 and 0.04 mg.mL-1, respectively. The IC50 exhibited by eucalyptol, carvacrol and thymol for AChE of the resistant strain larvae were 0.36, 0.28, and 0.13 mg.mL-1, respectively. This was the first study to investigate the action of terpenes on AChE from susceptible and resistant R. (B.) microplus. As not all terpenes with acaridical activity showed AChE inhibition, the participation of AChE in the acaricidal activity of terpenes needs further investigation.

Multiple acaricide-resistant Rhipicephalus microplus in the semi-arid region of Paraíba State, Brazil.

Rhipicephalus microplus is responsible for major economic losses in cattle ranching in Brazil, and the development of multiple acaricide resistance is a major concern to control this ectoparasite. To determine the resistance of R. microplus to acaricides in 26 ranches in the semi-arid region of Paraíba State, Brazil, larval assays were performed with cypermethrin, chlorpyriphos, amitraz, ivermectin and fipronil and resistance was observed in all locations. The frequency of populations resistant to cypermethrin, chlorpyriphos, amitraz and ivermectin was, respectively, 96 %, 72 %, 83 % and 92 % while none was resistant to fipronil. Resistance to one, two, three and four acaricides was recorded, respectively, in 4 %, 15 %, 31 % and 50 % of the farms. Errors in the control management of R. microplus determined the development of resistance to acaricides in all evaluated properties pointing towards development of new strategies for its control. In order to control multiple acaricide-resistant cattle tick populations in this area it is recommended to apply suppressive treatments with fipronil and/ or fluazuron.

Wild carnivores, domestic dogs and ticks: shared parasitism in the Brazilian Cerrado.

The increase of contact between natural and rural areas is prominent in Brazil, due to agricultural activities and concern with the environmental conservation. In this context, domestic animals, wild fauna and humans are exposed to mutual exchange of parasites, microorganisms and diseases. We studied tick parasitism of wild carnivores and domestic dogs, and the environmental of questing ticks, in extensive cattle ranch areas intermingled with natural vegetation, and in a natural reserve, both in a region of Cerrado biome, Midwestern Brazil. From 2008 to 2015, we inspected 119 wild carnivores from nine species, and collected six tick species (Amblyomma sculptum, Amblyomma ovale, Amblyomma dubitatum, Amblyomma tigrinum, Dermacentor nitens and Rhipicephalus microplus). The most numerous and infested hosts were Cerdocyon thous, Lycalopex vetulus, Chrysocyon brachyurus, Puma concolor and Conepatus amazonicus. From 139 domestic dogs, we collected A. sculptum, Rhipicephalus sanguineus and R. microplus. From vegetation, samplings resulted in A. sculptum, A. dubitatum, A. ovale, Amblyomma rotundatum and R. microplus, with dominance of A. sculptum. Domestics and wild animals presented high overlapping of infestations by A. sculptum, a generalist and anthropophilic tick species. This tick is the most important vector of the Brazilian spotted fever, a lethal human disease. This fact elicits attention and requires efforts to monitor the presence of pathogens vectored by ticks circulating in this type of agroecosystem, including in other regions of the Brazil, because the most of the natural vegetation remaining have been increasingly immersed in pastures and agricultural matrix.

Oogenesis and embryogenesis inhibition induced by two new ethyl-carbamates in the cattle tick Rhipicephalus microplus.

The purpose of this work was to contribute to the understanding of the mechanism of action of two new ixodicides. The histological and ultrastructural alterations of Rhipicephalus microplus oocytes (San Alfonso strain) treated with two new ethyl-carbamates (ethyl-4-bromophenyl carbamate and ethyl-4-chlorophenyl carbamate) by the adult immersion test were evaluated by light microscopy and transmission electron microscopy. The effects of the carbamates on embryogenesis in eggs were evaluated by fluorescence microscopy using DAPI staining. Both ethyl-carbamates inhibited the maturation of most oocytes and induced a concentration-dependent decrease (r2  = 0.5, p < 0.05) in the embryonation percentage in the small number of eggs oviposited by treated ticks. Evident ultrastructural alterations were observed in the oocytes from ticks exposed to the ethyl-carbamates, including modification of the chorion structure, myelinic bodies and autophagic vacuoles that were associated with degenerated organelles (mitochondria, endoplasmic reticulum and yolk granules), nucleolus fragmentation and chromatin clumping in germinal vesicles. In conclusion, these ethyl-carbamates affect the reproductive potential of R. microplus due to their negative effects on oogenesis and their repercussions for embryonic development.

Resistance profile and molecular characterization of pyrethroid resistance in a Rhipicephalus microplus strain from Colombia.

Intensive use of chemical acaricides for the control of cattle ticks (Rhipicephalus microplus) has led to the development of multiple acaricide resistance in Colombia. The present study aimed to characterize, using toxicological bioassays and molecular biology techniques, the resistance profile of a tick strain isolated from the Arauca state, Northeast Colombia. Commercial acaricides were used in adult immersion tests to determine its in vitro efficacies. Deltamethrin showed very low activity (4-7.3%), a mixture of cypermethrin and chlorpyrifos had intermediate efficacy (64-75.2%), and ethion presented the highest activity (88.5-100%). A colony (Arauquita strain) was established and larval immersion tests confirmed high resistance level to deltamethrin (241-fold) and susceptibility to ivermectin. A quantitative polymerase chain reaction-high resolution melt technique was used to identify single nucleotide polymorphisms (SNPs) in the para-sodium channel gene. All of the genotyped individuals were mutant, presenting one (n = 7), two (n = 7) or three (n = 9) SNPs previously associated with pyrethroid resistance. Sequencing revealed a novel mutation (F712L), that was found for the first time in R. microplus ticks from South America. This is the first description of mutations associated with pyrethroid resistance in R. microplus from Colombia. The acaricide resistance pattern found in the Arauquita strain is similar to other parts of Colombia.

Interpretation of the larval immersion test with ivermectin in populations of the cattle tick Rhipicephalus (Boophilus) microplus from Colombian farms.

Interpreting in vitro bioassays used to determine resistance against acaricides in Rhipicephalus microplus can be challenging without parallel in vivo studies that assess for lost efficacy. The larval immersion test (LIT) is currently the most widely used bioassay to detect ivermectin resistance. The objective of this study was to compare results of the LIT and a field trial using ivermectin in naturally infested cattle. Criteria to consider ticks as resistant with the LIT were based on discriminating doses (DD) and the ratio of lethal concentration (LC) in test populations over the LC of the susceptible Deutch strain, known as the resistance ratio (RR). Ticks were collected from 4 farms, two where ivermectin provided good control of tick infestations and two that claimed lack of efficacy. In two farms where administration of a long-acting ivermectin formulation reduced body tick counts to 45 and 25% of the initial counts at 10-days post-treatment, the RR50 and RR99 were approximately 6 and 20, respectively. The LC50 value approximated the DD for the two farms with claimed resistance, suggesting that about half of the population in each farm was resistant. These LIT values are equal to those reported for the most resistant ticks, which supports the use of the LIT to predict control failure in field situations. The two farms where ivermectin provided good control of tick infestations had LC50s similar to the susceptible strain, although for one farm the LC99 and RR99 suggested incipient resistance.

Activation of the ambusher tick Rhipicephalus microplus (Acari: Ixodidae) exposed to different stimuli.

The present study aimed to evaluate the behaviour of larvae of Rhipicephalus microplus exposed to different stimuli. A Y-olfactometer was positioned vertically and R. microplus larvae were exposed to environmental air, CO2 alone, N,N-diethyl-3-methylbenzamide (DEET) alone, and CO2 combined with the repellents DEET and (E)-2-octenal. Tests were also conducted with the olfactometer positioned horizontally; in this case, however, only CO2 was tested. In all tests conducted with the Y-olfactometer positioned vertically, CO2 activated R. microplus larvae even in the presence of DEET and (E)-2-octenal, although activation was lower when these repellents were used. In the absence of CO2 , larval behaviour against DEET was similar to that of the larvae in the control group. In the tests performed with the olfactometer positioned horizontally, the larvae had no significant response to the presence of CO2 . The larvae were not attracted to or repelled by any compound tested in either the vertical or horizontal position of the olfactometer. The lack of horizontal displacement, attraction or repellence may have been a result of the ambush behaviour of this tick species. However, when larvae were exposed to stimuli and the olfactometer was positioned vertically, the interference of attractant and repellent stimuli in larval behaviour was assessed.

Expression dynamics of autophagy-related genes in the cattle tick Rhipicephalus microplus during embryonic development and under increasing larval starvation.

Rhipicephalus microplus is a hematophagous ectoparasite that significantly affects parasitized cattle. As a one-host tick its entire life cycle consists of free-living and parasitic forms. Its extraordinary ability to survive during prolonged off-host periods has been related to the process of cytoplasmic degradation called autophagy. In order to deepen our understanding of this process during R. microplus non-parasitic stages, we determined the expression dynamics of a set of five autophagy-related genes (ATG genes) during embryonic development and over an increasing larval starvation period of 50 days. We found two apparent successive waves of ATG genes transcriptional activation, which paralleled key embryonic changes such as cellularization and organogenesis, as well as nutrient utilization. Moreover, during increasing larval starvation, ATG genes were up-regulated cyclically every 10-15 days. Taken together, our results suggest that autophagy is playing a major role in embryo development and energy metabolism during starvation in R. microplus.

Serologic and molecular survey of Rickettsia spp. in dogs, horses and ticks from the Atlantic rainforest of the state of Bahia, Brazil.

Tick-borne spotted fever in Brazil is known to be caused by two agents, Rickettsia rickettsii and Rickettsia parkeri. Nothing was known about these agents in one area of the Atlantic rainforest biome of Bahia state, where during March to June 2016, 356 rural dogs and 69 horses were sampled and their sera were processed through indirect immunofluorescence assay against antigens of R. rickettsii, R. parkeri, Rickettsia amblyommatis and Rickettsia bellii. Ticks collected from these dogs and horses were molecularly tested for the presence of rickettsial DNA. Overall, 16.4% (58/356) dogs and 24.6% (17/69) horses were seroreactive to Rickettsia spp. Five tick species, Rhipicephalus sanguineus sensu lato (s.l.), Amblyomma ovale, A. sculptum, R. microplus, and A. naponense, were collected from dogs, whereas horses were infested by A. sculptum and Dermacentor nitens. A total of 242 ticks from dogs and 62 from horses were analyzed by PCR targeting rickettsiae, which were detected in only 4/27 (14.8%) A. ovale. Fragments of the rickettsial gltA and ompA genes from these four ticks were 100% identical to the Atlantic rainforest strain of R. parkeri. The presence of A. ovale on dogs was positively associated with local canine seroreactivity to R. parkeri. Our results provide evidence for the transmission of R. parkeri strain Atlantic rainforest from A. ovale to domestic dogs within the rural area of Ilhéus, similarly to other areas of the Atlantic rainforest biome of Brazil, where human cases of R. parkeri-caused spotted fever have been reported.

Evaluation of Tagetes patula (Asteraceae) as an ecological alternative in the search for natural control of the cattle tick Rhipicephalus (Boophilus) microplus (Acari: Ixodidae).

The cattle tick Rhipicephalus microplus is a great sanitary problem and causes huge losses to livestock, being a vector of important diseases. The aim of this work was to verify the action of plant-derived material obtained from Tagetes patula on the life cycle of R. microplus, as well as to analyze the action of these extracts on ovary cells of engorged females. In the Adult Immersion Test, the crude ethanolic extract (TpEtOH) showed an efficacy of 99.2% (LC50 = 18.60 mg mL-1). From the repellency test, it was found that both the TpEtOH and essential oil were 100% efficient on the larval stage. In addition, in the Larval Package Test, a mortality of 98.37% was obtained with TpEtOH (LC50 = 3.798 mg mL-1). From the microscopic analysis of ovary cells, morphological changes in the chorion and cytoplasm of oocytes were observed, with vacuolization around the germinal vesicle and disorganization of the pedicular cells, suggesting an interference in the normal embryogenic formation of the larvae. These results demonstrate that T. patula extracts interfere at all stages of development of the ixodid, from eggs to adults forms, and have a pronounced repellent effect. In addition, the results of the cytotoxicity assays performed on keratinocytes, as well as previous information on oral and dermal acute toxicity (LD50 > 4000 mg kg-1), attest that T. patula can be a safe alternative for ectoparasitic control, representing an alternative for development of a formulation used to help control populations of ticks in the field.

Repellent Effects of Encapsulated Carvacrol on the Rhipicephalus (Boophilus) microplus (Acari: Ixodidae).

Rhipicephalus (Boophilus) microplus (Canestrini) is a problem for livestock production systems, and its control has become challenging due to the selection of tick populations resistant to synthetic chemical acaricides. The use of repellent compounds prevents contact between the arthropod parasite and the host and can, thus, contribute to increases in the efficacy of these acaricides. Carvacrol monoterpenic phenol is a possible alternative method for controlling R. (B.) microplus; however, this compound is highly volatile, and its volatilization can be decreased through microencapsulation, which results in the timed release of the compound. The cell wall of Saccharomyces cerevisiae can be utilized for the protection of volatile molecules. The aim of this study was to evaluate the in vitro repellent effect of yeast cell wall-encapsulated carvacrol on susceptible R. (B.) microplus larvae. Specifically, the vertical filter paper bioassay was employed to analyze the repellent activity of encapsulated carvacrol, nonencapsulated carvacrol, and N,N-diethyl-meta-toluamide at concentrations ranging from 0.75 to 0.001 mg/cm2, and the repellent activities were evaluated. Both carvacrol and encapsulated carvacrol exhibited repellent effects on R. (B.) microplus larvae, and the encapsulated compound showed the highest repellent activities at the lowest concentrations. Carvacrol encapsulated exhibited a low repellent concentration in all times (≤0.05 mg/cm2), whereas the carvacrol nonencapsulated ranged CR50 from 0.13 to 0.27 mg/cm2 at 1- to 6-h posttreatment. The present paper provides the first description of the use of a microencapsulation technique for achieving the highest repellent effect of carvacrol and indicates that this technique might be used to obtain new delivery systems for volatile and hydrophobic compounds.

Antiparasitic activities of hydroethanolic extracts of Ipomoea imperati (Vahl) Griseb. (Convolvulaceae).

Ipomoea imperati is widely used in tropical areas to treat several pathological conditions. The effect of this plant against parasitic species has not been investigated even being used for this purpose in the Brazilian northeastern. This study aimed to evaluate the anthelmintic and acaricide potential of a hydroethanolic extract of I. imperati leaves and stolons. I. imperati leaves and stolons were crushed and subjected to maceration in ethanol 70% (v/v), after which the solvent was removed using a rotary evaporator. The chromatographic profile of the extract was obtained by UV Spectrum high-performance liquid chromatography and compounds were identified by liquid chromatography/electrospray ionization tandem mass spectrometry. Identification of the compounds present in the extract was achieved by comparing their retention times and UV spectra with data in the literature. Anthelmintic activity was evaluated by larval exsheathment inhibition assays using Haemonchus contortus larvae and five concentrations of each extract ranging from 0.07 to 1.2 mg/mL. Acaricide activity was evaluated via larval immersion of Rhipicephalus microplus in eight concentrations of each extract ranging from 5.0 to 25.0 mg/mL. Live and dead larvae were counted after 24 hours. The median inhibitory concentration (IC50) for H. contortus larvae and the median lethal concentration (LC50) for R. microplus larvae were calculated. Twelve compounds were observed in the hydroethanolic extract of leaves, with a predominance of the aglycone form of flavonoids and tannins. This extract was effective against H. contortus larvae, presenting an average inhibitory concentration of 0.22 mg/mL, but showed no activity toward R. microplus larvae. The stolon hydroethanolic extract presented 11 compounds, with phenolic acids and glycosylated flavonoids prevailing. This extract showed low activity on R. microplus and no effect on inhibiting H. contortus larval exsheathment at the concentrations tested. This study is the first to assess the anthelmintic and acaricidal activities of I. imperati. Data reported confirm promising potential of I. imperati leaves hydroethanolic extract against H. contortus. This effect could be due to its secondary compounds presents in this extract, such as procyanidin, kaempferol, isoquercitrin and rutin.

Anatomical distribution and population dynamics of Rhipicephalus (Boophilus) microplus in cattle in the municipality of Óleo, São Paulo / Distribuição anatômica e dinâmica populacional de Rhipicephalus (Boophilus) microplus em bovinos do município de Óleo, São Paulo

Rhipicephalus (Boophilus) microplus infestation is responsible for significant economic losses in cattle industry. The use of acaricides at strategic times and selection of resistant animals can minimize economic losses. This study aimed to determine the anatomical distribution and population dynamics of R. microplus in the municipality of Óleo, state of São Paulo. This study was conducted using eight male cattle, crossbred zebu-taurine (in accordance with the regional herd). Female R. (B.) microplus, greater than 4 mm, present in the right side of the animals were counted between April 2012 and March 2013. The highest tick number per animal was noted in October (128.25), and the lowest in April (35.5), with three infestation peaks during the study, in May, October, and December, which were influenced by the minimum and maximum mean temperature and rainfall. Among the cranio-caudal regions (anterior,median, and posterior), the posterior had the highest mean tick number, and the anterior had the lowest. The minimum temperature, rainfall, and solar radiation interfered with infestation of dorsoventral regions: in cold months, the lowest infestations were found in the mixed area (ear and tail); in the warm months, in the high region (head, neck, upper ribs, flank, and rump). The lower region (dewlap, cranial limb, lower ribs, abdomen, prepuce, scrotum, and lower limb) always had the highest mean tick number.The tick numbers of the caudal limb and crotch showed strong correlation with the total tick number (+ 0.88 and + 0.70, respectively). These data, grouped according to median, posterior, and crotch regions, showed a stronger correlation with the total tick number (+ 0.96, + 0.90, and + 0.70, respectively).(AU)

A infestação pelo Rhipicephalus (Boophilus) microplus é responsável por significativas perdas econômicas na pecuária bovina. O uso de acaricidas de forma estratégica e a seleção de animais mais resistentes podem minimizar os prejuízos econômicos. O objetivo deste estudo foi determinar a distribuição anatômica e a dinâmica populacional do carrapato em bovinos do município de Óleo, SP. O estudo foi conduzido com 08 bovinos machos, mestiços zebu-taurino (conforme rebanho regional). Foram contadas as fêmeas de R. (B.) microplus, maiores que 4 mm, existentes no lado direito do animal, a cada 4 semanas, no período entre abril de 2012 e março de 2013. Obteve-se o maior número de carrapatos por animal em outubro (128,25), e o menor em abril (35,5), com a existência de três picos de infestação, sendo eles nos meses de maio, outubro e dezembro. Verificou-se a existência de três gerações anuais de carrapatos, que foram influenciadas pela precipitação pluviométrica e temperatura média, mínima e máxima. Das regiões Crânio-caudal (anterior, mediana e posterior), a posterior obteve o maior número médio de carrapatos e a anterior o menor. A temperatura mínima, a pluviosidade e a radiação solar interferiram nas infestações das regiões Dorso-ventral, nos meses frios, as menores infestações foram encontradas na área mista (orelha e cauda) e nos meses quentes na alta (cabeça, pescoço, costado alto, flanco e garupa). A região baixa (barbela, membro cranial, costado baixo, ventre,prepúcio, membro caudal e bolsa escrotal) se manteve sempre com a maior média. Das diferentes regiões anatômicas estabelecidas, o número de carrapatos do membro caudal e entrepernas mostraram ter forte correlação com o total, + 0,88 e + 0,70, respectivamente. Esses mesmos dados, agrupados segundo regiões previamente estabelecidas, obtiveram correlação mais forte com o total de carrapatos, sendo + 0,96 (mediana), + 0,90 (posterior) e + 0,70 (entrepernas).(AU)