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dc.contributor.author Lagares, Antonio
dc.contributor.author Agaras, Betina Cecilia
dc.contributor.author Bettiol, Marisa
dc.contributor.author Gatti, Blanca
dc.contributor.author Valverde, Claudio Fabián
dc.date.available 2018-04-16T16:23:24Z
dc.date.issued 2015-05
dc.identifier.citation Lagares, Antonio; Agaras, Betina Cecilia; Bettiol, Marisa; Gatti, Blanca; Valverde, Claudio Fabián; A cultivation-independent PCR-RFLP assay targeting oprF gene for detection and identification of Pseudomonas spp. in samples from fibrocystic paediatric patients; Elsevier Science; Journal of Microbiological Methods; 114; 5-2015; 66-74
dc.identifier.issn 0167-7012
dc.identifier.uri http://hdl.handle.net/11336/42125
dc.description.abstract Species-specific genetic markers are crucial to develop faithful and sensitive molecular methods for the detection and identification of Pseudomonas aeruginosa (Pa). We have previously set up a PCR-RFLP protocol targeting oprF, the gene encoding the genus-specific outer membrane porin F, whose strong conservation and marked sequence diversity allowed detection and differentiation of environmental isolates (Agaras et al., 2012). Here, we evaluated the ability of the PCR-RFLP assay to genotype clinical isolates previously identified as Pa by conventional microbiological methods within a collection of 62 presumptive Pa isolates from different paediatric clinical samples and different sections of the Hospital de Niños ?Sor María Ludovica? from La Plata, Argentina. All isolates, but one, gave an oprF amplicon consistent with that from reference Pa strains. The sequence of the smaller-sized amplicon revealed that the isolate was in fact a P. mendocina strain. The oprF RFLP pattern generated with TaqI or HaeIII nucleases matched those of reference Pa strains for 59 isolates (96%). The other two Pa isolates (4%) revealed a different RFLP pattern based on HaeIII digestion, although oprF sequencing confirmed that Pa identification was correct. We next tested the effectiveness of the PCR-RFLP to detect pseudomonads on clinical samples of paediatric fibrocystic patients directly without sample cultivation. The expected amplicon and its cognate RFLP profile were obtained for all samples in which Pa was previously detected by cultivation-dependent methods. Altogether, these results provide the basis for the application of the oprF PCR-RFLP protocol to directly detect and identify Pa and other non-Pa pseudomonads in fibrocystic clinical samples.
dc.format application/pdf
dc.language.iso eng
dc.publisher Elsevier Science
dc.rights info:eu-repo/semantics/restrictedAccess
dc.rights.uri https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.subject CYSTIC FIBROSIS
dc.subject OPRF
dc.subject PCR-RFLP
dc.subject PSEUDOMONAS AERUGINOSA
dc.subject.classification Otras Ciencias Biológicas
dc.subject.classification Ciencias Biológicas
dc.subject.classification CIENCIAS NATURALES Y EXACTAS
dc.subject.classification Otras Ciencias Biológicas
dc.subject.classification Ciencias Biológicas
dc.subject.classification CIENCIAS NATURALES Y EXACTAS
dc.title A cultivation-independent PCR-RFLP assay targeting oprF gene for detection and identification of Pseudomonas spp. in samples from fibrocystic paediatric patients
dc.type info:eu-repo/semantics/article
dc.type info:ar-repo/semantics/artículo
dc.type info:eu-repo/semantics/publishedVersion
dc.date.updated 2018-04-10T17:52:35Z
dc.journal.volume 114
dc.journal.pagination 66-74
dc.journal.pais Países Bajos
dc.journal.ciudad Amsterdam
dc.description.fil Fil: Lagares, Antonio. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina
dc.description.fil Fil: Agaras, Betina Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina
dc.description.fil Fil: Bettiol, Marisa. Sala de Microbiología HIAEP “Sor María Ludovica”; Argentina
dc.description.fil Fil: Gatti, Blanca. Sala de Microbiología HIAEP “Sor María Ludovica”; Argentina
dc.description.fil Fil: Valverde, Claudio Fabián. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina
dc.journal.title Journal of Microbiological Methods
dc.relation.alternativeid info:eu-repo/semantics/altIdentifier/doi/http://dx.doi.org/10.1016/j.mimet.2015.05.008
dc.relation.alternativeid info:eu-repo/semantics/altIdentifier/url/https://linkinghub.elsevier.com/retrieve/pii/S0167701215001475
dc.conicet.fuente unificacion


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info:eu-repo/semantics/restrictedAccess Excepto donde se diga explícitamente, este item se publica bajo la siguiente descripción: Creative Commons Attribution-NonCommercial-ShareAlike 2.5 Unported (CC BY-NC-SA 2.5)