O papel das imunidades nas relações parasito-hospedeiro: o carrapato Rhipicephalus (Boophilus ) microplus e bovinos resistentes ou susceptíveis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Aplicação do fungo Metarhizium anisopliae em pastagem visando o controle do carrapato Boophilus microplus em bovinos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Prospecção de biocida em plantas amazônicas e exóticas, visando seu uso racional

O principal método de controle do carrapato bovino Boophilus microplus é o uso de carrapaticidas sintéticos. Entretanto, o rápido desenvolvimento de resistência ao princípio ativo, os riscos de resíduo nos produtos de origem animal e a contaminação ambiental tem incentivado a busca por fontes alternativas de controle desse parasita. Dentro desse contexto, o uso de fitoterápicos aponta como uma alternativa no controle do carrapato, podendo reduzir os impactos econômicos e ambientais ao uso de produtos químicos sintéticos. Assim, o objetivo deste trabalho foi avaliar “in vitro” o efeito dos óleos essenciais de capim-salsar (Cymbopogon interanius Jowitt) e priprioca (Cyperus articulatus L.), substâncias químicas isoladas do timbó (Derris nicou; Derris urucu): rotenona, rotenolona e deguelina e óleos de andiroba (Carapa guianensis Aublet.) e nim (Azadirachta indica A. Juss.), no controle do carrapato. As pesquisas foram desenvolvidas nas instalações da Unidade de Pesquisa Animal “Sen. Álvaro Adolpho”, da Empresa Brasileira de Pesquisa Agropecuária – Embrapa Amazônia Oriental, no município de Belém, estado do Pará, no período de março a julho de 2008. As fêmeas ingurgitadas de Boophilus microplus foram colhidas manualmente, de bovinos naturalmente infestados, em propriedade particular localizada à uma hora de Belém. Após a coleta, foram transportadas para o laboratório em vasilhames plásticos, identificados, limpos e aerados. No laboratório, foram lavadas em água corrente, secas em papel absorvente e pesadas em balança analítica (precisão de 0,0001g). A metodologia utilizada foi a imersão de teleóginas por tempo de cinco minutos. Após a imersão, os carrapatos foram secos em papel absorvente, fixados em placas de Petri e mantidos em temperatura ambiente para realização da postura. Durante todo período experimental a temperatura e umidade relativa do ar em laboratório foi aferida com auxílio de um termohigrometro. O delineamento experimental utilizado foi o inteiramente casualizado. Os percentuais de mortalidade de fêmeas ingurgitadas de B. microplus ,no dia 5 e 15, variaram de: 45% a 90%; 37% a 90% e 60 a 90%; 60% a 90%, para capim-salsar e priprioca, respectivamente. Os resultados referentes a postura não evidenciaram diferença estatística entre as concentrações 2% e 10%, em relação ao controle (água + etanol). Entretanto, houve diferença na concentração 50%, tanto para capim-salsar quanto priprioca. Para capim-salsar, nesta concentração, foi observada postura de ovos inférteis. Com relação aos percentuais de eficiência do produto, os valores obtidos foram de: 100%; 95,63%; 98,77%; 100% e 99,30%; 96,97%; 99,85% e 100%, para os efeitos do óleo essencial de capim-salsar e priprioca nas concentrações 2%, 5%, 10%, 50%, respectivamente. As taxas de mortalidade, no dia 5, variaram de 37% a 41%, 49% a 64% e 52% a 41% para rotenona, rotenolona e deguelina, da menor para maior concentração, respectivamente. Todas as substâncias testadas apresentaram taxa de mortalidade significativamente maior do que as obtidas pelo grupo controle (água + etanol). Os melhores resultados de percentual da postura foram de 49% e 26% para rotenona e deguelina a 200ppm, respectivamente e 34% para rotenolona a 100ppm. Os percentuais de eficiência nos testes com rotenona foram de 95,24% e 98,68%, para rotenolona da ordem de 97,29%; 98,08%; 100% e deguelina 95,45%; 98,23% e 100%, em 100ppm, 200ppm e 300ppm, respectivamente. Para andiroba e nim, o percentual de mortalidade foi observado no dia 15 e demonstrou um comportamento natural do ciclo biológico dos carrapatos, pois não houve diferença estatística significativa entre o controle (água + etanol) e os tratamentos nas diferentes concentrações. Para andiroba e nim a maior eficiência obtida foi de 98,01% e 100%, na concentração de 50%, respectivamente.

Aspectos ecológicos de Boophilus microplus (CANESTRINI, 1887) (ACARINA: IXODIDAE) no município de Franca, nordeste de São Paulo

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Efficacy of fipronil (1.0 mg/kg) against Rhipicephalus (Boophilus) microplus strains resistant to ivermectin (0.63 mg/kg)

The present study aimed to evaluate the efficacy of fipronil (1 mg/kg) against three strains of ivermectin-resistant Rhipicephalus (Boophilus) microplus (R. (B.) microplus), naturally infesting cattle from different states of Brazil. Three rural properties with a history of macrocyclic lactones ineffectiveness against the cattle tick, and low frequency use of fipronil in the herd, were selected for the study. The animals were randomized according to the mean tick counts, performed on days 3, 2 and I, into three groups with 10 animals each: T01, control (saline solution); T02, subcutaneous ivermectin (0.63 mg/kg) and T03, topical fipronil (1 mg/kg). Treatment was performed on day 0. Counts of partially engorged female ticks were performed on days 3,7 and 14 post-treatment (DPT), and then every 7 days until the 49th DPT. In all three experiments, it was possible to diagnose resistance of R. (B.) microplus to ivermectin (0.63 mg/kg). The maximum efficacy (arithmetic mean) obtained for ivermectin was 64% in experiment II. On the other hand, the formulation containing fipronil (1 mg/kg) reached high efficacy values (>= 97%) in all three experiments. The results from all experiments in this study demonstrate the high efficacy of 1 mg/kg fipronil, administered pour-on in naturally infested cattle, against strains of R. (B.) microplus that are resistant to 630 mcg/kg ivermectin. (C) 2014 Elsevier B.V. All rights reserved.

Effect of a spray formulation on the reproductive parameters of a susceptible population of Rhipicephalus (Boophilus) microplus

The number of studies emphasizing the possible damage that acaricidal spray formulations can cause on engorged female ticks'reproductive parameters is small. The present study evaluated the deleterious effects of a spray formulation (dichlorvos 60% + chlorpyrifos 20%) on the reproductive parameters of a susceptible population of Rhipicephalus (B.) microplus females, using the Stall Test. The ticks were allocated randomly to treatments according to the mean numbers of females detached from each cow on days -3, -2 and -1 and the cattle pen location. The numbers of engorged female ticks that naturally detached from the cattle were counted daily from day 1 to day 30. For each group, 20 detached engorged female ticks or the available number collected daily were evaluated regarding reproductive parameters. Associations of organophosphates demonstrated elevated acaricidal efficacy, as well as deleterious effects on the reproductive parameters of R. (B.) microplus females. The engorged female weight (days 1 to 7), weight of egg masses (days 5 to 10) and larval hatching percentage (days 5 to 19) were decreased (P <= 0.05). It is possible that a formulation can lead to deleterious effects on R. (B.) microplus females when the tick population analyzed shows elevated sensitivity towards a particular formulation. However, further studies need to be conducted.

Purification and characterization of Hb 98-114: A novel hemoglobin-derived antimicrobial peptide from the midgut of Rhipicephalus (Boophilus) microplus

The antimicrobial activity of hemoglobin fragments (hemocidins) has been reported in a variety of models. The cattle tick Rhipicephalus (Boophilus) microplus is a blood sucking arthropod from where the first in vivo-generated hemocidin was characterized (Hb 33-61). In the present work we identified a novel antimicrobial peptide from the midgut of fully engorged R. (B.) microplus females, which comprises the amino acids 98-114 of the alpha subunit of bovine hemoglobin, and was designated Hb 98-114. This peptide was active against several yeast and filamentous fungi, although no activity was detected against bacteria up to 50 mu M of the synthetic peptide. Hb 98-114 was capable of permeabilizing Candida albicans cell membrane and had a fungicidal effect against this yeast. Circulardichroism (CD) and nuclear magnetic resonance (NMR) experiments showed that Hb 98-114 has a random conformation in aqueous solution but switches to an alpha-helical conformation in the presence of sodium dodecyl sulfate (SDS). This alpha helix adopts an amphipathic structure which may be the mechanism of cell membrane permeabilization. Importantly, Hb 98-114 may play an important role in defending the tick midgut against fungal pathogens and is the first hemocidin with specific antifungal activity to be characterized. (C) 2012 Elsevier Inc. All rights reserved.

Pathogenicity of Metarhizium anisopliae towards Rhipicephalus (Boophilus) microplus under laboratory and field conditions

Metarhizium anisopliae is one of the most studied agents of biological control of several arthropod plagues, including the cattle tick Rhipicephalus (Boophilus) microplus. Studies have been conducted to assess the fungal complex infection process towards its hosts. To accomplish that, mutant strains overexpressing or lacking assumed determinant genes for the process were constructed over the years. A fundamental experiment to demonstrate a particular gene or set of genes participation is the bioassay. The comparison of bioassays using wild and engineered strains is an essential tool to affirm a given gene is crucial in the process. Therefore, the in vitro bioassays should mimic the results obtained in tests under field conditions. In this study, tests under laboratory and filed conditions were done and a correlation analysis was performed in order to statistically validate in vitro bioassays. Tick egg laying, larvae hatching and host mortality were recorded in each experiment through 21 days, both under laboratory and field conditions. In all cases, M. anisopliae treatments were statistically different from the control treatments. A linear regression analysis was performed between the cases. Laboratory results showed a statistically significant correlation with the field conditions using the Pearson's Correlation Test (P < 0.01 host mortality - 0.969, tick egg laying - 0.977 and larvae hatching - 0.956). These results legitimize the in vitro bioassays and, therefore, constitute them as a valid tool for studying this fungus behavior, so they can be used to infer M. anisopliae response towards R. (Boophilus) microplus.

Applicability of in vitro bioassays for the diagnosis of ivermectin resistance in Rhipicephalus microplus (Acari: Ixodidae)

The applicability of laboratory bioassays to diagnose ivermectin (IVM) resistance in Rhipicephalus microplus was evaluated. Adult immersion tests (AITs), larval immersion tests (LITs) and larval packet tests (LPTs) were performed to characterise the effects of ivermectin toxicity on adults and larvae of a susceptible reference strain. The AIT was determined to be a reasonable assay but requires a large number of individuals to attain interpretable results. The LIT and LPT were validated with an IVM resistant strain, revealing resistance ratios (RRs) of 6.73 and 1.49, respectively. In a field survey, nine different populations of cattle tick from the states of Sao Paulo and Mato Grosso do Sul, Brazil, were analysed with the LIT. Populations without previous exposure to ivermectin exhibited RRs between 0.87 and 1.01. Populations previously exposed to IVM showed RRs between 1.83 and 4.62. The LIT was more effective at discriminating between resistant and susceptible populations than the LPT. The use of the LIT is recommended for the diagnosis of ivermectin resistance in R microplus. (C) 2011 Elsevier B.V. All rights reserved.

La garrapata que produce la fiebre bovina o tristeza. Lo que todo niño campesino debe saber acerca de la garrapata que ataca al ganado vacuno. Libro ilustrado con excelentes grabados que muestran cómo las garrapatas que transmiten dicha fiebre le roban leche, carne y dinero a los agricultores y ganaderos, y les matan miles de cabezas de ganado.

Issued by the Bureau of Animal Industry.

Sequencing a new target genome: The Boophilus microplus (Acari : Ixodidae) genome project

The southern cattle tick, Boophilus microplus (Canestrini), causes annual economic losses in the hundreds of millions of dollars to cattle producers throughout the world, and ranks as the most economically important tick from a global perspective. Control failures attributable to the development of pesticide resistance have become commonplace, and novel control technologies are needed. The availability of the genome sequence will facilitate the development of these new technologies, and we are proposing sequencing to a 4-6X draft coverage. Many existing biological resources are available to facilitate a genome sequencing project, including several inbred laboratory tick strains, a database of approximate to 45,000 expressed sequence tags compiled into a B. microplus Gene Index, a bacterial artificial chromosome (BAC) library, an established B. microplus cell line, and genomic DNA suitable for library synthesis. Collaborative projects are underway to map BACs and cDNAs to specific chromosomes and to sequence selected BAC clones. When completed, the genome sequences from the cow, B. microphis, and the B. microphis-borne pathogens Babesia bovis and Anaplasma marginale will enhance studies of host-vector-pathogen systems. Genes involved in the regeneration of amputated tick limbs and transitions through developmental stages are largely unknown. Studies of these and other interesting biological questions will be advanced by tick genome sequence data. Comparative genomics offers the prospect of new insight into many, perhaps all, aspects of the biology of ticks and the pathogens they transmit to farm animals and people. The B. microplus genome sequence will fill a major gap in comparative genomics: a sequence from the Metastriata lineage of ticks. The purpose of the article is to synergize interest in and provide rationales for sequencing the genome of B. microplus and for publicizing currently available genomic resources for this tick.

Productivity and health effects of anaplasmosis and babesiosis on Bos indicus cattle and their crosses, and the effects of differing intensity of tick control in Australia

Tick fever is an important disease of cattle where Rhipicephalus (Boophilus) microplus acts as a vector for the three causal organisms Babesia bovis, Babesia bigemina and Anaplasma marginale. Bos indicus cattle and their crosses are more resistant to the clinical effects of infection with B. bovis and B. bigemina than are Bos taurus cattle. Resistance is not complete, however, and herds of B. indicus-cross cattle are still at risk of babesiosis in environments where exposure to B. bovis is light in most years but occasionally high. The susceptibility of B. indicus cattle and their crosses to infection with A. marginale is similar to that of B. taurus cattle. In herds of B. indicus cattle and their crosses the infection rate of Babesia spp. and A. marginale is lowered because fewer ticks are likely to attach per day due to reduced numbers of ticks in the field (long-term effect on population, arising from high host resistance) and because a smaller proportion of ticks that do develop to feed on infected cattle will in turn be infected (due to lower parasitaemia). As a consequence, herds of B. indicus cattle are less likely than herds of B. taurus cattle to have high levels of population immunity to babesiosis or anaplasmosis. The effects of acaricide application on the probability of clinical disease due to anaplasmosis and babesiosis are unpredictable and dependent on the prevalence of infection in ticks and in cattle at the time of application. Attempting to manipulate population immunity through the toleration of specific threshold numbers of ticks with the aim of controlling tick fever is not reliable and the justification for acaricide application should be for the control of ticks rather than for tick fever. Vaccination of B. indicus cattle and their crosses is advisable in all areas where ticks exist, although vaccination against B. bigemina is probably not essential in pure B. indicus animals.

A genome-wide association study of tick burden and milk composition in cattle.

To study the genetic basis of tick burden and milk production and their interrelationship, we collected a sample of 1961 cattle with multiple tick counts from northern Australia of which 973 had dairy production data in the Australian Dairy Herd Information Service database. We calculated heritabilities, genetic and phenotypic correlations for these traits and showed a negative relationship between tick counts and milk and milk component yield. Tests of polymorphisms of four genes associated with milk yield, ABCG2, DGAT1, GHR and PRLR, showed no statistically significant effect on tick burden but highly significant associations to milk component yield in these data and we confirmed separate effects for GHR and PRLR on bovine chromosome 20. To begin to identify some of the molecular genetic bases for these traits, we genotyped a sample of 189 of these cattle for 7397 single nucleotide polymorphisms in a genome-wide association study. Although the allele effects for adjusted milk fat and protein yield were highly correlated (r = 0.66), the correlations of allele effects of these milk component yields and tick burden were small (|r| <= 0.10). These results agree in general with the phenotypic correlations between tick counts and milk component yield and suggest that selection on markers for tick burden or milk component yield may have no undesirable effect on the other trait.

Is endemic stability of tick-borne disease in cattle a useful concept?

Endemic stability is a widely used term in the epidemiology of ticks and tick-borne diseases. It is generally accepted to refer to a state of a host tick pathogen interaction in which there is a high level of challenge of calves by infected ticks, absence of clinical disease in calves despite infection, and a high level of immunity in adult cattle with consequent low incidence of clinical disease. Although endemic stability is a valid epidemiological concept, the modelling studies that underpinned subsequent studies on the epidemiology of tick-borne diseases were specific to a single host tick pathogen system, and values derived from these models should not be applied in other regions or host tick pathogen systems.

Efficacy of Rhipicephalus (Boophilus) microplus Bm86 against Hyalomma dromedarii and Amblyomma cajennense tick infestations in camels and cattle

The recombinant Bm86-based tick vaccines have shown their efficacy for the control of cattle ticks, Rhipicephalus (Boophilus) microplus and R. annulatus infestations. However, cattle ticks often co-exist with multi-host ticks such as Hyalomma and Amblyomma species, thus requiring the control of multiple tick infestations for cattle and other hosts. Vaccination trials using a R. microplus recombinant Bm86-based vaccine were conducted in cattle and camels against Hyalomma dromedarii and in cattle against Amblyomma cajennense immature and adult ticks. The results showed an 89% reduction in the number of H. dromedarii nymphs engorging on vaccinated cattle, and a further 32% reduction in the weight of the surviving adult ticks. In vaccinated camels, a reduction of 27% and 31% of tick engorgement and egg mass weight, respectively was shown, while egg hatching was reduced by 39%. However, cattle vaccination with Bm86 did not have an effect on A. cajennense tick infestations. These results showed that Bm86 vaccines are effective against R. microplus and other tick species but improved vaccines containing new antigens are required to control multiple tick infestations. (C) 2012 Elsevier Ltd. All rights reserved.