Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples
Background: Pseudomonas aeruginosa is a major cause of hospital-acquired and chronic infections, characterised by an extraordinary capacity to develop antimicrobial resistance through the selection of chromosomal mutations, leading to treatment failure. Here, we designed and tested a hybridisation-b...
| Autores: | , , , , , , , , , , , , , , |
|---|---|
| Tipo de recurso: | artículo |
| Estado: | Versión publicada |
| Fecha de publicación: | 2024 |
| País: | España |
| Institución: | Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya) |
| Repositorio: | Recercat. Dipósit de la Recerca de Catalunya |
| OAI Identifier: | oai:recercat.cat:10230/68493 |
| Acceso en línea: | http://hdl.handle.net/10230/68493 http://dx.doi.org/10.1016/j.ebiom.2024.105367 |
| Access Level: | acceso abierto |
| Palabra clave: | Antimicrobial resistance development Chronic infections Cystic fibrosis Nosocomial infections Pseudomonas aeruginosa Resistome |
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Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samplesCortes-Lara, SaraMedina Reatiga, PaolaBarrio Tofiño, Ester delGomis Font, María A.Cabot, GabrielGómez Romano, FernandoAyestarán, Nicolás EduardoColomar, AsunciónPalou Rotger, AlexandreOteo Iglesias, JesúsCampo, Rosa delCantón, RafaelHorcajada Gallego, Juan PabloLópez-Causapé, CarlaOliver, AntonioAntimicrobial resistance developmentChronic infectionsCystic fibrosisNosocomial infectionsPseudomonas aeruginosaResistomeBackground: Pseudomonas aeruginosa is a major cause of hospital-acquired and chronic infections, characterised by an extraordinary capacity to develop antimicrobial resistance through the selection of chromosomal mutations, leading to treatment failure. Here, we designed and tested a hybridisation-based capture system for the enrichment of genes of interest before sequencing to monitor resistant populations genomics directly from clinical samples. Methods: A panel for enrichment before sequencing of close to 200 genes related to P. aeruginosa antimicrobial resistance, multilocus sequence typing, mutability or virulence was designed, synthesised (KAPA HyperCap, Roche) and initially validated in vitro using a multidrug-resistant ST175 isolate and representative isolates from major P. aeruginosa clades. In vivo testing included ventilator associated pneumonia by MDR P. aeruginosa in ICU (3-10 sequential samples from 3 patients) and chronic respiratory infection by hypermutable P. aeruginosa in cystic fibrosis (8 sequential samples from a single patient covering a 4-year period). Results from direct sequencing with the enrichment panel were compared with those of whole genome sequencing (WGS) and phenotypic profiling of 10 isolated colonies per sample. Findings: In vitro assays confirmed the selectivity of the enrichment panel and the correct identification of the vast mutational resistome of ST175, including specific mutations even when introduced in a 1:100 proportion. In vivo performance was at least equivalent to sequencing 10 colonies per sample, including the accurate identification of the sequence types and the basal and acquired mutational resistome. To note, specific resistance mutations, such as those in ampC leading to resistance to novel β-lactams, could be traced even at frequencies of 1%. Moreover, the coselection of mutator populations and antibiotic resistance mutations, predicted in theoretical and in vitro studies, was evidenced in vivo. Interpretation: This proof-of-concept study demonstrates that resistance genomics of P. aeruginosa can be analysed directly from clinical samples, determining not only a considerable reduction in turnaround time and cost from a diagnostics perspective, but also an unprecedented potency for accurate monitoring of in vivo population dynamics in bacterial infections. Funding: Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación and Unión Europea-NextGenerationEU.Elsevier202420242024info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionapplication/pdfapplication/pdfhttp://hdl.handle.net/10230/68493http://dx.doi.org/10.1016/j.ebiom.2024.105367reponame:Recercat. Dipósit de la Recerca de Catalunyainstname:Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya)InglésEBioMedicine. 2024 Oct;108:105367© 2024 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).http://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/openAccessoai:recercat.cat:10230/684932026-05-29T05:05:01Z |
| dc.title.none.fl_str_mv |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| title |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| spellingShingle |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples Cortes-Lara, Sara Antimicrobial resistance development Chronic infections Cystic fibrosis Nosocomial infections Pseudomonas aeruginosa Resistome |
| title_short |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| title_full |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| title_fullStr |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| title_full_unstemmed |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| title_sort |
Monitoring of Pseudomonas aeruginosa mutational resistome dynamics using an enrichment panel for direct sequencing of clinical samples |
| dc.creator.none.fl_str_mv |
Cortes-Lara, Sara Medina Reatiga, Paola Barrio Tofiño, Ester del Gomis Font, María A. Cabot, Gabriel Gómez Romano, Fernando Ayestarán, Nicolás Eduardo Colomar, Asunción Palou Rotger, Alexandre Oteo Iglesias, Jesús Campo, Rosa del Cantón, Rafael Horcajada Gallego, Juan Pablo López-Causapé, Carla Oliver, Antonio |
| author |
Cortes-Lara, Sara |
| author_facet |
Cortes-Lara, Sara Medina Reatiga, Paola Barrio Tofiño, Ester del Gomis Font, María A. Cabot, Gabriel Gómez Romano, Fernando Ayestarán, Nicolás Eduardo Colomar, Asunción Palou Rotger, Alexandre Oteo Iglesias, Jesús Campo, Rosa del Cantón, Rafael Horcajada Gallego, Juan Pablo López-Causapé, Carla Oliver, Antonio |
| author_role |
author |
| author2 |
Medina Reatiga, Paola Barrio Tofiño, Ester del Gomis Font, María A. Cabot, Gabriel Gómez Romano, Fernando Ayestarán, Nicolás Eduardo Colomar, Asunción Palou Rotger, Alexandre Oteo Iglesias, Jesús Campo, Rosa del Cantón, Rafael Horcajada Gallego, Juan Pablo López-Causapé, Carla Oliver, Antonio |
| author2_role |
author author author author author author author author author author author author author author |
| dc.subject.none.fl_str_mv |
Antimicrobial resistance development Chronic infections Cystic fibrosis Nosocomial infections Pseudomonas aeruginosa Resistome |
| topic |
Antimicrobial resistance development Chronic infections Cystic fibrosis Nosocomial infections Pseudomonas aeruginosa Resistome |
| description |
Background: Pseudomonas aeruginosa is a major cause of hospital-acquired and chronic infections, characterised by an extraordinary capacity to develop antimicrobial resistance through the selection of chromosomal mutations, leading to treatment failure. Here, we designed and tested a hybridisation-based capture system for the enrichment of genes of interest before sequencing to monitor resistant populations genomics directly from clinical samples. Methods: A panel for enrichment before sequencing of close to 200 genes related to P. aeruginosa antimicrobial resistance, multilocus sequence typing, mutability or virulence was designed, synthesised (KAPA HyperCap, Roche) and initially validated in vitro using a multidrug-resistant ST175 isolate and representative isolates from major P. aeruginosa clades. In vivo testing included ventilator associated pneumonia by MDR P. aeruginosa in ICU (3-10 sequential samples from 3 patients) and chronic respiratory infection by hypermutable P. aeruginosa in cystic fibrosis (8 sequential samples from a single patient covering a 4-year period). Results from direct sequencing with the enrichment panel were compared with those of whole genome sequencing (WGS) and phenotypic profiling of 10 isolated colonies per sample. Findings: In vitro assays confirmed the selectivity of the enrichment panel and the correct identification of the vast mutational resistome of ST175, including specific mutations even when introduced in a 1:100 proportion. In vivo performance was at least equivalent to sequencing 10 colonies per sample, including the accurate identification of the sequence types and the basal and acquired mutational resistome. To note, specific resistance mutations, such as those in ampC leading to resistance to novel β-lactams, could be traced even at frequencies of 1%. Moreover, the coselection of mutator populations and antibiotic resistance mutations, predicted in theoretical and in vitro studies, was evidenced in vivo. Interpretation: This proof-of-concept study demonstrates that resistance genomics of P. aeruginosa can be analysed directly from clinical samples, determining not only a considerable reduction in turnaround time and cost from a diagnostics perspective, but also an unprecedented potency for accurate monitoring of in vivo population dynamics in bacterial infections. Funding: Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación and Unión Europea-NextGenerationEU. |
| publishDate |
2024 |
| dc.date.none.fl_str_mv |
2024 2024 2024 |
| dc.type.none.fl_str_mv |
info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion |
| format |
article |
| status_str |
publishedVersion |
| dc.identifier.none.fl_str_mv |
http://hdl.handle.net/10230/68493 http://dx.doi.org/10.1016/j.ebiom.2024.105367 |
| url |
http://hdl.handle.net/10230/68493 http://dx.doi.org/10.1016/j.ebiom.2024.105367 |
| dc.language.none.fl_str_mv |
Inglés |
| language_invalid_str_mv |
Inglés |
| dc.relation.none.fl_str_mv |
EBioMedicine. 2024 Oct;108:105367 |
| dc.rights.none.fl_str_mv |
http://creativecommons.org/licenses/by-nc-nd/4.0/ info:eu-repo/semantics/openAccess |
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http://creativecommons.org/licenses/by-nc-nd/4.0/ |
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openAccess |
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application/pdf application/pdf |
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Elsevier |
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Elsevier |
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Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya) |
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Recercat. Dipósit de la Recerca de Catalunya |
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