An improved in-house lysis-filtration protocol for bacterial identification from positive blood culture bottles with high identification rates by MALDI-TOF MS

Tsuchida, Sachio; Murata, Syota; Miyabe, Akiko; Satoh, Mamoru; Takiwaki, Masaki; Matsushita, Hiroshi; Nomura, Fumio

doi:10.1016/j.mimet.2018.03.014

Cited by 13 publications

(6 citation statements)

References 25 publications

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“…It is worth noting that, compared to existing methods, including similar lysis and centrifugation approach, our protocol shows superiority. The identification success rates of our method are similar to more complicated methods but higher than most of the nonmodified methods (Table S1) (Campigotto et al, 2018;Lin et al, 2018;Pan et al, 2018;Tsuchida et al, 2018;Azrad et al, 2019). We noticed that Simon's research group has used the lysis and centrifugation method for blood culture broth extraction.…”

Section: Discussionmentioning

confidence: 66%

Evaluation of a Rapid and Simplified Protocol for Direct Identification of Microorganisms From Positive Blood Cultures by Using Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS)

Dai

Xue

et al. 2021

Front. Cell. Infect. Microbiol.

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Early and rapid identification of microorganisms is critical for reducing the mortality rate caused by bloodstream infections (BSIs). The accuracy and feasibility of directly identifying pathogens in positive blood cultures by matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been intensely confirmed. In this study, we combined density centrifugation and extra chemical lysis-extraction to develop an optimized method in the blood culture process, which significantly improved the effectiveness of direct identification by MALDI-TOF MS. The accuracy was evaluated by 2,032 positive blood culture samples (115 species of microorganism). The overall MALDI-TOF MS based identification rate with scores ≥ 1.700 was 87.60%. 94.06% of gram-negative bacteria were identified consistently to the genus level, followed by anaerobes (93.33%), gram-positive bacteria (84.46%), and fungi (60.87%). This protocol could obtain results within 10–20 min at a cost of less than $0.1 per sample, which saved up to 24 h in identifying 87.60% of the microorganism from positive blood cultures. This rapid and simplified protocol facilitates the direct identification of microorganism in positive blood cultures, and exhibits the advantages of cost-effective, time-saving, and easy-to-use. It could provide the causative organism of the patient to clinicians in time for targeted treatment and reduce mortality.

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Section: Discussionmentioning

confidence: 66%

Evaluation of a Rapid and Simplified Protocol for Direct Identification of Microorganisms From Positive Blood Cultures by Using Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS)

Dai

Xue

et al. 2021

Front. Cell. Infect. Microbiol.

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“…Pretreatment of samples using MS methods is necessary for direct identification of microorganisms from blood cultures. We previously investigated the functionality of various methods and kits [ 57 , 62 , 63 ]. Among these, the MALDI Sepsityper ® kit (Bruker Daltonics) is extensively used [ 64 , 65 ].…”

Section: Direct Identification From a Positive Blood Culture Bottlmentioning

confidence: 99%

Current Status of Matrix-Assisted Laser Desorption/Ionization–Time-of-Flight Mass Spectrometry (MALDI-TOF MS) in Clinical Diagnostic Microbiology

2020

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Mass spectrometry (MS), a core technology for proteomics and metabolomics, is currently being developed for clinical applications. The identification of microorganisms in clinical samples using matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry (MALDI-TOF MS) is a representative MS-based proteomics application that is relevant to daily clinical practice. This technology has the advantages of convenience, speed, and accuracy when compared with conventional biochemical methods. MALDI-TOF MS can shorten the time used for microbial identification by about 1 day in routine workflows. Sample preparation from microbial colonies has been improved, increasing the accuracy and speed of identification. MALDI-TOF MS is also used for testing blood, cerebrospinal fluid, and urine, because it can directly identify the microorganisms in these liquid samples without prior culture or subculture. Thus, MALDI-TOF MS has the potential to improve patient prognosis and decrease the length of hospitalization and is therefore currently considered an essential tool in clinical microbiology. Furthermore, MALDI-TOF MS is currently being combined with other technologies, such as flow cytometry, to expand the scope of clinical applications.

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“…This approach was originally described for the Vitek MS blood culture kit [37]. Because the Vitek MS blood culture kit is not readily accessible for non-Vitek MS users in Japan, we described an in-house lysis filtration method for use with the MALDI-Biotyper system [52]. Our in-house method is similar to the original lysis filtration method [37], but differs from the previous technique in four ways.…”

Section: Laboratory-developed Tests For Bacterial Collection and Purimentioning

confidence: 99%

Mass spectrometry-based microbiological testing for blood stream infection

et al. 2020

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Background: The most successful application of mass spectrometry (MS) in laboratory medicine is identification (ID) of microorganisms using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in blood stream infection. We describe MALDI-TOF MS-based bacterial ID with particular emphasis on the methods so far developed to directly identify microorganisms from positive blood culture bottles with MALDI-TOF MS including our own protocols. We touch upon the increasing roles of Liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS) as well. Main body: Because blood culture bottles contain a variety of nonbacterial proteins that may interfere with analysis and interpretation, appropriate pretreatments are prerequisites for successful ID. Pretreatments include purification of bacterial pellets and short-term subcultures to form microcolonies prior to MALDI-TOF MS analysis. Three commercial protocols are currently available: the Sepsityper ® kit (Bruker Daltonics), the Vitek MS blood culture kit (bioMerieux, Inc.), and the rapid BACpro ® II kit (Nittobo Medical Co., Tokyo). Because these commercially available kits are costly and bacterial ID rates using these kits are not satisfactory, particularly for Gram-positive bacteria, various home-brew protocols have been developed: 1. Stepwise differential sedimentation of blood cells and microorganisms, 2. Combination of centrifugation and lysis procedures, 3. Lysis-vacuum filtration, and 4. Centrifugation and membrane filtration technique (CMFT). We prospectively evaluated the performance of this CMFT protocol compared with that of Sepsityper ® using 170 monomicrobial positive blood cultures. Although preliminary, the performance of the CMFT was significantly better than that of Sepsityper ® , particularly for Gram-positive isolates. MALDI-TOF MS-based testing of polymicrobial blood specimens, however, is still challenging. Also, its contribution to assessment of susceptibility and resistance to antibiotics is still limited. For this purpose, liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS) should be more useful because this approach can identify as many as several thousand peptide sequences. Conclusion: MALDI-TOF MS is now an essential tool for rapid bacterial ID of pathogens that cause blood stream infection. For the purpose of assessment of susceptibility and resistance to antibiotics of the pathogens, the roles of liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS) will increase in the future.

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An improved in-house lysis-filtration protocol for bacterial identification from positive blood culture bottles with high identification rates by MALDI-TOF MS

Cited by 13 publications

References 25 publications

Evaluation of a Rapid and Simplified Protocol for Direct Identification of Microorganisms From Positive Blood Cultures by Using Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS)

Evaluation of a Rapid and Simplified Protocol for Direct Identification of Microorganisms From Positive Blood Cultures by Using Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS)

Current Status of Matrix-Assisted Laser Desorption/Ionization–Time-of-Flight Mass Spectrometry (MALDI-TOF MS) in Clinical Diagnostic Microbiology

Mass spectrometry-based microbiological testing for blood stream infection

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