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2023-03-15T21:53:41Z2023-03-15T21:53:41Z2021-02-0411https://doi.org/10.3389/fimmu.2020.6111041664-3224http://hdl.handle.net/1843/50932https://orcid.org/0000-0003-1772-2336https://orcid.org/0000-0002-9771-3183https://orcid.org/0000-0003-3357-2497https://orcid.org/0000-0003-4843-0088https://orcid.org/0000-0002-5668-3934https://orcid.org/0000-0002-7924-8038https://orcid.org/0000-0002-5589-9450https://orcid.org/0000-0003-4181-7546https://orcid.org/0000-0002-4713-575Xhttps://orcid.org/0000-0003-0974-7357https://orcid.org/0000-0002-1272-4386Amblyomma sculptum is the main tick associated with human bites in Brazil and the main vector of Rickettsia rickettsii, the causative agent of the most severe form of Brazilian spotted fever. Molecules produced in the salivary glands are directly related to feeding success and vector competence. In the present study, we identified sequences of A. sculptum salivary proteins that may be involved in hematophagy and selected three proteins that underwent functional characterization and evaluation as vaccine antigens. Among the three proteins selected, one contained a Kunitz_bovine pancreatic trypsin inhibitor domain (named AsKunitz) and the other two belonged to the 8.9 kDa and basic tail families of tick salivary proteins (named As8.9kDa and AsBasicTail). Expression of the messenger RNA (mRNA) encoding all three proteins was detected in the larvae, nymphs, and females at basal levels in unfed ticks and the expression levels increased after the start of feeding. Recombinant proteins rAs8.9kDa and rAsBasicTail inhibited the enzymatic activity of factor Xa, thrombin, and trypsin, whereas rAsKunitz inhibited only thrombin activity. All three recombinant proteins inhibited the hemolysis of both the classical and alternative pathways; this is the first description of tick members of the Kunitz and 8.9kDa families being inhibitors of the classical complement pathway. Mice immunization with recombinant proteins caused efficacies against A. sculptum females from 59.4% with rAsBasicTail immunization to more than 85% by immunization with rAsKunitz and rAs8.9kDa. The mortality of nymphs fed on immunized mice reached 70–100%. Therefore, all three proteins are potential antigens with the possibility of becoming a new tool in the control of A. sculptum.Amblyomma sculptum é o principal carrapato associado a picadas humanas no Brasil e o principal vetor da Rickettsia rickettsii, agente causador da forma mais grave da febre maculosa brasileira. Moléculas produzidas nas glândulas salivares estão diretamente relacionadas ao sucesso alimentar e competência vetorial. No presente estudo, identificamos sequências de proteínas salivares de A. sculptum que podem estar envolvidas na hematofagia e selecionamos três proteínas que passaram por caracterização funcional e avaliação como antígenos vacinais. Entre as três proteínas selecionadas, uma continha um domínio inibidor de tripsina pancreática Kunitz_bovine (denominada AsKunitz) e as outras duas pertenciam às famílias 8,9 kDa e cauda básica das proteínas salivares de carrapatos (denominadas As8.9kDa e AsBasicTail). A expressão do RNA mensageiro (mRNA) que codifica todas as três proteínas foi detectada nas larvas, ninfas e fêmeas em níveis basais em carrapatos não alimentados e os níveis de expressão aumentaram após o início da alimentação. As proteínas recombinantes rAs8.9kDa e rAsBasicTail inibiram a atividade enzimática do fator Xa, trombina e tripsina, enquanto rAsKunitz inibiu apenas a atividade da trombina. Todas as três proteínas recombinantes inibiram a hemólise das vias clássica e alternativa; esta é a primeira descrição de carrapatos das famílias Kunitz e 8,9kDa como inibidores da via clássica do complemento. A imunização de camundongos com proteínas recombinantes causou eficácia contra fêmeas de A. sculptum de 59,4% com imunização rAsBasicTail para mais de 85% com imunização com rAsKunitz e rAs8.9kDa. A mortalidade de ninfas alimentadas com camundongos imunizados atingiu 70-100%. Portanto, todas as três proteínas são potenciais antígenos com possibilidade de se tornarem uma nova ferramenta no controle de A. sculptum.CNPq - Conselho Nacional de Desenvolvimento Científico e TecnológicoFAPEMIG - Fundação de Amparo à Pesquisa do Estado de Minas GeraisCAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorOutra AgênciaengUniversidade Federal de Minas GeraisUFMGBrasilICB - DEPARTAMENTO DE MORFOLOGIAICB - DEPARTAMENTO DE PARASITOLOGIAFrontiers in ImmunologyAmblyommaCarrapatosSalivaDoenças transmitidas por carrapatosInibidores de proteasesInativadores do complementoVacinasAmblyomma sculptumSalivaAntihemostaticComplement inhibitorsVaccineAmblyomma sculptum salivary protease inhibitors as potential anti-tick vaccinesInibidores de protease salivar de Amblyomma sculptum como potenciais vacinas anticarrapatosinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articlehttps://www.frontiersin.org/articles/10.3389/fimmu.2020.611104/fullGabriel Cerqueira Alves CostaIzabela Cosso Tavares RibeiroOtoni Alves de Oliveira Melo JúniorNelder de Figueiredo GontijoMaurício Roberto Viana Sant'AnnaMarcos Horacio PereiraGrasielle Caldas D`Ávila PessoaLeonardo Barbosa KoerichFabiano OliveiraJesus G. ValenzuelaRodolfo Cordeiro GiunchettiRicardo Toshio FujiwaraDaniella Castanheira BartholomeuRicardo Nascimento Araújoapplication/pdfinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UFMGinstname:Universidade Federal de Minas Gerais (UFMG)instacron:UFMGLICENSELicense.txtLicense.txttext/plain; charset=utf-82042https://repositorio.ufmg.br/bitstream/1843/50932/1/License.txtfa505098d172de0bc8864fc1287ffe22MD51ORIGINALAmblyomma sculptum Salivary Protease Inhibitors as Potential Anti-Tick Vaccines.pdfAmblyomma sculptum Salivary Protease Inhibitors as Potential Anti-Tick Vaccines.pdfapplication/pdf2452750https://repositorio.ufmg.br/bitstream/1843/50932/2/Amblyomma%20sculptum%20Salivary%20Protease%20Inhibitors%20as%20Potential%20Anti-Tick%20Vaccines.pdfcfcd61e3388271db577e219dc0aeabf1MD521843/509322023-03-15 19:04:42.9oai:repositorio.ufmg.br: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Repositório InstitucionalPUBhttps://repositorio.ufmg.br/oaiopendoar:2023-03-15T22:04:42Repositório Institucional da UFMG - Universidade Federal de Minas Gerais (UFMG)false
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