RESUMO
Rhipicephalus (Boophilus) microplus has substantial economic impact on the cattle breeding industry and, chemical control and tick resistance development are the major concern. There is a worldwide search for new options, and control using vaccines has been the main focus nowadays. Studies performed in Brazil found that Bm86-based immunization of bovines reduced the infestation of R. (B.) microplus of vaccinated bovines by 45% to 60%. Native Boophilusmicroplus tripsin inhibitors (BmTIs) with trypsin-, kallikrein-, and elastase-inhibiting activities have been used as immunogens in bovines reaching 72.8.% of efficacy. The reverse vaccinology approach has also been used for antigen search using transcriptome analysis to identify and characterize potential antigens. Study has generated more than 600 million sequences using RNA-seq of larvae, nymphs, salivary glands, intestines, and ovaries of the tick R. (B) microplus. Based on the set of transcripts obtained using this strategy, a total of 20,326 protein sequences have been identified. A pipeline analysis built in house identified the protein sequences that were most likely to be immunogenic based on the overall structural characteristic analysis.
Assuntos
Doenças dos Bovinos/prevenção & controle , Doenças dos Bovinos/parasitologia , Rhipicephalus/genética , Rhipicephalus/imunologia , Infestações por Carrapato/veterinária , Vacinas/imunologia , Animais , Brasil , Bovinos , Expressão Gênica , Infestações por Carrapato/prevenção & controle , Inibidores da Tripsina/imunologia , Vacinação/veterinária , Vacinas/administração & dosagemRESUMO
The cattle tick, Rhipicephalus microplus, is regarded as the most economically important ectoparasite of livestock globally. Control is achieved primarily through the use of acaricides. This approach is hampered by the development of resistance to commercial acaricides among cattle tick populations. Vaccination against R. microplus infestation is another technology that can be integrated for effective cattle tick control. Proteins belonging to the Kunitz-BPTI family are abundant in cattle tick salivary glands, midgut, and ovaries. These organs are attractive targets for the development of a novel cattle tick vaccine. Efficacy assessment against cattle tick infestation in bovines using a vaccine containing the recombinant form of a member of the Kunitz family from R. microplus produced in a yeast expression system is reported for the first time here. The yeast Pichia pastoris was bioengineered to produce the recombinant version of a trypsin inhibitor that is expressed in cattle tick larvae (rRmLTI). Immunization with rRmLTI afforded 32% efficacy against R. microplus. The estimated molecular weight of rRmLTI was 46 kDa. Structural homology to the native form of the larval trypsin inhibitor was documented by recognition of rRmLTI in Western-blots using polyclonal antibodies from mice immunized with cattle tick larval extract or rRmLTI. Bioinformatics analysis of the partial nucleotide and deduced amino acid sequences indicated that the rRmLTI closely resembles BmTI-6, which is a three-headed Kunitz protein present in cattle tick ovary and fat tissue.
Assuntos
Doenças dos Bovinos/prevenção & controle , Rhipicephalus/imunologia , Infestações por Carrapato/veterinária , Inibidores da Tripsina/imunologia , Vacinas/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Bovinos , Doenças dos Bovinos/parasitologia , Feminino , Imunidade Humoral , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Pichia/metabolismo , Proteínas Recombinantes/imunologia , Infestações por Carrapato/prevenção & controle , Vacinação/veterináriaRESUMO
Cattle tick control remains a serious problem in cattle farms worldwide, due to the limited success achieved with chemicals. Although the use of vaccines for tick control may open possibilities for an integrated approach, the search for other protective antigens is still necessary to improve control. Boophilus microplus is a rich source of BmTIs, serine proteinase inhibitors that are present throughout the tick's lifecycle. The present paper reports a pen trial conducted to evaluate the performance of a synthetic BmTI N-terminal fragment as antigen against the tick in bovines. The trial was conducted with two groups of eight crossbred cattle under controlled infestation: one group was vaccinated with BmTI peptide using saponin as adjuvant and the other was kept as control. Challenge was performed with 15,000 larvae. The specific IgG response was measured by ELISA, with successful outcomes for vaccinated animals. Vaccination resulted in an 18.4% level of efficacy when compared with the control group.
Assuntos
Antígenos/imunologia , Doenças dos Bovinos/prevenção & controle , Rhipicephalus/imunologia , Infestações por Carrapato/veterinária , Inibidores da Tripsina/imunologia , Animais , Bovinos , Doenças dos Bovinos/parasitologia , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Imunoglobulina G/biossíntese , Imunoglobulina G/sangue , Calicreínas/antagonistas & inibidores , Cinética , Infestações por Carrapato/prevenção & controle , Inibidores da Tripsina/síntese química , Vacinação/veterináriaRESUMO
Boophilus microplus trypsin inhibitors (BmTIs) present in larvae were preliminarily characterized as active proteins, approximately 10-18 kDa, by SDS-PAGE. BmTIs showed trypsin inhibitory activity on reverse zymography containing gelatin (0.03%) and also inhibited others serine proteinases (human neutrophil elastase and human plasma kallikrein). Bos indicus, Nelore breed calves, previously sensitized with BmTIs and challenged with tick larvae (20,000 larvae/animal), showed 72.8% efficacy to interfere in tick development with 69.7% and 71.3% reduction of both tick number and egg weight, respectively. Cattle BmTls antiserum titer was approximately 1:8000. The maximum level of BmTls antibody production was detected 40 days after the first immunization by ELISA. Our preliminary results suggest that B. microplus serine proteinase inhibitors may play a role in the tick larvae fixation and feeding processes. Therefore, the development of antibodies against BmTIs might impair the normal parasitism.