Orf virus como plataforma para vacinas vetoriais para suínos e bovinos
Autor(a) principal: | |
---|---|
Data de Publicação: | 2018 |
Tipo de documento: | Tese |
Idioma: | por |
Título da fonte: | Manancial - Repositório Digital da UFSM |
Texto Completo: | http://repositorio.ufsm.br/handle/1/14083 |
Resumo: | Parapoxvirus ovis (PPVO) or orf virus (ORFV) is the causative agent of contagious ecthyma (or orf), a mucocutaneous disease that affects mainly sheep and goats. The ORFV belongs to the family Poxviridae, subfamily Chordopoxvirinae and genus Parapoxvirus. The virions are enveloped and their genome consist of a linear, double stranded DNA molecule of approximately 138 kb and contains at least 131 genes. ORFV has several biological and genomic properties that make it an attractive candidate for a vaccine vector. Therefore, it has been proposed as a vaccine platform to carry heterologous genes of veterinary interest. The first study reports the construction and characterization of two ORFV recombinants out of the parental strain ORFV IA82, expressing the rabies virus glycoprotein G (RABVG) at the loci of ORFV024 or ORFV121 genes, respectively, and an evaluation of their immunogenicity in swine and cattle. The in vitro characterization showed that the recombinants ORFVΔ024RABV-G and ORFVΔ121RABV-G retained their replicative capacity in cell culture, compared to the parental virus. The recombinant viruses expressed RABV-G efficiently even after 10 passages in cell culture, demonstrating the stability of the inserts. Immunization of swine and cattle with ORFVΔ024RABV-G and ORFVΔ121RABV-G resulted in a robust neutralizing antibody response against the rabies virus (RABV). The neutralizing antibodies titers induced by ORFVΔ121RABV-G in swine and cattle were higher than those induced ORFVΔ024RABV-G, indicating a higher efficiency of ORFVΔ121 as a vector in these species. In a second study, ORFV mutants with individual deletions of genes ORFV112, ORFV117 or ORFV127 were constructed, characterized in vitro and inoculated experimentally in lambs. When characterized in vitro, the deletion mutants replicated in ovine fetal turbinate (OFTu) cells with the same efficiency of the parental virus, with no changes in replication kinetics, plaque size and morphology. Experimental inoculation of lambs at the mucocutaneous junction of the oral commissure demonstrated that individual deletions of ORFV112, ORFV117 or ORFV127 genes did not result in obvious changes in virulence, since the mutants produced lesions as severe as those induced by the parental virus. However, resolution of the lesions apparently occurred more rapidly in the animals inoculated with the mutant virus than in those inoculated with the parental virus. In addition, the mutant viruses were excreted from the lesions in lower titers than those of the parental virus. These results demonstrated that genes ORFV112, ORFV117 and ORFV127 are not essential for viability of ORFV in vitro and interfere only partially in ORFV virulence in lambs. A third study was conducted to compare the magnitude and duration of the serological response induced in cattle by four commercial vaccines against RABV. After two vaccinations 30-days apart, the animals received a booster vaccination one year later, and were subjected to serological tests for virus neutralizing (VN) antibodies at different intervals after vaccination and booster. The four vaccines were found to be suitably immunogenic, e.g., induced high titers of VN antibodies in the animals after primary vaccination. However, a marked decrease in antibody levels was observed prior to the annual booster. Therefore, it is recommended that annual revaccination protocols be reviewed, since part of the vaccinated animals no longer have adequate antibodies levels one year after vaccination. In summary, the experiments presented in this thesis demonstrate that ORFV represents a promising platform for vectoring vaccine antigens in swine and cattle and that, in addition to the ORFV024 and ORFV121 genes, loci ORFV112, ORF117 and ORFV127 can also be used for heterologous gene insertion. |
id |
UFSM-20_39669feda5b325bb74be269c152b7b79 |
---|---|
oai_identifier_str |
oai:repositorio.ufsm.br:1/14083 |
network_acronym_str |
UFSM-20 |
network_name_str |
Manancial - Repositório Digital da UFSM |
repository_id_str |
3913 |
spelling |
2018-08-23T13:10:22Z2018-08-23T13:10:22Z2018-02-26http://repositorio.ufsm.br/handle/1/14083Parapoxvirus ovis (PPVO) or orf virus (ORFV) is the causative agent of contagious ecthyma (or orf), a mucocutaneous disease that affects mainly sheep and goats. The ORFV belongs to the family Poxviridae, subfamily Chordopoxvirinae and genus Parapoxvirus. The virions are enveloped and their genome consist of a linear, double stranded DNA molecule of approximately 138 kb and contains at least 131 genes. ORFV has several biological and genomic properties that make it an attractive candidate for a vaccine vector. Therefore, it has been proposed as a vaccine platform to carry heterologous genes of veterinary interest. The first study reports the construction and characterization of two ORFV recombinants out of the parental strain ORFV IA82, expressing the rabies virus glycoprotein G (RABVG) at the loci of ORFV024 or ORFV121 genes, respectively, and an evaluation of their immunogenicity in swine and cattle. The in vitro characterization showed that the recombinants ORFVΔ024RABV-G and ORFVΔ121RABV-G retained their replicative capacity in cell culture, compared to the parental virus. The recombinant viruses expressed RABV-G efficiently even after 10 passages in cell culture, demonstrating the stability of the inserts. Immunization of swine and cattle with ORFVΔ024RABV-G and ORFVΔ121RABV-G resulted in a robust neutralizing antibody response against the rabies virus (RABV). The neutralizing antibodies titers induced by ORFVΔ121RABV-G in swine and cattle were higher than those induced ORFVΔ024RABV-G, indicating a higher efficiency of ORFVΔ121 as a vector in these species. In a second study, ORFV mutants with individual deletions of genes ORFV112, ORFV117 or ORFV127 were constructed, characterized in vitro and inoculated experimentally in lambs. When characterized in vitro, the deletion mutants replicated in ovine fetal turbinate (OFTu) cells with the same efficiency of the parental virus, with no changes in replication kinetics, plaque size and morphology. Experimental inoculation of lambs at the mucocutaneous junction of the oral commissure demonstrated that individual deletions of ORFV112, ORFV117 or ORFV127 genes did not result in obvious changes in virulence, since the mutants produced lesions as severe as those induced by the parental virus. However, resolution of the lesions apparently occurred more rapidly in the animals inoculated with the mutant virus than in those inoculated with the parental virus. In addition, the mutant viruses were excreted from the lesions in lower titers than those of the parental virus. These results demonstrated that genes ORFV112, ORFV117 and ORFV127 are not essential for viability of ORFV in vitro and interfere only partially in ORFV virulence in lambs. A third study was conducted to compare the magnitude and duration of the serological response induced in cattle by four commercial vaccines against RABV. After two vaccinations 30-days apart, the animals received a booster vaccination one year later, and were subjected to serological tests for virus neutralizing (VN) antibodies at different intervals after vaccination and booster. The four vaccines were found to be suitably immunogenic, e.g., induced high titers of VN antibodies in the animals after primary vaccination. However, a marked decrease in antibody levels was observed prior to the annual booster. Therefore, it is recommended that annual revaccination protocols be reviewed, since part of the vaccinated animals no longer have adequate antibodies levels one year after vaccination. In summary, the experiments presented in this thesis demonstrate that ORFV represents a promising platform for vectoring vaccine antigens in swine and cattle and that, in addition to the ORFV024 and ORFV121 genes, loci ORFV112, ORF117 and ORFV127 can also be used for heterologous gene insertion.O Parapoxvirus ovis (PPVO) ou vírus da orf (ORFV) é o agente do ectima contagioso (ou orf), uma enfermidade mucocutânea que afeta principalmente ovinos e caprinos. O ORFV pertence à família Poxviridae, subfamília Chordopoxvirinae e gênero Parapoxvirus. Os vírions possuem envelope e o genoma consiste de uma molécula de DNA linear e de fita dupla, com aproximadamente 138 quilobases (kb) e contém pelo menos 131 genes. O ORFV possui várias propriedades biológicas e genômicas que o tornam um atraente candidato a vetor vacinal. Por isso, tem sido proposto como plataforma vacinal para vetorar genes heterólogos de interesse veterinário. Em um primeiro estudo, descreve-se a construção e caracterização de dois recombinantes do ORFV, a partir da cepa parental ORFV IA82, que expressam a glicoproteína G do vírus da raiva (RABV-G) nos loci dos genes ORFV024 ou ORFV121, respectivamente, e a avaliação da sua imunogenicidade em suínos e bovinos. A caracterização in vitro demonstrou que os recombinantes ORFVΔ024RABV-G e ORFVΔ121RABV-G não apresentaram alterações na capacidade replicativa em cultivo celular, comparando-se ao vírus parental. Foi demonstrado que os recombinantes expressam a RABV-G com eficiência, mesmo após 10 passagens em cultivo celular, demonstrando a estabilidade dessas inserções. A imunização de suínos e bovinos com o ORFVΔ024RABV-G e ORFVΔ121RABV-G resultou em resposta robusta de anticorpos neutralizantes contra o vírus da raiva (RABV). Os títulos de anticorpos induzidos pelo recombinante ORFVΔ121RABV-G em suínos e bovinos foram superiores aos induzidos pelo ORFVΔ024RABV-G, indicando uma maior eficiência do recombinante ORFVΔ121 como vetor nessas espécies. Em um segundo estudo, mutantes do ORFV com deleções individuais nos genes ORFV112, ORFV117 ou ORFV127 foram construídos, caracterizados in vitro e inoculados experimentalmente em cordeiros. Quando caracterizados in vitro, os mutantes de deleção replicaram em células de corneto etmoidal ovino (OFTu) com a mesma eficiência do vírus parental, sem alterações na cinética de replicação, tamanho e morfologia de placas virais. A inoculação experimental de cordeiros na junção mucocutânea da comissura oral demonstrou que as deleções individuais dos genes ORFV112, ORFV117 ou ORFV127 não resultaram em alterações evidentes de virulência, pois os mutantes produziram lesões tão severas quanto as induzidas pelo vírus parental. No entanto, a resolução das lesões aparentemente ocorreu de forma mais rápida nos animais inoculados com os vírus mutantes do que naqueles inoculados com o vírus parental. Além disso, os vírus mutantes foram excretados das lesões em títulos inferiores aos do vírus parental. Estes resultados demonstraram que os genes ORFV112, ORFV117 e ORFV127 não são essenciais para a viabilidade do ORFV in vitro ou in vivo e interferem apenas parcialmente na virulência em ovinos. Um terceiro estudo foi conduzido para comparar a magnitude e duração da resposta sorológica induzida em bovinos por quatro vacinas comerciais contra o RABV. Após duas vacinações com intervalo de 30 dias, os animais receberam reforço vacinal um ano após, sendo submetidos a pesquisa e quantificação de anticorpos neutralizantes a diferentes intervalos após a vacinação e reforço. Verificou-se que as quatro vacinas são adequadamente imunogênicas, ou seja, induziram altos títulos de anticorpos neutralizantes nos animais após a primovacinação. Porém, verificou-se um decréscimo acentuado nos níveis de anticorpos antes do reforço, de forma que vários animais já não apresentavam níveis adequados de anticorpos por ocasião do reforço. Assim, recomenda-se que os protocolos de revacinação anual sejam revistos, antecipando-se a data prevista para reforçar a vacinação. Em resumo, os experimentos apresentados na presente tese demonstram que o ORFV representa uma plataforma promissora para vetorar antígenos vacinais em suínos e bovinos e que, além dos genes ORFV024 e ORFV121, os loci ORFV112, ORFV117 e ORFV127 também podem ser utilizados para a inserção de genes heterólogos de interesse.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPESporUniversidade Federal de Santa MariaCentro de Ciências RuraisPrograma de Pós-Graduação em Medicina VeterináriaUFSMBrasilMedicina VeterináriaAttribution-NonCommercial-NoDerivatives 4.0 Internationalhttp://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/openAccessOrf vírus recombinanteVacina recombinantePatogenicidadeRaivaRecombinant orf virusRecombinant vaccinePathogenicityRabiesCNPQ::CIENCIAS AGRARIAS::MEDICINA VETERINARIAOrf virus como plataforma para vacinas vetoriais para suínos e bovinosOrf virus as a platform for vector vaccines for swine and cattleinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesisWeiblen, Rudihttp://lattes.cnpq.br/7946350215388090Diel, Diego Gustavohttp://lattes.cnpq.br/0783755116551811Cargnelutti, Juliana Felipettohttp://lattes.cnpq.br/5180338810182471Brum, Mário Celso Sperottohttp://lattes.cnpq.br/9761857774819478Lima, Marcelo dehttp://lattes.cnpq.br/1435610308662740http://lattes.cnpq.br/2726377135890208Martins, Mathias50050000000760002776f4b-7154-4d1a-9f55-d68a031b94bc88e3e298-87c8-4195-a90e-220eee184bec5de97f57-5591-4503-81ef-a682eeb937c89a82a3b4-b1a0-4cbc-a69f-1b4d58dd0cb3b64de51e-e56f-48d7-b038-84dc2dec314bcc49bcf9-df7b-4e2e-8cdc-fd1ad6226e3breponame:Manancial - Repositório Digital da UFSMinstname:Universidade Federal de Santa Maria (UFSM)instacron:UFSMORIGINALTES_PPGMV_2018_MARTINS_MATHIAS.pdfTES_PPGMV_2018_MARTINS_MATHIAS.pdfTese de Doutoradoapplication/pdf7608313http://repositorio.ufsm.br/bitstream/1/14083/1/TES_PPGMV_2018_MARTINS_MATHIAS.pdf0e5a32ae66273a99b0c0d9a485bb585cMD51CC-LICENSElicense_rdflicense_rdfapplication/rdf+xml; charset=utf-8804http://repositorio.ufsm.br/bitstream/1/14083/2/license_rdfc1efe8e24d7281448e873be30ea326ffMD52LICENSElicense.txtlicense.txttext/plain; charset=utf-81956http://repositorio.ufsm.br/bitstream/1/14083/3/license.txt2f0571ecee68693bd5cd3f17c1e075dfMD53TEXTTES_PPGMV_2018_MARTINS_MATHIAS.pdf.txtTES_PPGMV_2018_MARTINS_MATHIAS.pdf.txtExtracted texttext/plain201137http://repositorio.ufsm.br/bitstream/1/14083/4/TES_PPGMV_2018_MARTINS_MATHIAS.pdf.txt5a88a5995bd716f953f343ebb5c9e584MD54THUMBNAILTES_PPGMV_2018_MARTINS_MATHIAS.pdf.jpgTES_PPGMV_2018_MARTINS_MATHIAS.pdf.jpgIM Thumbnailimage/jpeg4308http://repositorio.ufsm.br/bitstream/1/14083/5/TES_PPGMV_2018_MARTINS_MATHIAS.pdf.jpgc6a327b41d2915bed6a1da3c584d7606MD551/140832018-08-23 10:10:22.687oai:repositorio.ufsm.br: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ório Institucionalhttp://repositorio.ufsm.br/PUBhttp://repositorio.ufsm.br/oai/requestopendoar:39132018-08-23T13:10:22Manancial - Repositório Digital da UFSM - Universidade Federal de Santa Maria (UFSM)false |
dc.title.por.fl_str_mv |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
dc.title.alternative.eng.fl_str_mv |
Orf virus as a platform for vector vaccines for swine and cattle |
title |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
spellingShingle |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos Martins, Mathias Orf vírus recombinante Vacina recombinante Patogenicidade Raiva Recombinant orf virus Recombinant vaccine Pathogenicity Rabies CNPQ::CIENCIAS AGRARIAS::MEDICINA VETERINARIA |
title_short |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
title_full |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
title_fullStr |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
title_full_unstemmed |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
title_sort |
Orf virus como plataforma para vacinas vetoriais para suínos e bovinos |
author |
Martins, Mathias |
author_facet |
Martins, Mathias |
author_role |
author |
dc.contributor.advisor1.fl_str_mv |
Weiblen, Rudi |
dc.contributor.advisor1Lattes.fl_str_mv |
http://lattes.cnpq.br/7946350215388090 |
dc.contributor.referee1.fl_str_mv |
Diel, Diego Gustavo |
dc.contributor.referee1Lattes.fl_str_mv |
http://lattes.cnpq.br/0783755116551811 |
dc.contributor.referee2.fl_str_mv |
Cargnelutti, Juliana Felipetto |
dc.contributor.referee2Lattes.fl_str_mv |
http://lattes.cnpq.br/5180338810182471 |
dc.contributor.referee3.fl_str_mv |
Brum, Mário Celso Sperotto |
dc.contributor.referee3Lattes.fl_str_mv |
http://lattes.cnpq.br/9761857774819478 |
dc.contributor.referee4.fl_str_mv |
Lima, Marcelo de |
dc.contributor.referee4Lattes.fl_str_mv |
http://lattes.cnpq.br/1435610308662740 |
dc.contributor.authorLattes.fl_str_mv |
http://lattes.cnpq.br/2726377135890208 |
dc.contributor.author.fl_str_mv |
Martins, Mathias |
contributor_str_mv |
Weiblen, Rudi Diel, Diego Gustavo Cargnelutti, Juliana Felipetto Brum, Mário Celso Sperotto Lima, Marcelo de |
dc.subject.por.fl_str_mv |
Orf vírus recombinante Vacina recombinante Patogenicidade Raiva |
topic |
Orf vírus recombinante Vacina recombinante Patogenicidade Raiva Recombinant orf virus Recombinant vaccine Pathogenicity Rabies CNPQ::CIENCIAS AGRARIAS::MEDICINA VETERINARIA |
dc.subject.eng.fl_str_mv |
Recombinant orf virus Recombinant vaccine Pathogenicity Rabies |
dc.subject.cnpq.fl_str_mv |
CNPQ::CIENCIAS AGRARIAS::MEDICINA VETERINARIA |
description |
Parapoxvirus ovis (PPVO) or orf virus (ORFV) is the causative agent of contagious ecthyma (or orf), a mucocutaneous disease that affects mainly sheep and goats. The ORFV belongs to the family Poxviridae, subfamily Chordopoxvirinae and genus Parapoxvirus. The virions are enveloped and their genome consist of a linear, double stranded DNA molecule of approximately 138 kb and contains at least 131 genes. ORFV has several biological and genomic properties that make it an attractive candidate for a vaccine vector. Therefore, it has been proposed as a vaccine platform to carry heterologous genes of veterinary interest. The first study reports the construction and characterization of two ORFV recombinants out of the parental strain ORFV IA82, expressing the rabies virus glycoprotein G (RABVG) at the loci of ORFV024 or ORFV121 genes, respectively, and an evaluation of their immunogenicity in swine and cattle. The in vitro characterization showed that the recombinants ORFVΔ024RABV-G and ORFVΔ121RABV-G retained their replicative capacity in cell culture, compared to the parental virus. The recombinant viruses expressed RABV-G efficiently even after 10 passages in cell culture, demonstrating the stability of the inserts. Immunization of swine and cattle with ORFVΔ024RABV-G and ORFVΔ121RABV-G resulted in a robust neutralizing antibody response against the rabies virus (RABV). The neutralizing antibodies titers induced by ORFVΔ121RABV-G in swine and cattle were higher than those induced ORFVΔ024RABV-G, indicating a higher efficiency of ORFVΔ121 as a vector in these species. In a second study, ORFV mutants with individual deletions of genes ORFV112, ORFV117 or ORFV127 were constructed, characterized in vitro and inoculated experimentally in lambs. When characterized in vitro, the deletion mutants replicated in ovine fetal turbinate (OFTu) cells with the same efficiency of the parental virus, with no changes in replication kinetics, plaque size and morphology. Experimental inoculation of lambs at the mucocutaneous junction of the oral commissure demonstrated that individual deletions of ORFV112, ORFV117 or ORFV127 genes did not result in obvious changes in virulence, since the mutants produced lesions as severe as those induced by the parental virus. However, resolution of the lesions apparently occurred more rapidly in the animals inoculated with the mutant virus than in those inoculated with the parental virus. In addition, the mutant viruses were excreted from the lesions in lower titers than those of the parental virus. These results demonstrated that genes ORFV112, ORFV117 and ORFV127 are not essential for viability of ORFV in vitro and interfere only partially in ORFV virulence in lambs. A third study was conducted to compare the magnitude and duration of the serological response induced in cattle by four commercial vaccines against RABV. After two vaccinations 30-days apart, the animals received a booster vaccination one year later, and were subjected to serological tests for virus neutralizing (VN) antibodies at different intervals after vaccination and booster. The four vaccines were found to be suitably immunogenic, e.g., induced high titers of VN antibodies in the animals after primary vaccination. However, a marked decrease in antibody levels was observed prior to the annual booster. Therefore, it is recommended that annual revaccination protocols be reviewed, since part of the vaccinated animals no longer have adequate antibodies levels one year after vaccination. In summary, the experiments presented in this thesis demonstrate that ORFV represents a promising platform for vectoring vaccine antigens in swine and cattle and that, in addition to the ORFV024 and ORFV121 genes, loci ORFV112, ORF117 and ORFV127 can also be used for heterologous gene insertion. |
publishDate |
2018 |
dc.date.accessioned.fl_str_mv |
2018-08-23T13:10:22Z |
dc.date.available.fl_str_mv |
2018-08-23T13:10:22Z |
dc.date.issued.fl_str_mv |
2018-02-26 |
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.uri.fl_str_mv |
http://repositorio.ufsm.br/handle/1/14083 |
url |
http://repositorio.ufsm.br/handle/1/14083 |
dc.language.iso.fl_str_mv |
por |
language |
por |
dc.relation.cnpq.fl_str_mv |
500500000007 |
dc.relation.confidence.fl_str_mv |
600 |
dc.relation.authority.fl_str_mv |
02776f4b-7154-4d1a-9f55-d68a031b94bc 88e3e298-87c8-4195-a90e-220eee184bec 5de97f57-5591-4503-81ef-a682eeb937c8 9a82a3b4-b1a0-4cbc-a69f-1b4d58dd0cb3 b64de51e-e56f-48d7-b038-84dc2dec314b cc49bcf9-df7b-4e2e-8cdc-fd1ad6226e3b |
dc.rights.driver.fl_str_mv |
Attribution-NonCommercial-NoDerivatives 4.0 International http://creativecommons.org/licenses/by-nc-nd/4.0/ info:eu-repo/semantics/openAccess |
rights_invalid_str_mv |
Attribution-NonCommercial-NoDerivatives 4.0 International http://creativecommons.org/licenses/by-nc-nd/4.0/ |
eu_rights_str_mv |
openAccess |
dc.publisher.none.fl_str_mv |
Universidade Federal de Santa Maria Centro de Ciências Rurais |
dc.publisher.program.fl_str_mv |
Programa de Pós-Graduação em Medicina Veterinária |
dc.publisher.initials.fl_str_mv |
UFSM |
dc.publisher.country.fl_str_mv |
Brasil |
dc.publisher.department.fl_str_mv |
Medicina Veterinária |
publisher.none.fl_str_mv |
Universidade Federal de Santa Maria Centro de Ciências Rurais |
dc.source.none.fl_str_mv |
reponame:Manancial - Repositório Digital da UFSM instname:Universidade Federal de Santa Maria (UFSM) instacron:UFSM |
instname_str |
Universidade Federal de Santa Maria (UFSM) |
instacron_str |
UFSM |
institution |
UFSM |
reponame_str |
Manancial - Repositório Digital da UFSM |
collection |
Manancial - Repositório Digital da UFSM |
bitstream.url.fl_str_mv |
http://repositorio.ufsm.br/bitstream/1/14083/1/TES_PPGMV_2018_MARTINS_MATHIAS.pdf http://repositorio.ufsm.br/bitstream/1/14083/2/license_rdf http://repositorio.ufsm.br/bitstream/1/14083/3/license.txt http://repositorio.ufsm.br/bitstream/1/14083/4/TES_PPGMV_2018_MARTINS_MATHIAS.pdf.txt http://repositorio.ufsm.br/bitstream/1/14083/5/TES_PPGMV_2018_MARTINS_MATHIAS.pdf.jpg |
bitstream.checksum.fl_str_mv |
0e5a32ae66273a99b0c0d9a485bb585c c1efe8e24d7281448e873be30ea326ff 2f0571ecee68693bd5cd3f17c1e075df 5a88a5995bd716f953f343ebb5c9e584 c6a327b41d2915bed6a1da3c584d7606 |
bitstream.checksumAlgorithm.fl_str_mv |
MD5 MD5 MD5 MD5 MD5 |
repository.name.fl_str_mv |
Manancial - Repositório Digital da UFSM - Universidade Federal de Santa Maria (UFSM) |
repository.mail.fl_str_mv |
|
_version_ |
1794523920670392320 |