Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures

ABSTRACT A positive blood culture is a critical result that requires prompt identification of the causative agent. This article describes a simple method to identify microorganisms from positive blood culture broth within the time taken to perform a Gram stain (<20 min). The method is based on in...

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Autores principales:	John D. Walsh, Jay M. Hyman, Larisa Borzhemskaya, Ann Bowen, Caroline McKellar, Michael Ullery, Erin Mathias, Christopher Ronsick, John Link, Mark Wilson, Bradford Clay, Ron Robinson, Thurman Thorpe, Alex van Belkum, W. Michael Dunne
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2013
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/c344f9f7412c48a78262d52fa3cb6e71
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spelling	oai:doaj.org-article:c344f9f7412c48a78262d52fa3cb6e712021-11-15T15:42:31ZRapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures10.1128/mBio.00865-132150-7511https://doaj.org/article/c344f9f7412c48a78262d52fa3cb6e712013-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00865-13https://doaj.org/toc/2150-7511ABSTRACT A positive blood culture is a critical result that requires prompt identification of the causative agent. This article describes a simple method to identify microorganisms from positive blood culture broth within the time taken to perform a Gram stain (<20 min). The method is based on intrinsic fluorescence spectroscopy (IFS) of whole cells and required development of a selective lysis buffer, aqueous density cushion, optical microcentrifuge tube, and reference database. A total of 1,121 monomicrobial-positive broth samples from 751 strains were analyzed to build a database representing 37 of the most commonly encountered species in bloodstream infections or present as contaminants. A multistage algorithm correctly classified 99.6% of unknown samples to the Gram level, 99.3% to the family level, and 96.5% to the species level. There were no incorrect results given at the Gram or family classification levels, while 0.8% of results were discordant at the species level. In 8/9 incorrect species results, the misidentified isolate was assigned to a species of the same genus. This unique combination of selective lysis, density centrifugation, and IFS can rapidly identify the most common microbial species present in positive blood cultures. Faster identification of the etiologic agent may benefit the clinical management of sepsis. Further evaluation is now warranted to determine the performance of the method using clinical blood culture specimens. IMPORTANCE Physicians often require the identity of the infective agent in order to make life-saving adjustments to empirical therapy or to switch to less expensive and/or more targeted antimicrobials. However, standard identification procedures take up to 2 days after a blood culture is signaled positive, and even most rapid molecular techniques take several hours to provide a result. Other techniques are faster (e.g., matrix-assisted laser desorption ionization–time of flight [MALDI-TOF] mass spectrometry) but require time-consuming manual processing steps and expensive equipment. There remains a clear need for a simple, inexpensive method to rapidly identify microorganisms directly from positive blood cultures. The promising new method described in this research article can identify microorganisms in minutes by optical spectroscopy, thus permitting the lab to simultaneously report the presence of a positive blood culture and the organism’s identity.John D. WalshJay M. HymanLarisa BorzhemskayaAnn BowenCaroline McKellarMichael UlleryErin MathiasChristopher RonsickJohn LinkMark WilsonBradford ClayRon RobinsonThurman ThorpeAlex van BelkumW. Michael DunneAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 4, Iss 6 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 John D. Walsh Jay M. Hyman Larisa Borzhemskaya Ann Bowen Caroline McKellar Michael Ullery Erin Mathias Christopher Ronsick John Link Mark Wilson Bradford Clay Ron Robinson Thurman Thorpe Alex van Belkum W. Michael Dunne Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures
description	ABSTRACT A positive blood culture is a critical result that requires prompt identification of the causative agent. This article describes a simple method to identify microorganisms from positive blood culture broth within the time taken to perform a Gram stain (<20 min). The method is based on intrinsic fluorescence spectroscopy (IFS) of whole cells and required development of a selective lysis buffer, aqueous density cushion, optical microcentrifuge tube, and reference database. A total of 1,121 monomicrobial-positive broth samples from 751 strains were analyzed to build a database representing 37 of the most commonly encountered species in bloodstream infections or present as contaminants. A multistage algorithm correctly classified 99.6% of unknown samples to the Gram level, 99.3% to the family level, and 96.5% to the species level. There were no incorrect results given at the Gram or family classification levels, while 0.8% of results were discordant at the species level. In 8/9 incorrect species results, the misidentified isolate was assigned to a species of the same genus. This unique combination of selective lysis, density centrifugation, and IFS can rapidly identify the most common microbial species present in positive blood cultures. Faster identification of the etiologic agent may benefit the clinical management of sepsis. Further evaluation is now warranted to determine the performance of the method using clinical blood culture specimens. IMPORTANCE Physicians often require the identity of the infective agent in order to make life-saving adjustments to empirical therapy or to switch to less expensive and/or more targeted antimicrobials. However, standard identification procedures take up to 2 days after a blood culture is signaled positive, and even most rapid molecular techniques take several hours to provide a result. Other techniques are faster (e.g., matrix-assisted laser desorption ionization–time of flight [MALDI-TOF] mass spectrometry) but require time-consuming manual processing steps and expensive equipment. There remains a clear need for a simple, inexpensive method to rapidly identify microorganisms directly from positive blood cultures. The promising new method described in this research article can identify microorganisms in minutes by optical spectroscopy, thus permitting the lab to simultaneously report the presence of a positive blood culture and the organism’s identity.
format	article
author	John D. Walsh Jay M. Hyman Larisa Borzhemskaya Ann Bowen Caroline McKellar Michael Ullery Erin Mathias Christopher Ronsick John Link Mark Wilson Bradford Clay Ron Robinson Thurman Thorpe Alex van Belkum W. Michael Dunne
author_facet	John D. Walsh Jay M. Hyman Larisa Borzhemskaya Ann Bowen Caroline McKellar Michael Ullery Erin Mathias Christopher Ronsick John Link Mark Wilson Bradford Clay Ron Robinson Thurman Thorpe Alex van Belkum W. Michael Dunne
author_sort	John D. Walsh
title	Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures
title_short	Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures
title_full	Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures
title_fullStr	Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures
title_full_unstemmed	Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures
title_sort	rapid intrinsic fluorescence method for direct identification of pathogens in blood cultures
publisher	American Society for Microbiology
publishDate	2013
url	https://doaj.org/article/c344f9f7412c48a78262d52fa3cb6e71
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Rapid Intrinsic Fluorescence Method for Direct Identification of Pathogens in Blood Cultures

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