Rhipicephalus (Boophilus) microplus embryo proteins as target for tick vaccine

Rhipicephalus (Boophilus) microplus is one of the most widely distributed tick in the world. The control of the parasite is based mainly on the use of chemical acaricides, which are produced from a limited set of molecules. These drugs induce selection of acaricide-resistant ticks, and are an important source of environmental pollution. An approach based on anti-tick vaccines may circumvent these obstacles. Characterization of the physiological function of tick molecules may be useful to develop new vaccines. Previously, we reported the ability of some tick proteins as inducers of protective immune response. Vaccination studies using tick proteins like native (nBYC), recombinant (rBYC) egg-yolk aspartic endopeptidase and cysteine endopeptidase (VTDCE) from R. microplus and glutathione S-transferase (Hl-GST) from Haemaphysalis longicornis demonstrated the immunogenicity and antigenicity of these proteins in bovines. Eventually, immunization with these proteins triggered a partial immune response against R. microplus infestation in cattle, manifested mainly as a reduction in egg fertility (7.7% and 13.9% for nBYC, 5.9% for rBYC; 4.7% for VTDCE, 7.9% for Hl-GST), and in the number of fully engorged ticks (18.2% for rBYC, 14.6% for VTDCE, 53% for Hl-GST). The data so far obtained suggest that these proteins have potential to be used as antigens in an anti-tick vaccine. Other proteins involved in tick embryogenesis also have this potential, like THAP and BmCl1, which are enzymes with key roles in vitellin and hemoglobin hydrolysis. Moreover, the identification of analogous proteins present in other tick species may bring information about the way to develop a vaccine against multiple tick species which can help to solve the problem faced by numerous countries where animals are parasitized by more than one tick species. The aim of the present review is to comprehensibly summarize the data obtained in the last few years by our collaborative research, discussing the efforts we have made to find antigens efficient enough for a cattle tick-controlling vaccine. This review discusses tick physiology studies aimed at the selection of possible targets, characterization of the selected proteins with emphasis on their biochemical and immunological aspects and results of vaccine trials on bovines.     

The inability of a South African Babesia bovis vaccine strain to infect Boophilus microplus

A strain of Babesia bovis that had been attenuated by rapid syringe passage through a series of 23 splenectomized calves was unable to infect its vector Boophilus microplus. An attempt to transmit the attenuated Australian Babesia bigemina G strain with a South African strain of B. microplus was likewise unsuccessful. The epidemiological implication of these observations in terms of babesiosis control is discussed.     

Biodegradable alginate microspheres as a delivery system for naked DNA.

Sodium alginate is a naturally occurring polysaccharide that can easily be polymerized into a solid matrix to form microspheres. These biodegradable microspheres were used to encapsulate plasmid DNA containing the bacterial beta-galactosidase (LacZ) gene under the control of either the cytomegalovirus (CMV) immediate-early promoter or the Rous sarcoma virus (RSV) early promoter. Mice inoculated orally with microspheres containing plasmid DNA expressed LacZ in the intestine, spleen and liver. Inoculation of mice with microspheres containing both the plasmid DNA and bovine adenovirus type 3 (BAd3) resulted in a significant increase in LacZ expression compared to those inoculated with microspheres containing only the plasmid DNA. Our results suggest that adenoviruses are capable of augumenting transgene expression by plasmid DNA both in vitro and in vivo.     

Evaluation of tickGARD(PLUS), a novel vaccine against Boophilus microplus, in lactating Holstein-Friesian cows

The effects of vaccination with the Bm 86 vaccine TickGARD(PLUS) against infestation with cattle tick (Boophilus microplus) and of holding cattle on a feedpad until 09:00 hours after the morning milking was tested on 40 mid lactation Holstein cattle using a factorial design. Vaccination resulted in a 56% reduction in tick numbers in the field over one generation, and a 72% reduction in laboratory measures of the reproductive efficiency of ticks. The liveweight gain of vaccinated cattle over 27 weeks was 18.6 kg higher than that of controls, and vaccinated cattle tended to have lower somatic cell count in milk (SCC). There were no other significant differences in measures of production. Cattle kept on the feedpad after the morning milking carried 26% more ticks than those returned immediately to their paddocks.     

Immunization of cattle with synthetic peptides derived from the Boophilus microplus gut protein (Bm86)

Three synthetic peptides (SBm4912, SBm7462 and SBm19733), derived from the Bm86 glycoprotein from Boophilus microplus gut, were constructed and used to immunize cattle from a tick-free area. The immunized animals received three subcutaneous doses of the peptides, with saponin as adjuvant, at 30-day intervals. The immune response was evaluated by IgG elicited against the peptides by the detection of anti-Bm86 specific antibodies in situ and by Western blotting analysis. After tick challenge, reduction in the number, weight and oviposition capacity of engorged females was observed in the tick population that had fed on immunized animals. The results pointed a high efficacy (81.05%) for the SBm7462 synthetic peptide in relation to the others (p<0.01), demonstrating the efficiency of the immune response elicited by synthetic peptides to control the cattle tick B. microplus.     

Lack of infectivity of a Brazilian Anaplasma marginale isolate for Boophilus microplus ticks

Previous studies have shown that one Brazilian Anaplasma marginale isolate presents an inclusion appendage (tail), while other isolates do not present such inclusion. Studies on tick transmission have been carried out with tailless isolates but little is known about transmission of tailed isolates by Boophilus microplus. Two splenectomized calves were experimentally inoculated with the tailed A. marginale isolate. During ascending rickettsemia, B. microplus larvae, free from hemoparasites, were fed on the calves and the resulting nymphs, adult males and engorged females were examined by optic and electronic microscopy. No A. marginale colonies were observed in the gut cells of engorged females and the larvae originated from them did not transmit A. marginale to susceptible calves. In addition, no colonies of A. marginale were seen in the gut cells or in salivary glands of adult males and nymphs. These results suggest that B. microplus is not the biological vector for this tailed isolate.     

Effect of acaricides on the activity of a Boophilus microplus glutathione S-transferase

In the present study, we report the effect of several acaricides on the enzyme activity of a Boophilus microplus recombinant glutathione S-transferase (rGST). GST was expressed in Escherichia coli and was purified with glutathione (GSH) affinity column chromatography. The kinetic constants were determined by reacting GST with the substrates 1-chloro-2,4-dinitrobenzene (CDNB) and glutathione. We report the effect of several acaricides on the enzyme activity of rGST. Some acaricides (ethion, amitraz, chlorpyrifos, DDT, cypermethrin, diazinon, ivermectin, deltamethrin and flumethrin) inhibited rGST. Contrarily, coumaphos had an activating effect. Although the accurate mechanisms of the B. microplus resistance to acaricides remain elusive, this work helps in understanding how acaricides can interact with GST.     

微球制剂在疫苗中的应用及研究进展

疫苗是一种用于预防和治疗传染病或某些慢性疾病的生物制品,实际生产中应用最广泛的是传统灭活疫苗和减毒活疫苗。近20年来,随着生物技术在生物制品领域应用的不断深化和发展,疫苗制造技术和免疫学技术得到迅猛发展,产生了多种新型疫苗,包括基因重组亚单位疫苗、活载体基因工程疫苗、     

Use of the Larval Tarsal Test to determine acaricide resistance in Rhipicephalus (Boophilus) microplus Brazilian field populations

Acaricide resistance of the cattle tick Rhipicephalus (Boophilus) microplus is widespread in most of the countries where this parasite is present. Bioassays are used to diagnose the level and pattern of resistance in tick populations. In the present study, we describe a detailed protocol of the Larval Tarsal Test (LTT) using simplified equipment and data on the resistance of 17 tick field populations originating from 5 Brazilian states. Nine acaricidal compounds from 5 major classes were tested: organophosphates (OP), synthetic pyrethroids (SP), macrocyclic lactones (ML), phenylpyrazols (PYZ) and amidines. For comparison, four of the tick populations were also tested with the Larval Packet Test (LPT) with one compound per class. The most common resistances were to SP, amitraz and OP, with frequencies of 94%, 88% and 82%, respectively. Resistance to PYZ was also found to be widespread (65%), suggesting a rapid development of fipronil resistance in Brazil. One case of ML resistance and 2 cases of suspected ML resistance were identified with the LTT. The LTT led to higher resistance ratios to all compounds than the LPT, reflecting its high sensitivity to detect resistance. Finally, the LTT allowed testing a larger number of compounds and doses with reduced labour in comparison to the LPT and turned out to be a reliable bioassay to detect resistance in field populations.     

Novel amoxicillin nanoparticles formulated as sustained release delivery system for poultry use

Amoxicillin is used in the treatment and prevention of a wide range of diseases in poultry breeding. However, its short half‐life and low bioavailability restrict its clinical application in these species. Entrapment of drugs into polymeric nanoparticles (nps) presents a means to improve gastrointestinal absorption and oral bioavailability of drugs. This study was aimed to overcome limitation of amoxicillin use in poultry breeding. Amoxicillin was loaded into sodium alginate‐polyvinyl alcohol (NaAlg‐PVA ) blend nps, and characterization of the prepared nps was performed. For pharmacokinetic study, commercial male broilers were used and comparative pharmacokinetics of free and nanoparticle form of amoxicillin were investigated. Twenty‐one broilers were divided into three groups. All groups received 10 mg/kg drug. Blood samples were collected, and drug plasma concentrations were determined by HPLC . The results demonstrated that the particle size, zeta potential, encapsulation efficiency, and loading capacity of the nps were 513.96 ± 19.46 nm, ?45.36 ± 1.35 mV , 43.66 ± 3.30, and 12.06 ± 0.83%, respectively. In vitro drug release exhibited a biphasic pattern with an initial burst release of 18% within 2 hr followed by a sustained release over 22 hr. The pharmacokinetic results showed that amoxicillin nps have higher bioavailability and longer plasma half‐life (p  <  .01) than free amoxicillin. These results indicate that amoxicillin nano formulation is suitable for oral administration in broilers.     

Cyhalothrin—a novel acaricidal and insecticidal synthetic pyrethroid for the control of the cattle tick (Boophilus microplus) and the buffalo fly (Haematobia irritans exigua)

SUMMARY Cyhalothrin, a novel synthetic pyrethroid, was evaluated for control of the major resistant strains of the cattle tick (Boophilus microplus) and for control of the buffalo fly (Haematobia irritans exigua) on cattle. In regulated treatment trials with 0.007% cyhalothrin, greater than 99% control of the Biarra, Mackay, Mt Alford, DDT resistant and Ulam cattle tick strains was obtained. Protective-period trials were conducted in which animals which had been sprayed with 0.007% cyhalothrin then received a continuing heavy challenge of the organophosphate-resistant Biarra tick strain. The first semi-engorged adult ticks appeared no earlier than 27 days after treatment, which corresponds to a minimum protective period against reinfestation of 7 days. Protective-periods ranging from 7 to 15 days were obtained in trials that were conducted.     

Compatibility of the fungus Metarhizium anisopliae and deltamethrin to control a resistant strain of Boophilus microplus tick

The tick Boophilus microplus causes economic impact to cattle producers and has a great capacity to develop resistance to chemical acaricides. It is very important the development of new techniques to complement the control of this parasitosis. Biological control is a promising option to maintain acceptable levels of tick populations in cattle. Therefore, the present study evaluated the association of deltamethrin and the entomopathogenic fungus Metarhizium anisopliae against B. microplus larvae resistant to pyrethroid. The synthetic pyrethroid, deltamethrin, was used at concentrations of 0.39, 0.78, 1.56, 3.12 and 6.12 ppm, M. anisopliae was used at concentrations of 10(5), 10(6), 10(7) and 10(8) conidia ml(-1). Their associations were also evaluated in in vitro tests. The assays showed that the tick strain is resistant to deltamethrin, but high mortality rates were observed when deltamethrin was associated with the entomopathogen. The larvae's mortality rates ranged from 7 to 36.5% for the different concentrations of deltamethrin, however, for the different concentrations of M. anisopliae, the mortality rates ranged from 10 to 96.9%. Mortality rates were proportional to the concentrations used for both deltamethrin and the fungus. Predominantly, the association of pyrethroid and fungus resulted in higher larvae mortalility rates than those obtained with the respective non-associated concentrations. Thus, the present study indicates that this association can be used as a tool for integrated control of the tick B. microplus.     

Assessment of a low virulence Australian isolate of Anaplasma marginale for pathogenicity, immunogenicity and transmissibility by Boophilus microplus

A 14-year-old cow (Dawn) born and kept in a Boophilus microplus-free region gave birth to a calf, which showed the presence of an Anaplasma marginale infection after splenectomy. The calf's grand dam was from a B. microplus infected area and we assume the infection originated via the transplacental route over two generations. An isolate, prepared from the calf, had similar or lower pathogenicity as Anaplasma centrale, and previously exposed steers were resistant to challenge by four A. marginale field isolates. Two attempts to transmit the isolate using B. microplus were unsuccessful. Our results indicate that Dawn A. marginale may be a useful vaccine in Australia and warrants larger scale validation of its safety and potency locally as well as of the protection it affords against African and New World isolates.     

Moxidectin pour-on for control of natural populations of the cattle tick Boophilus microplus (Acarina: Ixodidae)

Fifty Bos taurus x Bos indicus heifers naturally infested with Boophilus microplus ticks were divided into two groups of 25 heifers each. Individuals of one group were treated with moxidectin 0.5% pour-on at a dosage of 500 microg of moxidectin/kg body weight and heifers from the other group remained as untreated controls. An efficacy higher than 95% was found on days 7-21 after treatment by using female ticks 4.5-8.0 mm long as the main infestation parameter. A lower, but significant efficacy (p < 0.05) was also found on days 1 (32.3% efficacy) and 27 (73.4% efficacy) post-treatment. Significantly (p < 0.05) lower numbers of immature ticks were also observed on heifers of the treated group from days 7 through 27 after treatment. A lower engorgement weight of female ticks from treated heifers was found on days 1 and 21 after treatment. Treatment also affected reproductive performance (oviposition, egg hatch and number of eggs laid) of female ticks collected on Day 1.     

Effect of different methods of maintenance on the pathogenicity and infectivity of Babesia bigemina for the vector Boophilus microplus

Observations were made on the effects of five different methods of laboratory maintenance on the infectivity and virulence of Babesia bigemina for the tick Boophilus microplus. The original isolate was highly infective and virulent, causing premature death of engorged female ticks and reduced egg production. Maintenance of the strain by syringe passage in unsplenectomised calves at six to 10 week intervals reduced both its infectivity and virulence for ticks. When slow passages were preceded by a series of rapid passages in splenectomised calves, the changes to the strain were less pronounced. The other three procedures, rapid syringe passage in splenectomised calves and tick passage in either splenectomised or intact calves, had no statistically significant effect on the characteristics measured.     

The application of moxidectin formulations for control of the cattle tick (Boophilus microplus) under Queensland field conditions

Objective To assess the control of the cattle tick (Boophilus microplus) and the performance of commercial cattle treated with the macrocylic lactone endectocide, moxidectin, formulated either as an injection or as a pour-on. Design Groups of 10–12 tick infested cattle were treated with moxidectin injection, moxidectin pour-on or remained untreated (28-day trials) or were treated with deltamethrinethion as a dip (140-day trials). The cattle were exposed to natural tick challenge under field conditions. Procedure Tick numbers on trial cattle were recorded in each trial before the initial treatment and in the 28-day trials at 7, 14, 21 and 28 days or in the 140-day trials, at 28-day intervals before each of the treatments and at the final inspection. Body weights of the cattle were also recorded prior to the initial treatment and at the termination of each trial. Cattle were observed on the day of each treatment and at each inspection for evidence of any reactions to treatment. Results 28-day trials: Significant reductions in tick counts were recorded in both treatment groups when compared with cattle in the untreated group. Weight advantage was recorded in the moxidectin treated groups. 140-day trials: All three treatments resulted in zero or low tick counts at each inspection with the exception of the pour-on treatment at week 8 in one trial and week 9 in the other trial. Additional weight gain was recorded for both the moxidectin treated groups, relative to the deltamethrin-ethion dip groups, but was significant only for the pour-on groups. There was no evidence of any local or systemic adverse reaction in any treated cattle in any trial. Conclusion Good to excellent control of the cattle tick (Boophilus microplus) was demonstrated with the moxidectin formulations in all trials, the injection being particularly effective. An improved performance was recorded in all trials in cattle treated with both moxidectin formulations when compared with the untreated cattle and with cattle treated with the deltamethrin-ethion dip. There was no evidence of any local or systemic adverse reaction to treatment with either moxidectin formulation.     

Critical evaluation of the modified-adult immersion test with discriminating dose bioassay for Boophilus microplus using American and Australian isolates

Similar adult immersion tests (AITs) for acaricide susceptibility of Boophilus microplus were done in Texas, USA (Mu?oz strain) and in Queensland, Australia (N-strain and Ultimo isolates). Engorged adult female ticks were immersed in one of a series of dilutions of commercial acaricide in water and then incubated at room temperature for 7 days. Data on oviposition were collected 7 days after exposure to acaricide and subjected to probit analysis. For most data, we observed poor fit to the probit model. Substantial differences in both LC50 and LC99 for the susceptible strains occurred between the respective laboratories and confidence intervals for all acaricides and all strains were unacceptably wide. For amitraz, the discriminating concentration (double the LC99.9 or LC99) recommended by FAO was 0.25%, but our estimates ranged from 0.46% to 9000%. For cypermethrin, the recommended DD was 0.0050%, with our estimates ranging from 0.00022% to 0.74%. For coumaphos the recommended DD was 0.50% but our estimates ranged from 0.66% to 130%. Finally, for moxidectin, the recommended DD was 0.10%, while our estimates ranged from <0.0001% to 5.9%. The method does not provide a means to discriminate between amitraz-susceptible and -resistant, nor between cypermethrin-susceptible and -resistant B. microplus.