Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis

Detalhes bibliográficos
Autor(a) principal: Campos-galvão, Maria Emilene Martino
Data de Publicação: 2012
Tipo de documento: Tese
Idioma: por
Título da fonte: LOCUS Repositório Institucional da UFV
Texto Completo: http://locus.ufv.br/handle/123456789/1568
Resumo: Quorum sensing (QS) is a population density dependent communication system where bacteria produce, detect and respond to signaling compounds of low molecular weight, called autoinducers (AI), which are released into the environment. Salmonella has QS systems regulated by three different AIs: AI-1, AI-2, and AI-3. In this study, we identified and characterized the QS systems mediated by AI-1 and 2 in the human pathogen Salmonella enterica serovar Enteritidis PT4 578, isolated from chicken carcasses. Analyzes were performed in vitro and in silico. We carried out a growth curve of Salmonella Enteritidis in the presence of synthetic AI-1: C6-HSL, C8-HSL, C10-HSL and C12-HSL, and we confirmed that AI-1 do not interfere in the growth. The confirmation that Salmonella Enteritidis is not able to synthesize AI-1 came from TLC. Analysis based on the genome of Salmonella Enteritidis PT4, previously deposited in the GenBank, and the described QS system for Salmonella enterica serovar typhimurium we proposed a model for QS regulation mediated by AI-2 for strain PT4 578. Analysis of the nucleotide sequence that encodes SdiA, the regulatory protein of QS mediated by AI-1, demonstrates the conservation of sdiA in this serovar. The knowledge of phenotypes regulated by QS system may contribute to elucidate important processes in food microbiology and to development control strategies for pathogenic and spoilage microorganism. Therefore we evaluated the biofilm formation by Salmonella on polystyrene microtiter plates in the presence of AI-1 (C6-HSL, C8-HSL, C10-HSL and C12-HSL) at final concentration of 50 nM in the culture medium. It was found that in the presence of AIs and on anaerobic conditions, Salmonella Enteritidis showed a higher number of cells adhered on polystyrene surfaces. Among the AHLs analyzed, C12-HSL showed greater interference with this phenotype. Strong biofilm formation in the presence of C12-HSL was also observed on polystyrene surface by scanning electron microscopy and confocal microscopy. We performed tests to evaluate the interference of AIs in combination. In an attempt to verify the regulation biofilm formation, by AI-1 we added a mix of furanones, wich are know QS inhibitors, and a reduced number of adhered cells was observed. Using Real time polymerase chain reaction analysis, it was found that C12-HSL increased the expression of the genes lpfA, fimF, fliF, glgC, wich are important for biofilm formation as well as the hilA, invA and invF virulence genes. These results demonstrate for the first time the function of SdiA for detection of signalling molecules produced by bacteria in the modulation of important phenotypes for the survival of Salmonella Enteritidis.
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spelling Campos-galvão, Maria Emilene Martinohttp://lattes.cnpq.br/0138653182683527Araujo, Elza Fernandes dehttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4783675E2Ribon, Andréa de Oliveira Barroshttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4727026E6Vanetti, Maria Cristina Dantashttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4783874H3Fantuzzi, Elisabetehttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4761222A8Santos, Miriam Teresinha doshttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4786872Z0Pinto, Uelinton Manoelhttp://lattes.cnpq.br/44169354056003002015-03-26T12:51:01Z2015-02-062015-03-26T12:51:01Z2012-06-25CAMPOS-GALVÃO, Maria Emilene Martino. Characterization of the signaling mechanism by quorum sensing in Salmonella enterica sorovar Enteritidis. 2012. 80 f. Tese (Doutorado em Associações micorrízicas; Bactérias láticas e probióticos; Biologia molecular de fungos de interesse) - Universidade Federal de Viçosa, Viçosa, 2012.http://locus.ufv.br/handle/123456789/1568Quorum sensing (QS) is a population density dependent communication system where bacteria produce, detect and respond to signaling compounds of low molecular weight, called autoinducers (AI), which are released into the environment. Salmonella has QS systems regulated by three different AIs: AI-1, AI-2, and AI-3. In this study, we identified and characterized the QS systems mediated by AI-1 and 2 in the human pathogen Salmonella enterica serovar Enteritidis PT4 578, isolated from chicken carcasses. Analyzes were performed in vitro and in silico. We carried out a growth curve of Salmonella Enteritidis in the presence of synthetic AI-1: C6-HSL, C8-HSL, C10-HSL and C12-HSL, and we confirmed that AI-1 do not interfere in the growth. The confirmation that Salmonella Enteritidis is not able to synthesize AI-1 came from TLC. Analysis based on the genome of Salmonella Enteritidis PT4, previously deposited in the GenBank, and the described QS system for Salmonella enterica serovar typhimurium we proposed a model for QS regulation mediated by AI-2 for strain PT4 578. Analysis of the nucleotide sequence that encodes SdiA, the regulatory protein of QS mediated by AI-1, demonstrates the conservation of sdiA in this serovar. The knowledge of phenotypes regulated by QS system may contribute to elucidate important processes in food microbiology and to development control strategies for pathogenic and spoilage microorganism. Therefore we evaluated the biofilm formation by Salmonella on polystyrene microtiter plates in the presence of AI-1 (C6-HSL, C8-HSL, C10-HSL and C12-HSL) at final concentration of 50 nM in the culture medium. It was found that in the presence of AIs and on anaerobic conditions, Salmonella Enteritidis showed a higher number of cells adhered on polystyrene surfaces. Among the AHLs analyzed, C12-HSL showed greater interference with this phenotype. Strong biofilm formation in the presence of C12-HSL was also observed on polystyrene surface by scanning electron microscopy and confocal microscopy. We performed tests to evaluate the interference of AIs in combination. In an attempt to verify the regulation biofilm formation, by AI-1 we added a mix of furanones, wich are know QS inhibitors, and a reduced number of adhered cells was observed. Using Real time polymerase chain reaction analysis, it was found that C12-HSL increased the expression of the genes lpfA, fimF, fliF, glgC, wich are important for biofilm formation as well as the hilA, invA and invF virulence genes. These results demonstrate for the first time the function of SdiA for detection of signalling molecules produced by bacteria in the modulation of important phenotypes for the survival of Salmonella Enteritidis.Quorum sensing (QS) é um sistema de comunicação entre bactérias dependente da densidade populacional, no qual as bactérias produzem, detectam e respondem a compostos sinalizadores de baixa massa molecular, os autoindutores (AIs). Salmonella apresenta sistemas de QS regulados por três diferentes AIs; AI-1, AI-2 e AI-3, que têm função ainda pouco elucidada. Neste trabalho foram identificados e caracterizados os sistemas de QS mediados pelos AIs 1 e 2 no patógeno humano Salmonella enterica sorovar Enteritidis PT4 578, isolado de carcaça de frango. As análises foram feitas in silico e in vitro. Foi acompanhado o crescimento de Salmonella Enteritidis na presença dos AIs-1sintéticos, as N-acil homoserina lactonas (AHLs): C6-HSL (homoserina lactona), C8-HSL, C10-HSL e C12-HSL e verificou-se que esses AIs não interferiram no crescimento deste patógeno. A confirmação de que Salmonella Enteritidis não sintetiza AI-1 foi feita por análise em cromatografia em camada fina. A partir do genoma de Salmonella Enteritidis PT4, previamente depositado no GenBank, foi confirmado que a regulação do QS mediado pelo AI-2 segue o modelo proposto para Salmonella enterica sorovar Typhimurium. A análise da sequência de nucleotídeos codificadora da proteína SdiA, reguladora do QS mediado pelo AI-1, demonstrou a conservação do gene sdiA neste sorovar. O conhecimento de fenótipos regulados pelo sistema QS pode contribuir para a elucidação de processos importantes na microbiologia de alimentos e permitir a elaboração de estratégias de controle deste patógeno. Portanto, foi avaliada a formação de biofilmes em microplaca de poliestireno por Salmonella na presença de AIs-1 (C6- HSL, C8-HSL, C10-HSL e C12-HSL) na concentração final de 50 nM no meio de cultivo. Foi constatado que, na presença de Ais e, em condições de cultivo em anaerobiose, Salmonella Enteritidis apresentou formação de biofilme mais denso e compacto em superfícies de poliestireno. A C12-HSL foi, dentre as AHLs analisadas, a que apresentou maior interferência nesse fenótipo. A formação de biofilme mais intensa na presença de C12-HSL em superfície de poliestireno também foi constatada por microscopia de varredura e microscopia confocal. Para comprovar a regulação do fenótipo de formação de biofilme pelo AI-1, o experimento foi repetido na presença de uma mistura de diferentes furanonas, inibidores de QS. A redução substancial de células aderidas na presença de furanonas confirma o efeito estimulador dos AIs-1 na formação de biofilmes por Salmonella Enteritidis PT4. Por meio da reação em cadeia da polimerase (PCR) em tempo real, foi comprovado que C12-HSL promoveu maior expressão dos genes lpfA, fimF, fliF e glgC, de importância para a formação de biofilmes, assim como, dos genes de virulência hilA, invA e invF. Estes resultados evidenciam, pela primeira vez, a função da SdiA na detecção de moléculas sinais produzidas por outras bactérias gram-negativas na modulação de fenótipos importantes para a sobrevivência e virulência de Salmonella Enteritidis.Conselho Nacional de Desenvolvimento Científico e Tecnológicoapplication/pdfporUniversidade Federal de ViçosaDoutorado em Microbiologia AgrícolaUFVBRAssociações micorrízicas; Bactérias láticas e probióticos; Biologia molecular de fungos de interesseSalmonella entericaComunicação celularBiofilmesAlimentos - MicrobiologiaSalmonella entericaCell communicationBiofilmsFood - MicrobiologyCNPQ::CIENCIAS AGRARIAS::CIENCIA E TECNOLOGIA DE ALIMENTOSCaracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar EnteritidisCharacterization of the signaling mechanism by quorum sensing in Salmonella enterica sorovar Enteritidisinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/embargoedAccessreponame:LOCUS Repositório Institucional da UFVinstname:Universidade Federal de Viçosa (UFV)instacron:UFVORIGINALtexto completo.pdfapplication/pdf1456260https://locus.ufv.br//bitstream/123456789/1568/1/texto%20completo.pdf50b25a62ec7395c28d37c9ce92c65e18MD51TEXTtexto completo.pdf.txttexto completo.pdf.txtExtracted texttext/plain161251https://locus.ufv.br//bitstream/123456789/1568/2/texto%20completo.pdf.txt0f286a6f14a676b2a5eb279ce39bb4e6MD52THUMBNAILtexto completo.pdf.jpgtexto completo.pdf.jpgIM Thumbnailimage/jpeg3608https://locus.ufv.br//bitstream/123456789/1568/3/texto%20completo.pdf.jpge3e8cca3ebd1c47f5d864c4ef16474d2MD53123456789/15682016-04-07 23:05:27.211oai:locus.ufv.br:123456789/1568Repositório InstitucionalPUBhttps://www.locus.ufv.br/oai/requestfabiojreis@ufv.bropendoar:21452016-04-08T02:05:27LOCUS Repositório Institucional da UFV - Universidade Federal de Viçosa (UFV)false
dc.title.por.fl_str_mv Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
dc.title.alternative.eng.fl_str_mv Characterization of the signaling mechanism by quorum sensing in Salmonella enterica sorovar Enteritidis
title Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
spellingShingle Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
Campos-galvão, Maria Emilene Martino
Salmonella enterica
Comunicação celular
Biofilmes
Alimentos - Microbiologia
Salmonella enterica
Cell communication
Biofilms
Food - Microbiology
CNPQ::CIENCIAS AGRARIAS::CIENCIA E TECNOLOGIA DE ALIMENTOS
title_short Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
title_full Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
title_fullStr Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
title_full_unstemmed Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
title_sort Caracterização do mecanismo de sinalização por quorum sensing em Salmonella enterica sorovar Enteritidis
author Campos-galvão, Maria Emilene Martino
author_facet Campos-galvão, Maria Emilene Martino
author_role author
dc.contributor.authorLattes.por.fl_str_mv http://lattes.cnpq.br/0138653182683527
dc.contributor.author.fl_str_mv Campos-galvão, Maria Emilene Martino
dc.contributor.advisor-co1.fl_str_mv Araujo, Elza Fernandes de
dc.contributor.advisor-co1Lattes.fl_str_mv http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4783675E2
dc.contributor.advisor-co2.fl_str_mv Ribon, Andréa de Oliveira Barros
dc.contributor.advisor-co2Lattes.fl_str_mv http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4727026E6
dc.contributor.advisor1.fl_str_mv Vanetti, Maria Cristina Dantas
dc.contributor.advisor1Lattes.fl_str_mv http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4783874H3
dc.contributor.referee1.fl_str_mv Fantuzzi, Elisabete
dc.contributor.referee1Lattes.fl_str_mv http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4761222A8
dc.contributor.referee2.fl_str_mv Santos, Miriam Teresinha dos
dc.contributor.referee2Lattes.fl_str_mv http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4786872Z0
dc.contributor.referee3.fl_str_mv Pinto, Uelinton Manoel
dc.contributor.referee3Lattes.fl_str_mv http://lattes.cnpq.br/4416935405600300
contributor_str_mv Araujo, Elza Fernandes de
Ribon, Andréa de Oliveira Barros
Vanetti, Maria Cristina Dantas
Fantuzzi, Elisabete
Santos, Miriam Teresinha dos
Pinto, Uelinton Manoel
dc.subject.por.fl_str_mv Salmonella enterica
Comunicação celular
Biofilmes
Alimentos - Microbiologia
topic Salmonella enterica
Comunicação celular
Biofilmes
Alimentos - Microbiologia
Salmonella enterica
Cell communication
Biofilms
Food - Microbiology
CNPQ::CIENCIAS AGRARIAS::CIENCIA E TECNOLOGIA DE ALIMENTOS
dc.subject.eng.fl_str_mv Salmonella enterica
Cell communication
Biofilms
Food - Microbiology
dc.subject.cnpq.fl_str_mv CNPQ::CIENCIAS AGRARIAS::CIENCIA E TECNOLOGIA DE ALIMENTOS
description Quorum sensing (QS) is a population density dependent communication system where bacteria produce, detect and respond to signaling compounds of low molecular weight, called autoinducers (AI), which are released into the environment. Salmonella has QS systems regulated by three different AIs: AI-1, AI-2, and AI-3. In this study, we identified and characterized the QS systems mediated by AI-1 and 2 in the human pathogen Salmonella enterica serovar Enteritidis PT4 578, isolated from chicken carcasses. Analyzes were performed in vitro and in silico. We carried out a growth curve of Salmonella Enteritidis in the presence of synthetic AI-1: C6-HSL, C8-HSL, C10-HSL and C12-HSL, and we confirmed that AI-1 do not interfere in the growth. The confirmation that Salmonella Enteritidis is not able to synthesize AI-1 came from TLC. Analysis based on the genome of Salmonella Enteritidis PT4, previously deposited in the GenBank, and the described QS system for Salmonella enterica serovar typhimurium we proposed a model for QS regulation mediated by AI-2 for strain PT4 578. Analysis of the nucleotide sequence that encodes SdiA, the regulatory protein of QS mediated by AI-1, demonstrates the conservation of sdiA in this serovar. The knowledge of phenotypes regulated by QS system may contribute to elucidate important processes in food microbiology and to development control strategies for pathogenic and spoilage microorganism. Therefore we evaluated the biofilm formation by Salmonella on polystyrene microtiter plates in the presence of AI-1 (C6-HSL, C8-HSL, C10-HSL and C12-HSL) at final concentration of 50 nM in the culture medium. It was found that in the presence of AIs and on anaerobic conditions, Salmonella Enteritidis showed a higher number of cells adhered on polystyrene surfaces. Among the AHLs analyzed, C12-HSL showed greater interference with this phenotype. Strong biofilm formation in the presence of C12-HSL was also observed on polystyrene surface by scanning electron microscopy and confocal microscopy. We performed tests to evaluate the interference of AIs in combination. In an attempt to verify the regulation biofilm formation, by AI-1 we added a mix of furanones, wich are know QS inhibitors, and a reduced number of adhered cells was observed. Using Real time polymerase chain reaction analysis, it was found that C12-HSL increased the expression of the genes lpfA, fimF, fliF, glgC, wich are important for biofilm formation as well as the hilA, invA and invF virulence genes. These results demonstrate for the first time the function of SdiA for detection of signalling molecules produced by bacteria in the modulation of important phenotypes for the survival of Salmonella Enteritidis.
publishDate 2012
dc.date.issued.fl_str_mv 2012-06-25
dc.date.accessioned.fl_str_mv 2015-03-26T12:51:01Z
dc.date.available.fl_str_mv 2015-02-06
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dc.identifier.citation.fl_str_mv CAMPOS-GALVÃO, Maria Emilene Martino. Characterization of the signaling mechanism by quorum sensing in Salmonella enterica sorovar Enteritidis. 2012. 80 f. Tese (Doutorado em Associações micorrízicas; Bactérias láticas e probióticos; Biologia molecular de fungos de interesse) - Universidade Federal de Viçosa, Viçosa, 2012.
dc.identifier.uri.fl_str_mv http://locus.ufv.br/handle/123456789/1568
identifier_str_mv CAMPOS-GALVÃO, Maria Emilene Martino. Characterization of the signaling mechanism by quorum sensing in Salmonella enterica sorovar Enteritidis. 2012. 80 f. Tese (Doutorado em Associações micorrízicas; Bactérias láticas e probióticos; Biologia molecular de fungos de interesse) - Universidade Federal de Viçosa, Viçosa, 2012.
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