Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus
Autor(a) principal: | |
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Data de Publicação: | 2017 |
Tipo de documento: | Tese |
Idioma: | por |
Título da fonte: | Biblioteca Digital de Teses e Dissertações da UFRRJ |
Texto Completo: | https://tede.ufrrj.br/jspui/handle/jspui/2587 |
Resumo: | This study aimed to perform the molecular characterization of the B. caballi rap-1 gene and to analyze the degree of heterogeneity of the T. equi 18S rDNA gene of naturally infected horses in the state of Rio de Janeiro. Complete sequences of the T. equi 18S rDNA gene (n = 20) were obtained. The PCR products were cloned and sequenced. Phylogenetic analyzes were performed on ARB-Silva basis. For the molecular characterization of a fragment of the B. caballi rap-1 gene, 32 cELISA and qPCR positive equine samples were used. A nested PCR was developed to amplify a fragment of the rap-1 gene and a conventional PCR, aiming the amplification of the complete gene. The products were sequenced. Sequences were submitted to the Maximum-Likelihood reconstruction method with Poisson model and 1000 replicates. In the analysis of T. equi, twelve different isolates were observed in equines from the state of Rio de Janeiro and were grouped into two of the three clades formed (I, II), most of them grouped in clade I (70%, n = 14/20). The sequences of T. equi are grouped within a clade different from other Theilerids. It was possible to amplify a fragment of the B. caballi rap-1 gene from 23 sequences using nested PCR, identified as single sequences. One of the sequences showed twelve mutation points. In cluster analysis, the Brazilian samples belonged to clade A, being analyzed as amino acids (152a) or nucleotides (1244 bp). The aim of the second chapter was to evaluate the dinamic of infection of T. equi in R. (B.) microplus fed in chronically infected horses. The larvae obtained from a hemoparasite-free colony were used in the infestation of an equine chronically infected by T. equi. Ticks and the horse blood were collected every two days. Ticks were dissected and stored in pools according to the pos-infestation day, evolutionary stage and gender. Some of these specimens were stored entire. Salivary gland and ovary samples were stored properly for Transmission Electron Microscopy. Some stages, prior to ecdysis, were stored in B.O.D. to molt. Samples were tested by qPCR. Regarding to entire tick samples, more than 90% of the samples were positive for T. equi. The mean of parasite load in entire ticks and salivary glands presented gradual increasing with the progress of the feeding period. Of the 45 salivary gland samples, 77.78% were positive for T. equi and had a higher infection rate at the end of the experimental period. It was found freshly molted ticks positives for T. equi by qPCR, indicating transestadial transmission. 84,38% of R. (B.) microplus ovary samples were considered positives for T. equi by molecular tools. Males of R. (B.) microplus were found parasiting the equine for 60 days. This fact associated with the high positivity for T. equi indicates males as an epidemiologically important gender. Transmission micrographs revealed morphologic structures compatible with a Piroplasmid in the acini of salivary glands and ovaries. The phylogenetic analysis of the T. equi 18S rDNA gene and of B. caballi rap-1 gene showed that different isolates co-exist in the same area in Rio de Janeiro, Brazil. Experimently, it was possible to observe that R. (B.) microplus could be infected with T. equi and that the forms of development of this parasite are capable of multiply inside salivary glands and, possibly, ovaries. |
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Massard, Carlos Luiz257.781.297-34Santos, Huarrisson Azevedo983.833.295-04Pires, Marcus Sandes055.166.387-12Massard, Carlos LuizLeite, Rom?rio CerqueiraLabruna, Marcelo BahiaDias, Roberto J?nio PedrosoSantos, Tiago Marques dos110.795.017-11http://lattes.cnpq.br/8817867478588076Peixoto, Maristela Peckle2019-01-31T18:26:42Z2017-02-22PEIXOTO, Maristela Peckle. Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus. 2017. 133 f. Tese (Doutorado em Ci?ncias Veterin?rias). Instituto de Veterin?ria, Departamento de Parasitologia Animal, Universidade Federal Rural do Rio de Janeiro, Serop?dica, RJ, 2017.https://tede.ufrrj.br/jspui/handle/jspui/2587This study aimed to perform the molecular characterization of the B. caballi rap-1 gene and to analyze the degree of heterogeneity of the T. equi 18S rDNA gene of naturally infected horses in the state of Rio de Janeiro. Complete sequences of the T. equi 18S rDNA gene (n = 20) were obtained. The PCR products were cloned and sequenced. Phylogenetic analyzes were performed on ARB-Silva basis. For the molecular characterization of a fragment of the B. caballi rap-1 gene, 32 cELISA and qPCR positive equine samples were used. A nested PCR was developed to amplify a fragment of the rap-1 gene and a conventional PCR, aiming the amplification of the complete gene. The products were sequenced. Sequences were submitted to the Maximum-Likelihood reconstruction method with Poisson model and 1000 replicates. In the analysis of T. equi, twelve different isolates were observed in equines from the state of Rio de Janeiro and were grouped into two of the three clades formed (I, II), most of them grouped in clade I (70%, n = 14/20). The sequences of T. equi are grouped within a clade different from other Theilerids. It was possible to amplify a fragment of the B. caballi rap-1 gene from 23 sequences using nested PCR, identified as single sequences. One of the sequences showed twelve mutation points. In cluster analysis, the Brazilian samples belonged to clade A, being analyzed as amino acids (152a) or nucleotides (1244 bp). The aim of the second chapter was to evaluate the dinamic of infection of T. equi in R. (B.) microplus fed in chronically infected horses. The larvae obtained from a hemoparasite-free colony were used in the infestation of an equine chronically infected by T. equi. Ticks and the horse blood were collected every two days. Ticks were dissected and stored in pools according to the pos-infestation day, evolutionary stage and gender. Some of these specimens were stored entire. Salivary gland and ovary samples were stored properly for Transmission Electron Microscopy. Some stages, prior to ecdysis, were stored in B.O.D. to molt. Samples were tested by qPCR. Regarding to entire tick samples, more than 90% of the samples were positive for T. equi. The mean of parasite load in entire ticks and salivary glands presented gradual increasing with the progress of the feeding period. Of the 45 salivary gland samples, 77.78% were positive for T. equi and had a higher infection rate at the end of the experimental period. It was found freshly molted ticks positives for T. equi by qPCR, indicating transestadial transmission. 84,38% of R. (B.) microplus ovary samples were considered positives for T. equi by molecular tools. Males of R. (B.) microplus were found parasiting the equine for 60 days. This fact associated with the high positivity for T. equi indicates males as an epidemiologically important gender. Transmission micrographs revealed morphologic structures compatible with a Piroplasmid in the acini of salivary glands and ovaries. The phylogenetic analysis of the T. equi 18S rDNA gene and of B. caballi rap-1 gene showed that different isolates co-exist in the same area in Rio de Janeiro, Brazil. Experimently, it was possible to observe that R. (B.) microplus could be infected with T. equi and that the forms of development of this parasite are capable of multiply inside salivary glands and, possibly, ovaries.Os objetivos deste trabalho foram caracterizar molecularmente Theileria equi e Babesia caballi de amostras de equinos do Rio de Janeiro, bem como realizar um estudo da din?mica da infec??o de T. equi em Rhipicephalus (Boophilus) microplus. No primeiro cap?tulo realizou-se a caracteriza??o molecular de um fragmento do gene rap-1 de B. caballi e a an?lise do grau de heterogeneidade do gene 18S rDNA em T. equi em amostras de equinos naturalmente infectados, do estado do Rio de janeiro. Foram obtidas vinte sequ?ncias do gene 18S rDNA de T. equi. As an?lises filogen?ticas foram realizadas baseadas no ARB-Silva. Para a caracteriza??o molecular de um fragmento do gene rap-1 de B. caballi foram utilizadas amostras positivas por cELISA e qPCR. Uma nested PCR foi desenvolvida para amplificar um fragmento do gene rap-1, e uma PCR convencional, para o gene completo. As sequ?ncias de amino?cidos e nucleot?deos sequenciados foram submetidas ao m?todo Maximum-Likelihood. Na an?lise das sequ?ncias do 18S rDNA de T. equi, doze diferentes isolados foram observados em equinos no estado do Rio de Janeiro, sendo agrupados em dois dos tr?s clados formados (I, II). As sequ?ncias de T. equi se agrupam dentro de um clado diferente dos demais Theiler?deos. Uma das 23 sequ?ncias de amino?cidos do gene rap-1 apresentou doze pontos de muta??o. Na an?lise de agrupamentos, as amostras brasileiras agruparam no clado A, sendo estas analisadas como amino?cidos (152 aas) ou nucleot?deos (1244 pb). No segundo cap?tulo foram avaliados aspectos da din?mica da infec??o de T. equi em R. (B.) microplus alimentados em equino cronicamente infectado. Larvas de uma col?nia livre de hemoparasitos foram utilizadas. Carrapatos e sangue do equino foram coletados de dois em dois dias. Os carrapatos foram dissecados e armazenados inteiros em ?pools? de acordo com o dia p?s-infesta??o, est?gio evolutivo e g?nero. Amostras de gl?ndula salivar e ov?rio foram armazenados para Microscopia Eletr?nica de Transmiss?o. As amostras foram testadas pela qPCR. Com rela??o aos carrapatos inteiros, mais de 90% das amostras foram positivas para T. equi. A carga parasit?ria m?dia das amostras de carrapatos inteiros e gl?ndulas salivares apresentaram aumento gradual com o avan?ar do per?odo de alimenta??o. Das 45 amostras de gl?ndula salivar, 77,78% foram positivas para T. equi e tiveram maior taxa de infec??o no final do per?odo experimental. Foi poss?vel observar positividade em carrapatos rec?m emergidos, ninfas e adultos, indicando a ocorr?ncia de transmiss?o transestadial. A positividade de amostras de ov?rios foi de 84,38%. A maior perman?ncia de machos sobre o equino, associado ? alta positividade deste tipo de amostra indicam que o macho pode ter participa??o significativa na transmiss?o. Visualizaram-se estruturas morfologicamente compat?veis com um piroplasma atrav?s de micrografia de transmiss?o em ov?rios e gl?ndulas salivares de em R. (B.) microplus. A an?lise do gene 18S rDNA de T. equi e do gene rap-1 de B. caballi de amostras brasileiras demonstrou que diferentes isolados coexistem na mesma ?rea. Experimentalmente, observou-se que R. (B.) microplus ? capaz de se infectar com T. equi e que as formas multiplicam-se nas gl?ndulas salivares e, possivelmente, em ov?rios.Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2019-01-31T18:26:42Z No. of bitstreams: 1 2017 - Maristela Peckle Peixoto.pdf: 10701979 bytes, checksum: faffb982c158894c0c385ae44c6a8b7c (MD5)Made available in DSpace on 2019-01-31T18:26:42Z (GMT). No. of bitstreams: 1 2017 - Maristela Peckle Peixoto.pdf: 10701979 bytes, checksum: faffb982c158894c0c385ae44c6a8b7c (MD5) Previous issue date: 2017-02-22CAPES - Coordena??o de Aperfei?oamento de Pessoal de N?vel SuperiorCNPq - Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gicoFAPERJ - Funda??o Carlos Chagas Filho de Amparo ? Pesquisa do Estado do Rio de Janeiroapplication/pdfhttps://tede.ufrrj.br/retrieve/54092/2017%20-%20Maristela%20Peckle%20Peixoto.pdf.jpgporUniversidade Federal Rural do Rio de JaneiroPrograma de P?s-Gradua??o em Ci?ncias Veterin?riasUFRRJBrasilInstituto de Veterin?riaDiversidade gen?ticapiroplasmose equinatheileriose equinatransmiss?o transestadialtransmiss?o transovarianaGenetic diversityequine piroplasmosisequine teileriosistransestadial transmissiontransovarian transmissionMedicina Veterin?riaAgentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplusEquine Piroplasmosis: genetic diversity of Theileria equi and Babesia caballi of naturally infected horses from the state of Rio de Janeiro and the dynamics of Theileria equi infection in Rhipicephalus (Boophilus) microplusinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/openAccessreponame:Biblioteca Digital de Teses e Dissertações da UFRRJinstname:Universidade Federal Rural do Rio de Janeiro (UFRRJ)instacron:UFRRJTHUMBNAIL2017 - Maristela Peckle Peixoto.pdf.jpg2017 - Maristela Peckle Peixoto.pdf.jpgimage/jpeg1943http://localhost:8080/tede/bitstream/jspui/2587/18/2017+-+Maristela+Peckle+Peixoto.pdf.jpgcc73c4c239a4c332d642ba1e7c7a9fb2MD518TEXT2017 - Maristela Peckle Peixoto.pdf.txt2017 - Maristela Peckle Peixoto.pdf.txttext/plain364328http://localhost:8080/tede/bitstream/jspui/2587/17/2017+-+Maristela+Peckle+Peixoto.pdf.txt531461974ced5f1e831703c16364cbe0MD517ORIGINAL2017 - Maristela Peckle Peixoto.pdf2017 - Maristela Peckle Peixoto.pdfapplication/pdf10701979http://localhost:8080/tede/bitstream/jspui/2587/2/2017+-+Maristela+Peckle+Peixoto.pdffaffb982c158894c0c385ae44c6a8b7cMD52LICENSElicense.txtlicense.txttext/plain; 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dc.title.por.fl_str_mv |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
dc.title.alternative.eng.fl_str_mv |
Equine Piroplasmosis: genetic diversity of Theileria equi and Babesia caballi of naturally infected horses from the state of Rio de Janeiro and the dynamics of Theileria equi infection in Rhipicephalus (Boophilus) microplus |
title |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
spellingShingle |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus Peixoto, Maristela Peckle Diversidade gen?tica piroplasmose equina theileriose equina transmiss?o transestadial transmiss?o transovariana Genetic diversity equine piroplasmosis equine teileriosis transestadial transmission transovarian transmission Medicina Veterin?ria |
title_short |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
title_full |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
title_fullStr |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
title_full_unstemmed |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
title_sort |
Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus |
author |
Peixoto, Maristela Peckle |
author_facet |
Peixoto, Maristela Peckle |
author_role |
author |
dc.contributor.advisor1.fl_str_mv |
Massard, Carlos Luiz |
dc.contributor.advisor1ID.fl_str_mv |
257.781.297-34 |
dc.contributor.advisor-co1.fl_str_mv |
Santos, Huarrisson Azevedo |
dc.contributor.advisor-co1ID.fl_str_mv |
983.833.295-04 |
dc.contributor.advisor-co2.fl_str_mv |
Pires, Marcus Sandes |
dc.contributor.advisor-co2ID.fl_str_mv |
055.166.387-12 |
dc.contributor.referee1.fl_str_mv |
Massard, Carlos Luiz |
dc.contributor.referee2.fl_str_mv |
Leite, Rom?rio Cerqueira |
dc.contributor.referee3.fl_str_mv |
Labruna, Marcelo Bahia |
dc.contributor.referee4.fl_str_mv |
Dias, Roberto J?nio Pedroso |
dc.contributor.referee5.fl_str_mv |
Santos, Tiago Marques dos |
dc.contributor.authorID.fl_str_mv |
110.795.017-11 |
dc.contributor.authorLattes.fl_str_mv |
http://lattes.cnpq.br/8817867478588076 |
dc.contributor.author.fl_str_mv |
Peixoto, Maristela Peckle |
contributor_str_mv |
Massard, Carlos Luiz Santos, Huarrisson Azevedo Pires, Marcus Sandes Massard, Carlos Luiz Leite, Rom?rio Cerqueira Labruna, Marcelo Bahia Dias, Roberto J?nio Pedroso Santos, Tiago Marques dos |
dc.subject.por.fl_str_mv |
Diversidade gen?tica piroplasmose equina theileriose equina transmiss?o transestadial transmiss?o transovariana |
topic |
Diversidade gen?tica piroplasmose equina theileriose equina transmiss?o transestadial transmiss?o transovariana Genetic diversity equine piroplasmosis equine teileriosis transestadial transmission transovarian transmission Medicina Veterin?ria |
dc.subject.eng.fl_str_mv |
Genetic diversity equine piroplasmosis equine teileriosis transestadial transmission transovarian transmission |
dc.subject.cnpq.fl_str_mv |
Medicina Veterin?ria |
description |
This study aimed to perform the molecular characterization of the B. caballi rap-1 gene and to analyze the degree of heterogeneity of the T. equi 18S rDNA gene of naturally infected horses in the state of Rio de Janeiro. Complete sequences of the T. equi 18S rDNA gene (n = 20) were obtained. The PCR products were cloned and sequenced. Phylogenetic analyzes were performed on ARB-Silva basis. For the molecular characterization of a fragment of the B. caballi rap-1 gene, 32 cELISA and qPCR positive equine samples were used. A nested PCR was developed to amplify a fragment of the rap-1 gene and a conventional PCR, aiming the amplification of the complete gene. The products were sequenced. Sequences were submitted to the Maximum-Likelihood reconstruction method with Poisson model and 1000 replicates. In the analysis of T. equi, twelve different isolates were observed in equines from the state of Rio de Janeiro and were grouped into two of the three clades formed (I, II), most of them grouped in clade I (70%, n = 14/20). The sequences of T. equi are grouped within a clade different from other Theilerids. It was possible to amplify a fragment of the B. caballi rap-1 gene from 23 sequences using nested PCR, identified as single sequences. One of the sequences showed twelve mutation points. In cluster analysis, the Brazilian samples belonged to clade A, being analyzed as amino acids (152a) or nucleotides (1244 bp). The aim of the second chapter was to evaluate the dinamic of infection of T. equi in R. (B.) microplus fed in chronically infected horses. The larvae obtained from a hemoparasite-free colony were used in the infestation of an equine chronically infected by T. equi. Ticks and the horse blood were collected every two days. Ticks were dissected and stored in pools according to the pos-infestation day, evolutionary stage and gender. Some of these specimens were stored entire. Salivary gland and ovary samples were stored properly for Transmission Electron Microscopy. Some stages, prior to ecdysis, were stored in B.O.D. to molt. Samples were tested by qPCR. Regarding to entire tick samples, more than 90% of the samples were positive for T. equi. The mean of parasite load in entire ticks and salivary glands presented gradual increasing with the progress of the feeding period. Of the 45 salivary gland samples, 77.78% were positive for T. equi and had a higher infection rate at the end of the experimental period. It was found freshly molted ticks positives for T. equi by qPCR, indicating transestadial transmission. 84,38% of R. (B.) microplus ovary samples were considered positives for T. equi by molecular tools. Males of R. (B.) microplus were found parasiting the equine for 60 days. This fact associated with the high positivity for T. equi indicates males as an epidemiologically important gender. Transmission micrographs revealed morphologic structures compatible with a Piroplasmid in the acini of salivary glands and ovaries. The phylogenetic analysis of the T. equi 18S rDNA gene and of B. caballi rap-1 gene showed that different isolates co-exist in the same area in Rio de Janeiro, Brazil. Experimently, it was possible to observe that R. (B.) microplus could be infected with T. equi and that the forms of development of this parasite are capable of multiply inside salivary glands and, possibly, ovaries. |
publishDate |
2017 |
dc.date.issued.fl_str_mv |
2017-02-22 |
dc.date.accessioned.fl_str_mv |
2019-01-31T18:26:42Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/doctoralThesis |
format |
doctoralThesis |
status_str |
publishedVersion |
dc.identifier.citation.fl_str_mv |
PEIXOTO, Maristela Peckle. Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus. 2017. 133 f. Tese (Doutorado em Ci?ncias Veterin?rias). Instituto de Veterin?ria, Departamento de Parasitologia Animal, Universidade Federal Rural do Rio de Janeiro, Serop?dica, RJ, 2017. |
dc.identifier.uri.fl_str_mv |
https://tede.ufrrj.br/jspui/handle/jspui/2587 |
identifier_str_mv |
PEIXOTO, Maristela Peckle. Agentes da Piroplasmose equina: variabilidade gen?tica de Theileria equi e Babesia caballi em equinos naturalmente infectados do estado do Rio de Janeiro e a din?mica da infec??o de Theileria equi em Rhipicephalus (Boophilus) microplus. 2017. 133 f. Tese (Doutorado em Ci?ncias Veterin?rias). Instituto de Veterin?ria, Departamento de Parasitologia Animal, Universidade Federal Rural do Rio de Janeiro, Serop?dica, RJ, 2017. |
url |
https://tede.ufrrj.br/jspui/handle/jspui/2587 |
dc.language.iso.fl_str_mv |
por |
language |
por |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Universidade Federal Rural do Rio de Janeiro |
dc.publisher.program.fl_str_mv |
Programa de P?s-Gradua??o em Ci?ncias Veterin?rias |
dc.publisher.initials.fl_str_mv |
UFRRJ |
dc.publisher.country.fl_str_mv |
Brasil |
dc.publisher.department.fl_str_mv |
Instituto de Veterin?ria |
publisher.none.fl_str_mv |
Universidade Federal Rural do Rio de Janeiro |
dc.source.none.fl_str_mv |
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Universidade Federal Rural do Rio de Janeiro (UFRRJ) |
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UFRRJ |
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UFRRJ |
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Biblioteca Digital de Teses e Dissertações da UFRRJ |
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