Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia
Autor(a) principal: | |
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Data de Publicação: | 2019 |
Tipo de documento: | Dissertação |
Idioma: | por |
Título da fonte: | Repositório Institucional da Universidade Federal do Espírito Santo (riUfes) |
Texto Completo: | http://repositorio.ufes.br/handle/10/10947 |
Resumo: | Introduction: Bacterial tolerance is characterized by the ability of a microbial population to survive the high concentrations of a bactericidal drug without alteration in the Minimum Inhibitory Concentration. At the end of the antimicrobial treatment, the tolerant strains multiply again, which may cause recalcitrant infections and facilitate resistance. Many studies relate antimicrobial exposure to tolerance induction. Although observed in several species, tolerance has not yet been investigated for the opportunistic bacillus Stenotrophomonas maltophilia, commonly exposed in the clinical scenario to antibiotics that have no action on this pathogen. Thus, our objective was to investigate whether the prior use of certain non-targeted antimicrobials induces cross-tolerance in S. maltophilia. Materials and methods: Qualitative and quantitative screening tests were performed to evaluate possible tolerance-inducing antimicrobials. In these assays, we exposed the S. maltophilia ATCC® 13637™ to daptomycin (DAP), vancomycin (VAN) and gentamicin (GEN) and evaluated the occurrence of cross-tolerance to the following drugs targeting S. maltophilia: sulfamethoxazole/trimethoprim (SXT), ceftazidime (CAZ) and levofloxacin (LVX). After the screening, strains with tolerance-induced characteristics were tested for genetic similarity by ERIC-PCR. To exclude the contamination hypothesis and confirm the genus and species of the strains, the MALDI-TOF MS was performed. To evaluate tolerance, the following tests were carried out: determination of the Minimum Bactericidal Concentration (MBC) / Minimum Inhibitory Concentration (MIC) ratio for LVX; adapted disk-diffusion test for SXT, CAZ, LVX, ticarcillin-clavulanate (TCC), chloramphenicol (CLO), minocycline (MIN) and polymyxin B (PXB) and time-kill curve for LVX, CAZ and ciprofloxacin (CIP). Results: After induction of the parental strain ATCC® 13637™ to DAP and VAN, three potentially tolerant strains (D25, V15.6 and V3.9) were isolated, whose genetic similarity to parental was confirmed by ERIC-PCR (100% similarity). The induced strains maintained the same MIC value of LVX as the parental strain (0.125 μg/mL), with the MBC/MIC ratio ranging from 2.64 to 3.04 among the induced strains. In the adapted disc-diffusion test, strain D25 showed high tolerance for LVX, CAZ and TCC. In the stationary phase time-kill curves against 50 times the MIC of LVX, strain D25 presented a minimum duration for killing 99,99% of all population (MDK99.99) higher than the parental. The survival rate was also higher for the D25 strain in the presence of CAZ (50xMIC). In general, strain D25 had a higher survival rate in all time-kill curves, except for CIP. Conclusions: Previous exposure of S. maltophilia to daptomycin induces cross-tolerance to levofloxacin, ceftazidime and ticarcillinclavulanate. Thus, erroneous empirical therapy with daptomycin could lead to the failure of these antimicrobials to eradicate S. maltophilia, culminating in chronic infections or death by a subpopulation of tolerant microorganisms. |
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Santos, Kênia Valéria dosFerreira, Mariana Abou MouradSilva, Rodrigo Cayô daTavares, Sarah Goncalves2019-03-15T02:11:22Z2019-03-142019-03-15T02:11:22Z2019-02-22Introduction: Bacterial tolerance is characterized by the ability of a microbial population to survive the high concentrations of a bactericidal drug without alteration in the Minimum Inhibitory Concentration. At the end of the antimicrobial treatment, the tolerant strains multiply again, which may cause recalcitrant infections and facilitate resistance. Many studies relate antimicrobial exposure to tolerance induction. Although observed in several species, tolerance has not yet been investigated for the opportunistic bacillus Stenotrophomonas maltophilia, commonly exposed in the clinical scenario to antibiotics that have no action on this pathogen. Thus, our objective was to investigate whether the prior use of certain non-targeted antimicrobials induces cross-tolerance in S. maltophilia. Materials and methods: Qualitative and quantitative screening tests were performed to evaluate possible tolerance-inducing antimicrobials. In these assays, we exposed the S. maltophilia ATCC® 13637™ to daptomycin (DAP), vancomycin (VAN) and gentamicin (GEN) and evaluated the occurrence of cross-tolerance to the following drugs targeting S. maltophilia: sulfamethoxazole/trimethoprim (SXT), ceftazidime (CAZ) and levofloxacin (LVX). After the screening, strains with tolerance-induced characteristics were tested for genetic similarity by ERIC-PCR. To exclude the contamination hypothesis and confirm the genus and species of the strains, the MALDI-TOF MS was performed. To evaluate tolerance, the following tests were carried out: determination of the Minimum Bactericidal Concentration (MBC) / Minimum Inhibitory Concentration (MIC) ratio for LVX; adapted disk-diffusion test for SXT, CAZ, LVX, ticarcillin-clavulanate (TCC), chloramphenicol (CLO), minocycline (MIN) and polymyxin B (PXB) and time-kill curve for LVX, CAZ and ciprofloxacin (CIP). Results: After induction of the parental strain ATCC® 13637™ to DAP and VAN, three potentially tolerant strains (D25, V15.6 and V3.9) were isolated, whose genetic similarity to parental was confirmed by ERIC-PCR (100% similarity). The induced strains maintained the same MIC value of LVX as the parental strain (0.125 μg/mL), with the MBC/MIC ratio ranging from 2.64 to 3.04 among the induced strains. In the adapted disc-diffusion test, strain D25 showed high tolerance for LVX, CAZ and TCC. In the stationary phase time-kill curves against 50 times the MIC of LVX, strain D25 presented a minimum duration for killing 99,99% of all population (MDK99.99) higher than the parental. The survival rate was also higher for the D25 strain in the presence of CAZ (50xMIC). In general, strain D25 had a higher survival rate in all time-kill curves, except for CIP. Conclusions: Previous exposure of S. maltophilia to daptomycin induces cross-tolerance to levofloxacin, ceftazidime and ticarcillinclavulanate. Thus, erroneous empirical therapy with daptomycin could lead to the failure of these antimicrobials to eradicate S. maltophilia, culminating in chronic infections or death by a subpopulation of tolerant microorganisms.Introdução: A tolerância bacteriana caracteriza-se pela habilidade de uma população microbiana em sobreviver a altas concentrações de uma droga bactericida sem que a Concentração Inibitória Mínima (CIM) seja alterada. Ao término do tratamento antimicrobiano, as cepas tolerantes voltam a se multiplicar, podendo causar infecções recalcitrantes e facilitar a resistência. Muitos estudos relacionam a exposição a antimicrobianos com a indução de tolerância. Embora observada em diversas espécies bacterianas, a tolerância ainda não foi investigada para o bacilo oportunista e emergente Stenotrophomonas maltophilia, comumente exposto a diversos antimicrobianos sem ação sobre este patógeno. Desta forma, nosso objetivo foi investigar se o uso prévio de determinados antimicrobianos não direcionados induz tolerância cruzada em S. maltophilia. Materiais e métodos: Para avaliar possíveis antimicrobianos indutores de tolerância foram feitos ensaios de triagem qualitativa e quantitativa. Nestes ensaios, expomos a cepa S. maltophilia ATCC® 13637 às drogas daptomicina (DAP), vancomicina (VAN) e gentamicina (GEN) e avaliamos a ocorrência de tolerância cruzada frente às drogas específicas contra S. maltophilia: sulfametoxazol/trimetoprim (SXT), ceftazidima (CAZ) e levofloxacina (LVX). Após a triagem, as cepas com características de tolerância induzida foram testadas quanto à similaridade genética por meio de ERIC-PCR. Para excluir a hipótese de contaminação e confirmar o gênero e a espécie das cepas, foi realizado o MALDI-TOF MS. Para avaliação da tolerância foram realizados os seguintes ensaios: determinação da razão Concentração Bactericida Mínima (CBM)/Concentração Inibitória Mínima (CIM) para LVX; teste de disco-difusão adaptado (para SXT, CAZ, LVX, ticarcilina-clavulanato [TCC], cloranfenicol [CLO], minociclina [MIN] e polimixina B [PXB]) e curva de tempo de morte (para LVX, CAZ e ciprofloxacina [CIP]). Resultados: Após indução da cepa parental ATCC® 13637 à DAP e à VAN, três cepas potencialmente tolerantes (D25, V15,6 e V3,9) foram isoladas, cuja similaridade genética com a parental foi comprovada por ERIC-PCR. As cepas induzidas mantiveram o mesmo valor da CIM de LVX que a cepa parental (0,125 μg/mL) com a razão CBM/CIM variando de 2,64 a 3,04 entre as cepas induzidas. No teste de disco-difusão adaptado, a cepa D25 mostrou alta tolerância para LVX, CAZ e TCC. Nas curvas de tempo de morte em fase estacionária frente à 50xCIM de LVX, a cepa D25 apresentou MDK99,99 maior que o da parental. A taxa de sobrevivência também foi maior para a cepa D25 na presença de CAZ (50xCIM). De modo geral, a cepa D25 apresentou maior taxa de sobrevivência em todas as curvas de tempo de morte, exceto para CIP. Conclusões: Encontramos resultados consistentes de que a exposição prévia de S. maltophilia à daptomicina induz tolerância cruzada à levofloxacina, ceftazidima e ticarcilina-clavulanato. Assim, uma terapia empírica equivocada com daptomicina poderia levar à falha destes antimicrobianos em erradicar S. maltophilia, culminando em infecções crônicas por uma subpopulação de microrganismos tolerantes.Texthttp://repositorio.ufes.br/handle/10/10947porUniversidade Federal do Espírito SantoMestrado em Ciências FarmacêuticasPrograma de Pós-Graduação em Ciências FarmacêuticasUFESBRCentro de Ciências da SaúdeTolerancePersistenceDaptomycinStenotrophomonas maltophiliaTolerânciaPersistênciaDaptomicinaTestes de sensibilidade bacterianaBactérias gram-negativasMicrobiologiaAgentes antiinfecciososMedicamentosFarmácia615.1Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophiliainfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da Universidade Federal do Espírito Santo (riUfes)instname:Universidade Federal do Espírito Santo (UFES)instacron:UFESORIGINALtese_11338_Dissertação Final - Mariana Abou Mourad Ferreira.pdftese_11338_Dissertação Final - Mariana Abou Mourad Ferreira.pdfapplication/pdf1825683http://repositorio.ufes.br/bitstreams/e3e1054c-d9c0-4656-a86c-e95f37eed08f/download746b7ec0bf1a927313accf3e33dbd53bMD5210/109472024-06-27 10:59:06.597oai:repositorio.ufes.br:10/10947http://repositorio.ufes.brRepositório InstitucionalPUBhttp://repositorio.ufes.br/oai/requestopendoar:21082024-06-27T10:59:06Repositório Institucional da Universidade Federal do Espírito Santo (riUfes) - Universidade Federal do Espírito Santo (UFES)false |
dc.title.none.fl_str_mv |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
title |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
spellingShingle |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia Ferreira, Mariana Abou Mourad Tolerance Persistence Daptomycin Stenotrophomonas maltophilia Tolerância Persistência Daptomicina Testes de sensibilidade bacteriana Bactérias gram-negativas Farmácia Microbiologia Agentes antiinfecciosos Medicamentos 615.1 |
title_short |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
title_full |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
title_fullStr |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
title_full_unstemmed |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
title_sort |
Tolerância induzida por drogas : efeito da associação in vitro de antimicrobianos em Stenotrophomonas maltophilia |
author |
Ferreira, Mariana Abou Mourad |
author_facet |
Ferreira, Mariana Abou Mourad |
author_role |
author |
dc.contributor.advisor1.fl_str_mv |
Santos, Kênia Valéria dos |
dc.contributor.author.fl_str_mv |
Ferreira, Mariana Abou Mourad |
dc.contributor.referee1.fl_str_mv |
Silva, Rodrigo Cayô da |
dc.contributor.referee2.fl_str_mv |
Tavares, Sarah Goncalves |
contributor_str_mv |
Santos, Kênia Valéria dos Silva, Rodrigo Cayô da Tavares, Sarah Goncalves |
dc.subject.eng.fl_str_mv |
Tolerance Persistence Daptomycin |
topic |
Tolerance Persistence Daptomycin Stenotrophomonas maltophilia Tolerância Persistência Daptomicina Testes de sensibilidade bacteriana Bactérias gram-negativas Farmácia Microbiologia Agentes antiinfecciosos Medicamentos 615.1 |
dc.subject.por.fl_str_mv |
Stenotrophomonas maltophilia Tolerância Persistência Daptomicina Testes de sensibilidade bacteriana Bactérias gram-negativas |
dc.subject.cnpq.fl_str_mv |
Farmácia |
dc.subject.br-rjbn.none.fl_str_mv |
Microbiologia Agentes antiinfecciosos Medicamentos |
dc.subject.udc.none.fl_str_mv |
615.1 |
description |
Introduction: Bacterial tolerance is characterized by the ability of a microbial population to survive the high concentrations of a bactericidal drug without alteration in the Minimum Inhibitory Concentration. At the end of the antimicrobial treatment, the tolerant strains multiply again, which may cause recalcitrant infections and facilitate resistance. Many studies relate antimicrobial exposure to tolerance induction. Although observed in several species, tolerance has not yet been investigated for the opportunistic bacillus Stenotrophomonas maltophilia, commonly exposed in the clinical scenario to antibiotics that have no action on this pathogen. Thus, our objective was to investigate whether the prior use of certain non-targeted antimicrobials induces cross-tolerance in S. maltophilia. Materials and methods: Qualitative and quantitative screening tests were performed to evaluate possible tolerance-inducing antimicrobials. In these assays, we exposed the S. maltophilia ATCC® 13637™ to daptomycin (DAP), vancomycin (VAN) and gentamicin (GEN) and evaluated the occurrence of cross-tolerance to the following drugs targeting S. maltophilia: sulfamethoxazole/trimethoprim (SXT), ceftazidime (CAZ) and levofloxacin (LVX). After the screening, strains with tolerance-induced characteristics were tested for genetic similarity by ERIC-PCR. To exclude the contamination hypothesis and confirm the genus and species of the strains, the MALDI-TOF MS was performed. To evaluate tolerance, the following tests were carried out: determination of the Minimum Bactericidal Concentration (MBC) / Minimum Inhibitory Concentration (MIC) ratio for LVX; adapted disk-diffusion test for SXT, CAZ, LVX, ticarcillin-clavulanate (TCC), chloramphenicol (CLO), minocycline (MIN) and polymyxin B (PXB) and time-kill curve for LVX, CAZ and ciprofloxacin (CIP). Results: After induction of the parental strain ATCC® 13637™ to DAP and VAN, three potentially tolerant strains (D25, V15.6 and V3.9) were isolated, whose genetic similarity to parental was confirmed by ERIC-PCR (100% similarity). The induced strains maintained the same MIC value of LVX as the parental strain (0.125 μg/mL), with the MBC/MIC ratio ranging from 2.64 to 3.04 among the induced strains. In the adapted disc-diffusion test, strain D25 showed high tolerance for LVX, CAZ and TCC. In the stationary phase time-kill curves against 50 times the MIC of LVX, strain D25 presented a minimum duration for killing 99,99% of all population (MDK99.99) higher than the parental. The survival rate was also higher for the D25 strain in the presence of CAZ (50xMIC). In general, strain D25 had a higher survival rate in all time-kill curves, except for CIP. Conclusions: Previous exposure of S. maltophilia to daptomycin induces cross-tolerance to levofloxacin, ceftazidime and ticarcillinclavulanate. Thus, erroneous empirical therapy with daptomycin could lead to the failure of these antimicrobials to eradicate S. maltophilia, culminating in chronic infections or death by a subpopulation of tolerant microorganisms. |
publishDate |
2019 |
dc.date.accessioned.fl_str_mv |
2019-03-15T02:11:22Z |
dc.date.available.fl_str_mv |
2019-03-14 2019-03-15T02:11:22Z |
dc.date.issued.fl_str_mv |
2019-02-22 |
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info:eu-repo/semantics/publishedVersion |
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info:eu-repo/semantics/masterThesis |
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http://repositorio.ufes.br/handle/10/10947 |
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Text |
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Universidade Federal do Espírito Santo Mestrado em Ciências Farmacêuticas |
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Programa de Pós-Graduação em Ciências Farmacêuticas |
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UFES |
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BR |
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Centro de Ciências da Saúde |
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Universidade Federal do Espírito Santo Mestrado em Ciências Farmacêuticas |
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UFES |
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Repositório Institucional da Universidade Federal do Espírito Santo (riUfes) |
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