Cell death during preoviposition period in Boophilus microplus tick

Programmed cell death (PCD) is present during the development of multicellular organisms and occurs from embryogenesis to death. In females of Boophilus microplus, the mass of several organs is reduced after the detachment from the host. In order to better characterize the cell death process that eliminates unnecessary tissues, the degeneration of salivary glands, ovaries and synganglia was investigated using DNA fragmentation in agarose gel, comet and TUNEL assays, and apoptosis activation pathway by the caspase assay. DNA fragmentation and enzymatic activity of caspase-3 were observed in salivary glands and ovaries at 48 and 72h after tick removal from the host; in synganglia these parameters were maintained at low levels upon 48h. The results obtained suggest that there is a refined control of tissue maintenance through apoptosis.     

一株白僵菌的分离鉴定及其对微小牛蜱的致病力试验

为了得到用于防控微小牛蜱的高毒力菌株,本试验从野外微小牛蜱饱血雌蜱体表分离出一株白僵菌,通过形态学和分子生物学方法进行了鉴定,确认该分离株为球孢白僵菌.致病力研究表明,当孢子浓度达到108孢子/mL,该菌株对微小牛蜱饱血成蜱的致死率即可达到100%,为微小牛蜱的生物防控试剂的研究及应用奠定了基础.     

Tests to determine LC50 and discriminating doses for macrocyclic lactones against the cattle tick, Boophilus microplus

Laboratory tests were carried out on larvae and adults of the cattle tick Boophilus microplus to determine the toxicity of macrocyclic lactone acaricides (MLs). Technical and commercial MLs were used in larval packet test (LPT), larval immersion test (LIT) and adult immersion test (AIT). In LIT and AIT the toxicity of MLs was much higher than for LPT. In the AIT, diluting the injectable formulation of MLs in water was as effective as dilution in ethanol+Triton X-100. LC50, LC99.9 and 95% confidence limits were determined so that a discriminating dose (DD) could be set for larval and adult tests in order to diagnose potential resistance to MLs in field samples of the tick. These DDs are for Australian strains of B. microplus and may not be suitable for other strains until further work is carried out.The value of these diagnostic tests can only be verified if or when resistance to MLs emerges in ticks.     

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.     

Resistance of Holstein-Friesian cows to infestation by the cattle tick (Boophilus microplus)

In two experiments, the milk production of 59 Holstein-Friesian cows in mid lactation was measured over 1 week before artificial infestation with 2,500 or 5,000 Boophilus microplus larvae. Host resistance, the proportion of female ticks applied but not engorging was estimated from weekly counts of engorging female ticks of 4.5-8. 0mm long. Mean host resistance was 79 and 67% in two experiments. Host resistance was not significantly related to milk yield before infestation, to early pregnancy, or to parity. Culling the 10% of cattle with the least resistance to ticks would result in removal of 19-21% of ticks in a herd. The results suggest that selection to improve the existing low levels of resistance to B. microplus might be undertaken without compromising milk production. However, low levels of resistance among the cattle studied and difficulties in assessment of resistance are likely to limit the usefulness of selection within the Holstein-Friesian breed.     

Synthetic pyrethroid resistance in field samples in the cattle tick (Boophilus microplus)

First evidence of resistance to synthetic pyrethroids in the cattle tick in Australia, other than the pre-existing cross-resistance in DDT-resistant ticks, has emerged. Three new strains have been investigated and characterised. The first, detected in central Queensland, showed increased ability to detoxify pyrethroids resulting in a marginal loss of efficacy of all pyrethroid tickicides tested, except flumethrin. Subsequently two more strains have been detected, with serious implications for chemical tick control. The first, from southern Queensland, has a high level of specific resistance to flumethrin. The second, from central Queensland, has a broad spectrum resistance encompassing all pyrethroids currently in use. The toxicology and biochemistry of these new resistant strains have been investigated and alternative tickicides evaluated. The significance of these latest developments is discussed.     

Progress in the epidemiology and diagnosis of amitraz resistance in the cattle tick Boophilus microplus

Amitraz is a rapidly acting acaricide that has been in use for the control of cattle ticks for more than 30 years. Resistance against amitraz was first reported in Boophilus microplus in Australia in 1980 but has been slow to spread in comparison to resistance against synthetic pyrethroids. The most recent estimate of prevalence of amitraz resistance in Australia is 10.8%. In Mexico, the development and distribution of amitraz appears to have been more rapid and the prevalence has been estimated to be 19.4% in Yucatan state. In New Caledonia, about 10% of properties were confirmed to have amitraz resistance. There is little reliable information on the prevalence of amitraz resistance in southern Africa. Risk factors have been identified, but the small sample sizes in the studies that have attempted to identify risk factors using survey data suggest caution in their interpretation. Regional variation in prevalence has been reported, as has a positive relationship with frequency of acaricide application. There is evidence to suggest that in Australia, amitraz resistance might have emerged on a small number of properties and been disseminated by cattle movements. There is also some evidence to suggest that amitraz resistance can diminish in the field when selection pressure is not applied. The mode of inheritance of amitraz resistance is uncertain and it has been suggested that it is a polygenic trait. The mechanism of amitraz resistance is unknown. Two possibilities have been proposed: octopamine receptor and monoamine oxidase. There is some equivocal support for both possibilities. The larval packet test bioassay is the most reliable method of diagnosing amitraz resistance in B. microplus, and this test has been modified by Miller to provide more accuracy and repeatability. Molecular tests are in development but will not eliminate the need for the bioassay.     

Effect of moxidectin against natural infestation of the cattle tick Boophilus microplus (Acarina: Ixodidae) in the Mexican tropics

The study was divided in to two trials and carried out in a ranch in eastern Yucatan state, Mexico. In the first trial, two groups of 15 BostaurusxBosindicus heifers, 6-12 month of age and naturally infested with Boophilus microplus ticks were used. Heifers in Group 1 were treated with a 1% injectable formulation of moxidectin at the dose of 0.20mg/kg body weight by subcutaneous injection. The other group remained as untreated controls. Number of immature and engorging female ticks were assessed on days 0, 7, 14, 21, 28 and 35 post-treatment (PT). The efficacy of moxidectin on adult ticks from day 7 to 28 PT was greater than 95%. The efficacy decreased to 74.9% by day 35. In the second trial, animals in Group 1 were treated with the moxidectin product as before, while cattle in Group 2 were treated according to the routine procedure for the control of ticks on that property (125 g/l amitraz as a dip). Treatment of all cattle was repeated four times at intervals of 28 days. The efficacy of the experimental moxidectin treatment was similar to that of the routine amitraz treatment, i.e., greater than 99%.     

In vitro acaricidal effect of tannin-rich plants against the cattle tick Rhipicephalus (Boophilus) microplus (Acari: Ixodidae)

The objectives of this study were to evaluate the in vitro acaricidal effects of lyophilized extracts of four tannin rich plants (Acacia pennatula, Piscidia piscipula, Leucaena leucocephala and Lysiloma latisiliquum) against diverse stages of Rhipicephalus (Boophilus) microplus, and to asses whether tannins were involved in the acaricidal effect using polyethylene glycol (PEG) to block tannins. Larval immersion (LIT) and adult immersion (AIT) tests were used to evaluate the acaricidal effect of each of the lyophilized extracts against larval and adult stages of R. microplus respectively. Larvae and adult ticks were exposed to increasing concentrations of each plant extract (0, 1200, 2400, 4800, 9600 and 19,200 μg ml(-1)) for 10 min. Larval mortality was recorded at 48 h post-incubation. Adult mortality was recorded daily over 14 days, at which point their reproductive efficiency was evaluated. PEG was added to the extracts to verify whether tannins were involved in the acaricidal effect. The effect on egg laying inhibition and larval mortality was analyzed using the GLM procedure in SAS. A Kruskal-Wallis test was used to assess the effect of PEG on LIT results. Calculation of the lethal concentration 50 (LC50) was performed using a probit analysis. All extracts reduced the viability of R. microplus larval stages (P<0.001), and viability was restored with the addition of PEG suggesting an important role of tannins in the acaricidal effect (P<0.001). The LC50 values of L. latisiliquum and P. piscipula plant extracts were 6.402 and 2.466 μg ml(-1). None of the tannin-rich plant extracts affected adult mortality (P>0.05). Lysiloma latisiliquum extract inhibited egg hatching of R. microplus (P<0.01). Tannin-rich plant extracts from A. pennatula, P. piscipula, L. leucocephala and L. latisiliquum showed potential acaricidal activity. Further in vivo studies are needed to confirm this finding.     

A new in vitro test to evaluate the resistance level against acaricides of the cattle tick, Rhipicephalus (Boophilus) microplus

In this article we present a new bioassay to assess the resistance status of ticks to acaricides. The Larval Tarsal Test (LTT) is a sensitive, highly time-effective in vitro test. It allows the investigation of a large number of compounds and doses on the cattle tick Rhipicephalus (Boophilus) microplus in a short period of time. The ability of the LTT to assess the lethal concentration at 50% mortality (LC(50)) and resistance ratios (RRs) of a susceptible and a resistant R. microplus strain was compared with the FAO-recommended Larval Packet Test (LPT). Representative compounds of the carbamate, organophosphate (OP), synthetic pyrethroid (SP), formamidine (FOR), macrocyclic lactone and pyrazole classes were used for this comparison. The resistance status against OP, SP and FOR of the resistant R. microplus strain was confirmed in vivo. The LTT resulted in resistance ratios comparable to those obtained with the LPT. However, the lethal concentrations were up to 150-fold lower in the LTT than in the LPT. The advantage of the LTT is to simplify the methodology by avoiding the handling of larvae and using multi-well plates. The LTT is therefore a suitable test for the assessment of the level of resistance of R. microplus and is very promising to evaluate the resistance profile of field strains. Additionally, the LTT is also suitable to test other ixodid species.     

Detection of genomic variability in different populations of the cattle tick Boophilus microplus in southern Brazil

DNA from seven isolates of the cattle tick Boophilus microplus was analyzed by restriction fragment length polymorphism (RFLP). Three different cDNA clones, named P-9, P-25 and CP-12, isolated from a B. microplus cDNA library, were used as DNA probes. DNA sequences of P-9 have high similarity to ribosomal genes, whereas P-25 does not show significant homology with known sequences within databases. CP-12 is a cDNA clone encoding a cysteine endopeptidase gene. A limited degree of polymorphism was detected with P-9 and P-25, while CP-12 showed a different pattern of bands for each tick isolate. These findings suggest the existence of a complex genotypic diversity of the tick B. microplus population in endemic regions.     

Attitudes and practices of Queensland dairy farmers to the control of the cattle tick, Boophilus microplus

Objective To determine practices for the control of cattle ticks on dairy farms in Queensland, the attitudes of farmers to tick infestations and to identify opportunities for and barriers against the introduction of non-chemical methods of tick control.
Design A survey of 199 dairy farmers from tick-infested parts of Queensland was undertaken by 20 dairy advisers and stock inspectors from October 1996 to June 1997. The sample was a proportional, random selection of dairy farms from four regions. A personal interview was conducted with each farmer and answers to 134 questions were obtained.
Results and conclusions Most farmers were not concerned by cattle ticks on their own farms, although they believed that ticks are important to the dairy industry. They were most concerned about the development of chemical resistance by cattle ticks. Inadequate facilities and lack of motivation appeared to be the factors most limiting to improving the methods of control. Most farmers claimed to have only small numbers of ticks at worst. Although a control program recom mended by the Queensland Dairyfarmers' Organisation was well regarded by farmers, few had adopted it. Many farmers saw no need to implement a strategic control program.