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Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing

For mass spectrometry-based diagnostics of microorganisms, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is currently routinely used to identify urinary tract pathogens. However, it requires a lengthy culture step for accurate pathogen identification, an...

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Autores principales: Svetličić, Ema, Dončević, Lucija, Ozdanovac, Luka, Janeš, Andrea, Tustonić, Tomislav, Štajduhar, Andrija, Brkić, Antun Lovro, Čeprnja, Marina, Cindrić, Mario
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9457756/
https://www.ncbi.nlm.nih.gov/pubmed/36080229
http://dx.doi.org/10.3390/molecules27175461
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author Svetličić, Ema
Dončević, Lucija
Ozdanovac, Luka
Janeš, Andrea
Tustonić, Tomislav
Štajduhar, Andrija
Brkić, Antun Lovro
Čeprnja, Marina
Cindrić, Mario
author_facet Svetličić, Ema
Dončević, Lucija
Ozdanovac, Luka
Janeš, Andrea
Tustonić, Tomislav
Štajduhar, Andrija
Brkić, Antun Lovro
Čeprnja, Marina
Cindrić, Mario
author_sort Svetličić, Ema
collection PubMed
description For mass spectrometry-based diagnostics of microorganisms, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is currently routinely used to identify urinary tract pathogens. However, it requires a lengthy culture step for accurate pathogen identification, and is limited by a relatively small number of available species in peptide spectral libraries (≤3329). Here, we propose a method for pathogen identification that overcomes the above limitations, and utilizes the MALDI-TOF/TOF MS instrument. Tandem mass spectra of the analyzed peptides were obtained by chemically activated fragmentation, which allowed mass spectrometry analysis in negative and positive ion modes. Peptide sequences were elucidated de novo, and aligned with the non-redundant National Center for Biotechnology Information Reference Sequence Database (NCBInr). For data analysis, we developed a custom program package that predicted peptide sequences from the negative and positive MS/MS spectra. The main advantage of this method over a conventional MALDI-TOF MS peptide analysis is identification in less than 24 h without a cultivation step. Compared to the limited identification with peptide spectra libraries, the NCBI database derived from genome sequencing currently contains 20,917 bacterial species, and is constantly expanding. This paper presents an accurate method that is used to identify pathogens grown on agar plates, and those isolated directly from urine samples, with high accuracy.
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spelling pubmed-94577562022-09-09 Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing Svetličić, Ema Dončević, Lucija Ozdanovac, Luka Janeš, Andrea Tustonić, Tomislav Štajduhar, Andrija Brkić, Antun Lovro Čeprnja, Marina Cindrić, Mario Molecules Article For mass spectrometry-based diagnostics of microorganisms, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is currently routinely used to identify urinary tract pathogens. However, it requires a lengthy culture step for accurate pathogen identification, and is limited by a relatively small number of available species in peptide spectral libraries (≤3329). Here, we propose a method for pathogen identification that overcomes the above limitations, and utilizes the MALDI-TOF/TOF MS instrument. Tandem mass spectra of the analyzed peptides were obtained by chemically activated fragmentation, which allowed mass spectrometry analysis in negative and positive ion modes. Peptide sequences were elucidated de novo, and aligned with the non-redundant National Center for Biotechnology Information Reference Sequence Database (NCBInr). For data analysis, we developed a custom program package that predicted peptide sequences from the negative and positive MS/MS spectra. The main advantage of this method over a conventional MALDI-TOF MS peptide analysis is identification in less than 24 h without a cultivation step. Compared to the limited identification with peptide spectra libraries, the NCBI database derived from genome sequencing currently contains 20,917 bacterial species, and is constantly expanding. This paper presents an accurate method that is used to identify pathogens grown on agar plates, and those isolated directly from urine samples, with high accuracy. MDPI 2022-08-25 /pmc/articles/PMC9457756/ /pubmed/36080229 http://dx.doi.org/10.3390/molecules27175461 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Svetličić, Ema
Dončević, Lucija
Ozdanovac, Luka
Janeš, Andrea
Tustonić, Tomislav
Štajduhar, Andrija
Brkić, Antun Lovro
Čeprnja, Marina
Cindrić, Mario
Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing
title Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing
title_full Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing
title_fullStr Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing
title_full_unstemmed Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing
title_short Direct Identification of Urinary Tract Pathogens by MALDI-TOF/TOF Analysis and De Novo Peptide Sequencing
title_sort direct identification of urinary tract pathogens by maldi-tof/tof analysis and de novo peptide sequencing
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9457756/
https://www.ncbi.nlm.nih.gov/pubmed/36080229
http://dx.doi.org/10.3390/molecules27175461
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