VITEK® MSSelection of Publications

2017 EDITION

The VITEK® MS system is a MALDI-TOF (Matrix Assisted Laser Desorption Ionization - Time of Flight) Mass Spectrometry system that has been designed to provide a rapid, accurate and reliable identification result to the microbiology laboratory. Moreover, the VITEK® MS system aids the physician in diagnosing and confirming microbial infections quickly which ultimately helps reduce the time for effective treatment and management of patients with infectious diseases.

MALDI-TOF MS allows for the detection of high-abundance soluble proteins, including ribosomal and other structural proteins, directly from intact microbial cells resulting in spectra that are analyzed with the VITEK® MS system. The VITEK® MS database has been created by collecting mass spectra from multiple isolates per species, geographically diverse isolates, different sample origins, all grown on different media types with different incubation times.The VITEK® MS system has a comprehensive database of clinically relevant species including mycobacteria, nocardia and molds (version 3.0), that allows the identification of organisms in a matter of minutes. Compared with conventional phenotype or PCR-based identification, MALDI-TOF MS provides rapid turnaround time (1-2 minutes per sample), low sample volume requirements, and low reagent costs.

VITEK® MS system

The VITEK® MS system includes the VITEK® MS Prep Station, VITEK® MS Acquisition Station and MYLA® software:• The VITEK® MS Prep Station securely links specimen information with each spot on the

target slide and to the VITEK® 2 Cassette containing the Susceptibility card. • The VITEK® MS Acquisition station receives the spectra data from the instrument which

are then sent to the MYLA® software. • MYLA® is connected to both the VITEK® MS Prep Station and Acquisition Station allowing

for complete traceability, and holds the compute engine software for identification.

VITEK® MS Plus system

The VITEK® MS Plus system combines both the robustness and accuracy of the VITEK® MS for routine patient (IVD) testing, but also allows users to expand their research capabilities using a broad and open research database in which they can easily record and save spectra to build their own personalized database. Users can perform research protocols such as strain typing, detection of bacterial toxins and resistance. With only one sample preparation, users can simply switch between the VITEK® MS IVD and VITEK® MS RUO database for maximum data generation from each sample while still meeting all compliance and IVD regulations.

MASS SPECTROMETRY POWERED BY MICROBIOLOGY

*ID/AST: Identification/Antimicrobial Susceptibility Testing

1. Barenfanger J., et. al. J Clin Microbiol, 1999; 37(5): 1415.2. Galar A., et. al. J Infect, 2012; 65(4): 302.3. Galar A., et. al. Eur J Clin Microbiol Infect Dis, 2012; 31 (9): 2445.4. LaBombardi V.J. bioMérieux White Paper, 2011.

MOVING MICROBIOLOGY FORWARD

VITEK® Solutions combines the optimized ID/AST* workflow and operational efficiency of VITEK® MS and VITEK® 2 to provide real time reporting of results

that support quicker treatment decisions.1,2,3,4

Same-day accurate results with:

• VITEK® MS IDs• VITEK® 2 ASTs

Seamless integration of ID/AST results from VITEK® MS and VITEK® 2 with MYLA®

Helps clinicians:

• quickly optimize patient therapy• reduce hospital length of stay• improve antibiotic stewardship• reduce healthcare costs

VITEK® SOLUTIONS

VITEK® Solutions is the only complete identification andantimicrobial susceptibility testing (ID/AST) platform

designed by microbiologists for microbiologists.

VITEK® MS VITEK® 2

MYLA®

CONTENTS➔ ARTICLES VITEK® MS IVD V3.0*

Routine identification of nocardia species by MALDI-TOF mass spectrometry.Girard V, Mailler S, Polsinelli S, Jacob D, Saccomani MC, Celliere B, Monnin V, van Belkum A, Hagenn F, Meis JF, Durand G.DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2017;87(1):7-10

Identification of mycobacterium spp. and nocardia spp. from solid and liquid cultures by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS).Girard V, Mailler S, Welker M, Arsac M, Cellière B, Cotte-Pattat PJ, Chatellier S, Durand G, Béni AM, Schrenzel J, Miller E, Dussoulier R, Dunne WM Jr, Butler-Wu S, Saubolle MA, Sussland D, Bell M, van Belkum A, Deol P.DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2016;86(3):277-283

Comparison of Sample Preparation Methods, Instrumentation Platforms, and Contemporary Commercial Databases for Identification of Clinically Relevant Mycobacteria by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry.Wilen CB, McMullen AR, Burnham C-AD.JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(7):2308-15

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry using the VITEK® MS system for rapid and accurate identification of dermatophytes on solid cultures.De Respinis S, Monnin V, Girard V, Welker M, Arsac M, Cellière B, Durand G, Bosshard PP, Farina C, Passera M, Van Belkum A, Petrini O, Tonolla M.JOURNAL OF CLINICAL MICROBIOLOGY 2014;52(12):4286-92

Rapid Inactivation of mycobacterium and nocardia species before identification using MALDI-TOF mass spectrometry.Wm. Michael Dunne Jr., Kirk Doing, Elizabeth Miller, Eric Miller, Erik Moreno, Mehdi Baghli, Sandrine Mailler, Victoria Girard, Alex van Belkum, and Parampal DeolJOURNAL OF CLINICAL MICROBIOLOGY 2014;52(10):3654-3659

*CE-Marked and FDA cleared. Check with your local representative for availability.

VITEK® MS IVD V2.0 (In Vitro Diagnostic Applications)

Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) for rapid identification of micro-organisms in the routine clinical microbiology laboratory. Wattal C, Oberoi JK, Goel N, Raveendran R, Khanna S.EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2016; doi:10.1007/s10096-016-2864-9.

Comparison of VITEK® MS and MALDI Biotyper for identification of Actinomycetaceae of clinical importance.Ferrand J, Hochard H, Girard V, Aissa N, Bogard B, Alauzet C, Lozniewski A.JOURNAL OF CLINICAL MICROBIOLOGY 2016;54(3):782-4.

MALDI-TOF mass spectrometry for differentiation between Streptococcus pneumoniae and Streptococcus pseudopneumoniae.van Prehn J, van Veen SQ, Schelfaut JJ, Wessels E.DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2016;85(1):9-11

Using Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) Complemented with Selected 16S rRNA and gyrB Genes Sequencing to Practically Identify Clinical Important Viridans Group Streptococci (VGS).Zhou M, Yang Q, Kudinha T, Zhang L, Xiao M, Kong F, Zhao Y, Xu YC.FRONTIERS IN MICROBIOLOGY. 2016 ;7:1328

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Assessment of Reproducibility of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry for Bacterial and Yeast Identification.Westblade LF, Garner OB, MacDonald K, Bradford C, Pincus DH, Mochon AB, Jennemann R, Manji R, Bythrow M, Lewinski MA, Burnham CD and Ginocchio CC.JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(7):2349-52

Comparative evaluation of two matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) systems, VITEK® MS and Microflex LT, for the identification of Gram-positive cocci routinely isolated in clinical microbiology laboratories. Lee M, Chung HS, Moon HW, Lee SH, Lee K. JOURNAL OF MICROBIOLOGICAL METHODS 2015;113:13–15

Performances and Reliability of Bruker Microflex LT and VITEK® MS MALDI-TOF Mass Spectrometry Systems for the Identification of Clinical Microorganisms. Bilecen K, Yaman G, Ciftci U, Laleli YR.BIOMEDICAL RESEARCH INTERNATIONAL 2015 Dec 17. doi:10.1155/2015/516410

Comparison and optimization of two MALDI-TOF MS platforms for the identification of medically relevant yeast species. Pence MA, McElvania TeKippe E, Wallace MA, Burnham CA.EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2014;33(10):1703-12

MULTICENTER EVALUATIONS OF THE VITEK® MS IVD V2.0 SYSTEM

Multicenter validation of the VITEK® MS v2.0 MALDI-TOF mass spectrometry system for the identification of fastidious gram-negative bacteria. Branda JA, Rychert J, Burnham CA, Bythrow M, Garner OB, Ginocchio CC, Jennemann R, Lewinski MA, Manji R, Mochon AB, Procop GW, Richter SS, Sercia LF, Westblade LF, Ferraro MJ.DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2014;78(2):129-31

Multicenter evaluation of the VITEK® MS system for mass spectrometric identification of non-Enterobacteriaceae Gram-negative bacilli. Manji R, Bythrow M, Branda JA, Burnham CA, Ferraro MJ, Garner OB, Jennemann R, Lewinski MA, Mochon AB, Procop GW, Richter SS, Rychert JA, Sercia L, Westblade LF, Ginocchio CC.EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2014; 33(3):337-46

Multicenter Evaluation of Mass Spectrometric Identification of Anaerobic Bacteria Using the VITEK® MS System. Garner OB, Mochon AB, Branda JA, Burnham CD, Bythrom M, Ferraro MJ, Ginocchio CC, Jennemann R, Manji R, Procop GW, Richter SS, Rychert JA, Sercia L, Westblade LF, Lewinski MA.CLINICAL MICROBIOLOGY AND INFECTION 2014;20(4):335-9

Multicenter Evaluation of the VITEK® MS Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry System for Identification of Gram-Positive Aerobic Bacteria. Rychert J, Burnham CD, Bythrow M, Garner OB, Ginocchio CC, Jennemann R, Lewinski MA, Manji R, Mochon A, Procop GW, Richter SS, Sercia L, Westblade LF, Ferraro MJ, Branda JA.JOURNAL OF CLINICAL MICROBIOLOGY 2013; 51(7):2225-31

Multicenter Study Evaluating the VITEK® MS System for Identification of Medically Important Yeasts.Westblade LF, Jennemann R, Branda JA, Bythrow M, Ferraro MJ, Gerner OB, Ginocchio CC, Lewinski MA, Manji R, Mochon AB, Procop GW, Richter SS, Rychert JA, Sercia L, Burnham CD.JOURNAL OF CLINICAL MICROBIOLOGY 2013; 51(7):2267-72

Identification of Enterobacteriaceae by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using the VITEK® MS system. Richter SS, Sercia L, Branda JÁ, Burnham CD, Bythrow M., Ferraro MJ, Garner OB, Ginocchio CC, Jennemann R, Lewinski MA, Manji R, Mochon AB, Rychert JA, Westblade LF, Procop GW.EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2013;32(12):1571-8

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VITEK® MS RUO (Research Use Only)

Comparison among four proposed direct blood culture microbial identification methods using MALDI-TOF MS. Bazzi AM, Rabaan AA, El Edaily Z, John S, Fawarah MM, Al-Tawfiq JA.JOURNAL OF INFECTION AND PUBLIC HEALTH 2016 Jun 13. pii: S1876-0341(16)30068-5

Rapid urine preparation prior to identification of uropathogens by MALDI-TOF MS. Veron L., Mailler S., Girard V., Muller B., LHostis G., Ducruix C., Lesenne A., Richez A., Rostaing H., Lanet V., Ghirardi S., Van Belkum A., Mallard F.EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY & INFECTIOUS DISEASES 2015;34(9):1787-95

Same Day Identification and Full Panel Antimicrobial Susceptibility Testing of Bacteria from Positive Blood Culture Bottles Made Possible by a Combined Lysis-Filtration Method with MALDI-TOF VITEK® Mass Spectrometry and the VITEK® 2 System. Machen A, Drake T, Wang YF. PLOS ONE 2014;9(2):E87870129-31

Identification and typing of the emerging pathogen Candida auris by matrix-assisted laser desorption ionisation time of flight mass spectrometry.Girard V, Mailler S, Chetry M, Vidal C, Durand G, van Belkum A, Colombo AL, Hagen F, Meis JF, Chowdhary A.MYCOSES 2016;59(8):535-8

Comparison of MALDI-TOF MS and AFLP for strain typing of ESBL-producing Escherichia coli. Veenemans J, Welker M, van Belkum A, Saccomani M.C., Girard V, Pettersson A, Verhulst C, Kluytmans-Vandenbergh M, Kluytmans J.EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2016; 35:829–838

Influence of Culture Media on Detection of Carbapenem Hydrolysis by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.Ramos AC, Carvalhaes CG, Cordeiro-Moura JR, Rockstroh AC, Machado AM, Gales AC.JOURNAL OF CLINICAL MICROBIOLOGY 2016;54(7):1896-8

Rapid and accurate identification of Streptococcus equi subspecies by MALDI-TOF MS.Kudirkiene E, Welker M, Knudsen NR, Bojesena AM.SYSTEMATIC AND APPLIED MICROBIOLOGY 2015;38(5):315-22

Fusobacterium nucleatum subspecies identification by matrix-assisted Laser desorption ionization-time of flight mass spectrometry.Nie S, Tian B, Wang X, Pincus DH, Welker M, Gilhuley K, Lu X, Han YW, Tang YW.JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(4):1399-402.

Mycobacterium abscessus Complex Identification with Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.Panagea T, Pincus DH, Grogono D, Jones M, Bryant J, Parkhill J, Floto RH, Gilligan P.JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(7):2355-8.

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VITEK® MS IVD V3.0

VITEK® MS IVD V3.0

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KEY POINTS

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VITEK® MS IVD V3.0

DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2016;86(3):277-283

Identification of mycobacterium spp. and nocardia spp. from solid and liquid cultures by matrix-assisted laser desorption ionization–time of flight mass spectrometry

(MALDI-TOF MS).Girard V1, Mailler S1, Welker M1, Arsac M1, Cellière B1, Cotte-Pattat PJ1, Chatellier S1, Durand G1, Béni AM2, Schrenzel J2,

Miller E3, Dussoulier R3, Dunne WM Jr3, Butler-Wu S4, Saubolle MA5, Sussland D5, Bell M6, van Belkum A7, Deol P3

(1)bioMérieux, Microbiology R&D, Route de Port Michaud, 38390, La Balme, Les Grottes, France.(2)Hôpitaux Universitaires de Genève, Laboratoire de Bactériologie, Rue Gabrielle Perret Gentil 4, 1211, Geneva 14, Switzerland.

(3)bioMérieux Inc., Microbiology R&D, 100 Rodolphe St, Durham, NC, 27712, USA.(4)Clinical Microbiology Laboratory, University of Washington, 1959 NE Pacific Street, Seattle, WA 98195, USA; Keck School of Medicine,

University of Southern California, 1975 Zonal Avenue, Los Angeles, CA, 90033, USA.(5)Banner University Medical Center, 1111 E. McDowell Rd., Phoenix, AZ, 85006, USA.

(6)Centers for Disease Control and Prevention, 1600 Clifton Road, Atlanta, GA, 30329-4027, USA.(7)bioMérieux, Microbiology R&D, Route de Port Michaud, 38390, La Balme, Les Grottes, France

The genus mycobacterium comprises a large number of species with a wide spectrum of pathogenicity. Mycobacterium tuberculosis is among the greatest public health threats in low-income countries. Nontuberculous mycobacteria are environmental organisms which occasionally act as opportunistic pathogens, leading to a wide array of clinical syndromes. Nocardia species are also involved in many different infections. Traditional identification of mycobacteria and nocardia species has mainly relied on phenotypic and biochemical traits and remains cumbersome. More recent methods involve molecular biology but are restricted to a limited number of species and could be relatively costly. The application of MALDI-TOF to the identification of mycobacteria and nocardia could provide a rapid and accurate way to identify these important species.

In this study, the VITEK® MS database was generated using 84 strains of nocardia and 410 strains of mycobacteria from clinically relevant species that were treated with a specific sample preparation protocol. A total of 1520 spectra were acquired for mycobacteria on 6 different media (liquid and solid). For nocardia, 875 spectra were acquired on 5 different solid media. For a number of organisms, species complexes or groups had to be defined comprising multiple species such as Mycobacterium tuberculosis complex or Mycobacterium fortuitum group.

A total of 183 mycobacterium and 213 nocardia unique isolates were used to challenge the database for validation purposes using the extraction protocol. Results were compared to nucleic acid sequence-based identifications. For mycobacteria, the VITEK® MS gave an accurate identification to the species or complex level for 164 of 183 isolates (90%). For nocardia, 187 of the 213 isolates (88%) were correctly identified by VITEK® MS.

In conclusion, the optimized VITEK® MS IVD database (version 3), when built with spectra acquired in different conditions and associated with a suitable sample preparation protocol enables rapid, reliable and robust identification of the main species of nocardia and mycobacteria of clinical interest.

“We here show that species belonging to the genera mycobacterium and nocardia can be adequately identified by

VITEK® MS when the recommendations for incubation and sample preparation are followed”

DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2017;87(1):7-10

Routine identification of nocardia species by MALDI-TOF mass spectrometry.

Girard Va, Mailler Sa, Polsinelli Sa, Jacob Da, Saccomani M.C.a, Celliere Ba, Monnin Va, van Belkum Aa, Hagen Fb, Meis J.F.b, c, Durand Ga

(a) R&D Microbiology, bioMérieux, La Balme Les Grottes, France(b) Canisius–Wilhelmina Hospital, Department of Medical Microbiology and Infectious Diseases, Nijmegen, The Netherlands

(c) Department of Medical Microbiology, Radboud University Medical Center, Nijmegen, The Netherlands

Nocardia have been involved in many different infections and their identification by phenotypic methods is cumbersome and time-consuming. Molecular identification is possible but not always available in routine practice in clinical labs. It appears important to develop and validate the use of a MALDI-TOF system such as VITEK® MS to identify those organisms.

In this study, the VITEK® MS database was generated using 84 strains from clinically relevant species using a specific sample preparation protocol. A total of 171 unique isolates (17 nocardia species) were analyzed for validation purpose using the extraction protocol and results were compared to nucleic acid sequence-based identifications. For 155 of 171 isolates (91%), the VITEK® MS gave an accurate identification to the species level. If only claimed species are considered, 155 of 164 isolates (95%) were correctly identified. No identification was obtained for 14 isolates (6%), but 7 isolates were unclaimed.

In conclusion, the optimized VITEK® MS IVD database (version 3), associated with a suitable sample preparation protocol, enables rapid, reliable and robust identification of the main nocardia spp. of clinical interest. This study also demonstrates the value of integrating MALDI-TOF MS technology into routine laboratory use for identification of nocardia spp.

“...Our data show that a reliable identification can be obtained using a large collection of nocardia spp. with the next version of the VITEK® MS IVD

database (version 3) without the need of a custom database”

KEY POINTS

➔ The VITEK® MS system is highly accurate for the identification of nocardia in the clinical laboratory setting when used with a suitable sample preparation protocol.

➔ Most nocardia species can be clearly distinguished.

➔ Shortening the time for nocardia identification will allow better patient care.

➔ Mycobacteria from solid and liquid cultures and nocardia from solid cultures can be accurately identified using VITEK® MS and a specific sample preparation protocol.

➔ Mycobacteria and nocardia can be identified to the species level in most cases.

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VITEK® MS IVD V3.0

JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(7):2308-15

Comparison of Sample Preparation Methods, Instrumentation Platforms, and Contemporary Commercial

Databases for Identification of Clinically Relevant Mycobacteria by Matrix-Assisted Laser Desorption

Ionization–Time of Flight Mass Spectrometry.Wilen CB1, McMullen AR1, Burnham C-AD1,2

(1) Department of Pathology & Immunology, Washington University School of Medicine in St. Louis, St. Louis, Missouri, USA(1) Department of Pediatrics, Washington University School of Medicine in St. Louis, St. Louis, Missouri, USA

When mycobacteria are recovered in clinical specimens, timely species-level identification is required to establish the clinical significance of the isolate and facilitate optimization of antimicrobial therapy. Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has recently been reported to be a reliable and expedited method for identification of mycobacteria, although various specimen preparation techniques and databases for analysis are reported across studies.

This study compared two MALDI-TOF MS instrumentation platforms and three databases: Bruker Biotyper Real Time Classification 3.1 (Biotyper), VITEK® MS Plus Saramis Premium (Saramis), and VITEK® MS v3.0. Two sample preparation techniques were evaluated and the study demonstrated that extraction methods are not interchangeable across different platforms or databases. Once testing parameters were established, a panel of 157 mycobacterial isolates (including 16 Mycobacterium tuberculosis isolates) was evaluated, demonstrating that with the appropriate specimen preparation, all three methods provide reliable identification for most species.

Using a score cutoff value of ≥1.8, the Biotyper correctly identified 133 (84.7%) isolates with no misidentifications. Using a confidence value of ≥90%, Saramis correctly identified 134 (85.4%) isolates with one misidentification and VITEK® MS v3.0 correctly identified 140 (89.2%) isolates with one misidentification. The levels of accuracy were not significantly different across the three platforms (P= 0.14). In addition, VITEK® MS v3.0 requires fewer repeat analyses than the Biotyper and Saramis methods (P= 0.04), which may have implications for laboratory workflow.

“This report represents the first description of the VITEK® MS v3.0 platform for identification of mycobacterium spp. and suggests that the VITEK® MS v3.0 may offer modest advantages over the Biotyper and Saramis, especially by reducing the necessity of

repeat identification attempts.”

KEY POINTS

➔ Even with authors’ modification of interpretation scores, VITEK® MS still outperformed Bruker Biotyper with 91.7% vs. 81.5% at scores ≥ 80% and ≥1.8, respectively. Using the manufacturer-recommended thresholds, there was an even greater difference in performance, i.e., only 70.7% with Bruker Biotyper at scores ≥ 2.0.

➔ A challenge set of 40 isolates showed better performance with 7H10 vs. LJ agar; 92.5% vs. 82.5% for Bruker Biotyper and 97.5% and 87.5% for VITEK® MS, respectively.

➔ VITEK® MS required nearly 50% less repeat extractions compared to Bruker Biotyper.

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JOURNAL OF CLINICAL MICROBIOLOGY 2014;52(12):4286-92

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry using the VITEK®

MS system for rapid and accurate identification of dermatophytes on solid cultures.

De Respinis S1, Monnin V2, Girard V2, Welker M2, Arsac M2, Cellière B2, Durand G2, Bosshard PP3, Farina C4, Passera M4, Van Belkum A2, Petrini O5, Tonolla M6.

(1) Laboratory of Applied Microbiology, University of Applied Sciences and Arts of Southern Switzerland, Bellinzona, Switzerland sophie.derespinis@supsi.ch.

(2) bioMérieux SA, Unit Microbiology, R&D Microbiology, La Balme Les Grottes, France.(3) Department of Dermatology, University Hospital of Zurich, Zurich, Switzerland.(4) Microbiology and Virology Unit, Papa Giovanni XXIII Hospital, Bergamo, Italy.

(5) POLE Pharma Consulting, Breganzona, Switzerland.(6) Laboratory of Applied Microbiology, University of Applied Sciences and Arts of Southern Switzerland, Bellinzona, Switzerland.

Identification of dermatophytes, a type of filamentous fungi, remains particularly challenging, and is usually based on macroscopic and microscopic observations. Morphological identification is a slow and complex process requiring highly skilled mycologists, and misidentifications may occur, even in experienced reference laboratories. Molecular identification is limited due to the cost and the time needed for sequencing. MALDI-TOF MS appears to be a promising method for dermatophyte identification but the heterogeneous morphological phenotypes of fungi, the impact of culture medium, as well as the duration of incubation affect the identification process.

In this study, a simple and rapid sample preparation was established to build the VITEK® MS database using a variety of strains, incubation times and growth conditions. A total of 1130 spectra acquired from 134 well-characterized strains belonging to 17 species were used to create the database. The strains were incubated on two different media from 5 to 10 days to capture growth heterogeneity.

The database was then evaluated using 131 clinical isolates. 125 out of the 131 isolates (95.4%) were identified when grown on sabouraud dextrose agar and 107 (81.7%) of the same isolates were identified when grown on sabouraud supplemented with antibiotics. On average, 88.5% of the isolates were identified to the species level. Some cross-identification was observed between Trychophyton rubrum and Trycophyton soudanense, due to the close relationship between those two taxa.

In conclusion, the study demonstrated that an accurate identification of dermatophytes at different stages of growth can be achieved using the VITEK® MS and a robust spectra library.

“Our study has demonstrated that the inactivation/extraction method is rapid and reliable and that the newly built VITEK® MS knowledge base provides a robust

tool for the reproducible identification to the species level of dermatophytes grown on common routine media.”

➔ Identification of dermatophytes using the VITEK® MS is possible with a rapid sample preparation protocol and a robust database.

➔ The method is suitable for research and diagnostic work with dermatophytes and filamentous fungi in general.

➔ The overall identification performance is high with good species differentiation.

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JOURNAL OF CLINICAL MICROBIOLOGY 2014;52(10):3654-3659

Rapid Inactivation of mycobacterium and nocardia species before identification using MALDI-TOF mass

spectrometry.W. Michael Dunne, Jr.a Kirk Doing,a Elizabeth Miller,a Eric Miller,a Erik Moreno,a Mehdi Baghli,a Sandrine Mailler,b

Victoria Girard,b Alex van Belkum,b Parampal Deolaa

(a) bioMérieux, Inc., Durham, North Carolina, USA;(b) bioMérieux, La Balme Les Grottes, France;

Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) is a reliable and rapid means for the identification of bacterial species, including mycobacteria. However, inactivation of the mycobacteria is essential prior to identification. This study describes a rapid, safe and convenient inactivation protocol, compatible with identification of mycobacterium and nocardia species, for use with MALDI-TOF MS.

The study included 28 strains covering 13 mycobacterium species, including five strains of M. tuberculosis with varied isoniazid (INH) and rifampin (RIF) susceptibility. In addition, five species of nocardia (N. cyriacigeorgica, 9 strains; N. farcinica, 4 strains; N. kruczakiae, 2 strains; N. nova, 7 strains; and N. otitidiscaviarum, 1 strain) were tested.

The authors demonstrated that a 15-minute inactivation protocol (a 5 minute bead-beating step in 70% ethanol (EtOH) followed by a 10 minute incubation period at room temperature) was rapidly bactericidal for a wide variety of mycobacterium and nocardia species and strains.

This protocol provided high-quality spectra compared to spectra obtained directly from growth on solid media and was selected by the authors to be used for all future studies involving the identification of mycobacteria using MALDI-TOF mass spectrometry.

“… a simple, rapid, and reliable [mycobacterial] inactivation process is essential for achieving the reproducibility of identification within a safe environment.”

KEY POINTS

➔ An effective and convenient 15-minute mycobacterial inactivation protocol compatible with identification of mycobacterium and nocardia species using MALDI-TOF MS has been developed.

➔ This protocol provides high-quality spectra compared to spectra obtained directly from growth on solid media.

➔ The inactivation process developed in this study allows safe inactivation for further handling of mycobacteria and nocardia.

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VITEK® MS IVD V2.0(In Vitro Diagnostic Applications)

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VITEK® MS IVD V2.0

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KEY POINTS

VITEK® MS IVD V2.0

➔ The use of VITEK® MS allows a rapid identification of aerobic bacteria and yeasts in clinical practice.

➔ VITEK® MS identified 95.8% of the isolates in the routine workload of an Indian hospital when evaluated on more than 10,000 prospective isolates.

EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2016; doi:10.1007/s10096-016-2864-9

Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) for rapid identification of micro-organisms in the routine clinical

microbiology laboratory.Wattal C, Oberoi JK, Goel N, Raveendran R, Khanna S.

Sir Ganga Ram Hospital, New Delhi, India

Early and accurate microbial identification is a critical requisite for early, adequate antibiotic treatment. Bacterial identification directly from colonies and samples using matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry (MS) has been described as a revolutionary tool perfectly adapted to the clinical microbiology laboratory. The aim of the study was to evaluate the performance of the MALDI-TOF VITEK® MS system in a routine diagnostic microbiology laboratory from a hospital in India.

In this study, a total of 12,003 isolates including bacteria, fungi and mycobacteria received during a one year period were identified using MALDI-TOF. The VITEK® MS identified 11,509 isolates (95.8%) to the species level. No ID was obtained in 2.93% of the cases (352 isolates). The table below describes the percentage of good ID obtained for each group of organisms.

Out of the 1,248 fungal isolates tested, 1,175 (92.6%) were identified using VITEK® MS.

In conclusion, VITEK® MS is a robust tool to identify the isolates encountered in a routine microbiology laboratory in India.

“We found that MALDI-TOF MS could accurately identify bacteria and yeasts to both the genus and species level for gram positive and

gram negative bacteria as well as commonly encountered yeasts”

KEY POINTS

➔ Most of the isolates (92.5%) belonging to species claimed in the database were identified at the species level, showing the robustness of this method.

➔ An additional sample preparation step such as on-plate formic acid treatment or ethanol extraction is not needed to obtain good performance on Actinomycetaceae compared to other systems.

➔ The identification of Actinomycetaceae using VITEK® MS is rapid, reliable, cost-effective and easy to implement in the laboratory.

JOURNAL OF CLINICAL MICROBIOLOGY 2016;54(3):782-4.

Comparison of VITEK® MS and MALDI Biotyper for identification of Actinomycetaceae of clinical importance.

Ferrand J1, Hochard H2, Girard V3, Aissa N4, Bogard B4, Alauzet C4, Lozniewski A4.(1) Laboratoire de Bactériologie, Centre Hospitalier Universitaire de Nancy, Nancy, France

(2) Laboratoire de Biologie Médicale, Centre Hospitalier Régional de Metz-Thionville, Metz, France(3) bioMérieux, R&D Microbiologie, La Balme les Grottes, France

(4) Laboratoire de Bactériologie, Centre Hospitalier Universitaire de Nancy, Nancy, France.

Anaerobic bacteria such as Actinomycetaceae are the main colonizers of human mucosal membranes. They can be predominantly found in the oral cavity and in the intestinal and genital tracts, causing infection when these barriers are damaged. Their detection in a routine microbiology laboratory might be difficult due to their special culture requirements. In addition, conventional bacteriological for anaerobic identification is time-consuming and not always effective.

Over the past years, MALDI-TOF MS technology has proven to be a rapid and inexpensive method for the accurate identification of different microorganisms. The aim of this study is to compare two MALDI-TOF systems for the identification of clinically relevant isolates of the Actinomycetaceae using different pretreatment protocols of the colonies: direct deposit (DD), on-plate formic acid treatment (DD-FA) and ethanol-formic acid extraction (EXT).

A total of 158 strains were tested on both the Bruker Biotyper (IVD version with 5627 entries) and the VITEK® MS IVD V2 with the three sample preparation protocols mentioned above. The conclusions are as follows:

• When direct transfer was used, the rates of correct identification were higher with the VITEK® MS system than with the Biotyper system at the species level (92.5% vs 28.5%) when considering solely the species included in the database.

• Neither the on-plate formic acid treatment nor the extraction process modified the rate of correct identification for VITEK® MS but provided higher identification rates at the genus and species level for the Biotyper.

In conclusion, the two systems were globally effective in identifying Actinomycetaceae. The Biotyper system requires an additional on-plate extraction step to reach acceptable performance levels.

“When direct transfer was used, the rate of species identification was significantly lower with the Biotyper system, despite

its higher species coverage, than that obtained with the VITEK® MS system”

Group of organisms Percentage of good ID to the genus and/ or species level

Enterobacteriacae 97.3%

Gram Negative bacteria other than Enterobacteriacae

98.1%

Enterococci 98.5%

Streptococci and related species 89.9%

Staphylococci and related species 98.3%

Gram positive aerobic bacilli 95%

17

19

KEY POINTS

VITEK® MS IVD V2.0

18

KEY POINTS

VITEK® MS IVD V2.0

➔ VITEK® MS can differentiate pneumococci from pseudopneumococci with high accuracy.

➔ VITEK® MS is a powerful tool to identify streptococci from the mitis group.

DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE 2016;85(1):9-11

MALDI-TOF mass spectrometry for differentiation between Streptococcus pneumoniae and Streptococcus

pseudopneumoniae.van Prehn J1, van Veen SQ2, Schelfaut JJ3, Wessels E3.

(1) Department of Medical Microbiology and Infection Control, VU University Medical Center, Amsterdam, The Netherlands; Department of Medical Microbiology, Onze Lieve Vrouwe Gasthuis, Amsterdam, The Netherlands.

(2) Department of Medical Microbiology, Onze Lieve Vrouwe Gasthuis, Amsterdam, The Netherlands.(3) Department of Medical Microbiology, Leiden University Medical Center, Leiden, The Netherlands

Reliable distinction of Streptococcus pneumoniae from other streptococci is important because of the different pathogenic properties of these organisms. Differentiation between S. pneumoniae and other closely related streptococci has always been challenging, even when using such modern methods as 16S rRNA gene sequencing or matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry.

In this study, two MALDI-TOF platforms, Bruker Biotyper (v 3.1.66, MBT DB-2567) and VITEK® MS IVD V2, were evaluated for their ability to identify a panel of 37 streptococci strains (11 pneumococci, 13 pseudopneumococci and 13 S.mitis/oralis). The direct smear method was used and interpretation of the identification results was made according to the manufacturer’s instructions.

Using the VITEK® MS platform, all strains were identified correctly except one S.mitis/oralis and one S.pneumoniae.

Using the Bruker platform, all S.mitis/oralis and 9 out of 11 S.pneumoniae were correctly identified but none of the S.pseudopneumoniae were correctly identified.

In conclusion, mass spectrometry is able to identify streptococci from the mitis group.

“Mass spectrometry has the ability to adequately distinguish relevant species within the mitis group streptococci. However, the Microflex platform

could not differentiate between pneumococci and pseudopneumococci whereas the VITEK® MS could”

➔ VITEK® MS can provide a very good alternative to phenotypic methods for VGS identification.

FRONTIERS IN MICROBIOLOGY2016;7:1328

Using Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) Complemented with Selected 16S rRNA and gyrB Genes Sequencing to Practically Identify Clinical Important Viridans Group Streptococci (VGS).

Zhou M1,2, Yang Q1, Kudinha T3, Zhang L1, Xiao M1, Kong F4, Zhao Y1, Xu YC5.(1) Department of Clinical Laboratory, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing, China

(2) Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China(3) School of Biomedical Sciences, Charles Sturt University, Orange, NSW, Australia

(4) Centre for Infectious Diseases and Microbiology Laboratory Services, Westmead Hospital, Westmead, NSW, Australia(5) Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, Beijing, China

The viridans group streptococci (VGS) are a heterogeneous group of organisms which exist as commensals in the oropharynx and the gut. They are capable of causing serious diseases when they gain entrance to sites that are usually sterile. Today, there are thirty recognized species of VGS which are classified into six major groups. Species differentiation of these organisms remains a challenge because of the closeness of the species within this group. Phenotypic identification methods have only 30 to 80% accuracy and sequence analysis targeting different genes has been used with various degrees of success. In this study, the performance of two MALDI-TOF systems, Bruker Biotyper and VITEK® MS V2 was evaluated for the identification of the species within the VGS group, using sequencing as a gold standard.

Spectra from 181 clinically relevant VGS isolates from different sources were acquired on both MALDI-TOF systems.

The results are summarized in the table below:

In conclusion, MALDI-TOF can provide a good alternative to phenotypic methods for VGS identification.

“Overall, the VITEK® MS system performed better than the Bruker Biotyper, accurately identifying 98.9% of the 181 VGS isolates”

Percentage of correct identification to the

species level

Percentage of correct identification to the

group levelPercentage of

misidentificationPercentage of No

identification

Bruker Biotyper 88.4% 0% 11.6% 0%

VITEK® MS 98.9% 0.55% 0.33% 0.55%

21

KEY POINTS

VITEK® MS IVD V2.0

20

KEY POINTS

VITEK® MS IVD V2.0

➔ Both initial and repeat spectra obtained from the strains are highly reproducible within and between laboratories.

➔ MALDI-TOF mass spectrometry-based microbial identification is highly reproducible in the setting of a clinical microbiology laboratory.

JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(7):2349-52

Assessment of Reproducibility of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

for Bacterial and Yeast Identification.Westblade LFa,b,c, Garner OBd, MacDonald Ke, Bradford Cf, Pincus DHf, Mochon ABd,g,h, Jennemann Ri, Manji Rj, Bythrow

Mj, Lewinski MAd,k, Burnham CDc and Ginocchio CCe,l.(a) Department of Pathology, Children’s Healthcare of Atlanta, Atlanta, Georgia, USA

(b) Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia, USA(c) Department of Pathology and Laboratory Medicine, Washington University in St. Louis School of Medicine, St. Louis, Missouri, USA

(d) Department of Pathology and Laboratory Medicine, David Geffen School of Medicine at UCLA, Los Angeles, California, USA(e) bioMérieux, Inc., Durham, North Carolina, USA

(f) bioMérieux, Inc., Hazelwood, Missouri, USA(g) Division of Infectious Diseases, Laboratory Sciences of Arizona/Sonora Quest Laboratories, Phoenix, Arizona, USA

(h) Department of Pathology, University of Arizona College of Medicine, Phoenix, Arizona, USA(i) Barnes-Jewish Hospital, St. Louis, Missouri, USA

(j) Department of Pathology and Laboratory Medicine, North Shore-LIJ Laboratories, Lake Success, New York, USA(k) Roche Molecular Systems, Inc., Pleasanton, California, USA

(l) Hofstra North Shore-LIJ School of Medicine, Hempstead, NY, USA

Identification of microorganisms using MALDI-TOF mass spectrometry is being used in laboratories all over the world based on the abundance of data showing very good identification performance. However, there is very limited data on the reproducibility of MALDI-TOF for microorganism identification. This multicenter study assessed the ability of the VITEK® MS version 2.0 to reproducibly identify bacterial and yeast isolates in three laboratories.

Over a five-day period, reproducibility studies were performed in three different laboratories in the US, one in New York, one in Missouri, and one in California. A collection of well characterized strains was used and the testing was performed by two different operators at each site. Three different lots of matrix solution, formic acid (for yeast extraction), and target slides were used by the operators.

For all sites, 598/600 (99.7%) initial identification results were correct to the species level. No isolates were misidentified. In one instance, a Candida albicans and a C. glabrata were not identified and in one instance a Klebsiella pneumoniae and a K. oxytoca were not identified.

On repeat testing of the 598 isolates, 595 isolates (99.5%) were correctly identified on the initial (and only) read. One isolate of Proteus mirabilis and one isolate of Pseudomonas aeruginosa were unidentified, but there were no misidentifications.

“We unambiguously demonstrate that bacteria and yeast identification using the VITEK® MS v2.0 system is highly reproducible and can tolerate

numerous variables including different testing environments, instruments, operators, reagent and target slide lots,

and sample positions.”

➔ VITEK® MS V2.0 is a rapid and reliable method to identify Gram-positive cocci in the clinical laboratory.

➔ VITEK® MS V2.0 is able to reliably distinguish Streptococcus pneumoniae from Streptococcus mitis/oralis.

JOURNAL OF MICROBIOLOGICAL METHODS2015;113:13–15

Comparative evaluation of two matrix-assisted laser desorption ionization time-of-flight mass spectrometry

(MALDI-TOF MS) systems, VITEK® MS and Microflex LT, for the identification of Gram-positive cocci routinely

isolated in clinical microbiology laboratories.Lee Ma, Chung HSa, Moon HWb, Lee SHc, Lee Kd.

(a) Department of Laboratory Medicine, Ewha Womans University School of Medicine, Republic of Korea(b) Department of Laboratory Medicine, Konkuk University School of Medicine, Republic of Korea

(c) Seegene Medical Foundation, Republic of Korea(d) Department of Laboratory Medicine, Research Institute of Antimicrobial Resistance, Yonsei University College of Medicine, Republic of Korea

The performance of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) for routine bacterial and yeast identification has been thoroughly evaluated but data on gram-positive cocci are limited.

In this study, two commercial MALDI-TOF MS devices were evaluated for the identification of 394 Gram-positive cocci collected over a 4-month period in Korea. 16S rRNA molecular identification was conducted to resolve discrepancies.

Overall, VITEK® MS V2.0 and Microflex LT correctly identified 97.2% and 94.7% of the isolates respectively. The percentage of correctly identified species for each group of organisms is detailed below:

In conclusion, VITEK® MS offers a reliable and rapid identification of clinically important Gram-positive cocci isolated in clinical laboratories.

“VITEK® MS was able to identify all S.pneumoniae and S.mitis/oralis isolates”

Organism group Percentage of correct identification to the species level

VITEK® MS Microflex LT

Staphylococci 97.2% 94.7%

Streptococci 96% 92.1%

Enterococci 100% 100%

23

KEY POINTS

VITEK® MS IVD V2.0

22

KEY POINTS

VITEK® MS IVD V2.0

➔ VITEK® MS shows a better identification accuracy compared to classical phenotypic identifications

➔ VITEK® MS is adapted to the routine identification of organisms commonly found in clinical settings.

BIOMEDICAL RESEARCH INTERNATIONAL 2015 Dec 17. doi:10.1155/2015/516410

Performances and Reliability of Bruker Microflex LT and VITEK® MS MALDI-TOF Mass Spectrometry Systems for

the Identification of Clinical Microorganisms.Bilecen K1, Yaman G2, Ciftci U3, Laleli YR3.

(1) Department of Research & Development, Duzen Laboratories Group, 06680 Ankara, Turkey(2) Department of Clinical Microbiology, Duzen Laboratories Group, 34387 Istanbul, Turkey(3) Department of Clinical Microbiology, Duzen Laboratories Group, 06680 Ankara, Turkey

Conventional identification methods rely on biochemical results and require lengthy incubation procedures. In comparison, MALDI-TOF MS can identify bacteria and yeast within minutes directly from colonies grown on culture plates. The aim of this study was to evaluate the performance of the two MS platforms currently marketed, Bruker Biotyper and VITEK® MS, on a total of 1341 isolates from a Turkish hospital.

The results for bacteria identification are summarized in the table below:

The results for yeast are summarized in the table below:

In conclusion, a correct species level identification of over 96% was obtained for bacteria and yeast on both MS platforms.

“Our results indicated that, a species level correct identification accuracy of over 96% could be achieved without any

prior information about the tested samples”

BACTERIAPercentage of

identification to the species level

Percentage of identification to the

genus level onlyPercentage of

misidentification

VITEK® MS 97.12% 1.61% 0.42%

Bruker 96.70% 2.37% 0.33%

YEASTPercentage of

identification to the species level

Percentage of identification to the

genus level onlyPercentage of

misidentification

VITEK® MS 98.12% 1.88% 0%

Bruker 97.50% 2.50% 0%

EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2014;33(10):1703-12

Comparison and optimization of two MALDI-TOF MS platforms for the identification of medically

relevant yeast species.Pence MA, McElvania TeKippe E, Wallace MA, Burnham CD.

Department of Pathology and Immunology, Division of Laboratory and Genomic Medicine, Washington University School of Medicine, St. Louis, MO, USA

Accurate species identification of yeasts is important for initiating early effective antifungal therapy, especially when susceptibility testing results are not promptly available. Furthermore, the emergence of less-common or “cryptic” non-albicans Candida species, as well as uncommon yeasts increases the need for a rapid and effective method for identification of those organisms. In this study, two matrix assisted laser desorption – time-of-flight (MALDI-TOF) platforms, VITEK® MS V2.0 and Bruker Biotyper version 3.1, were evaluated for the identification of 117 isolates of clinically relevant yeasts. Sequence analysis was used as the gold standard for isolate identification. Isolates were tested after 24h and 48h of incubation on four different media and spectra were acquired using a direct smear method with formic acid (FA) on-plate extraction.

The percentages of correct identification using the different methods and following manufacturer’s instructions for result interpretation are summarized below:

The study shows that additional incubation time has no impact on yeast identification. The media used to grow the isolates did not lead to significant differences in the identification rates.

In conclusion, this study demonstrated that MALDI-TOF is a rapid and reliable method to identify yeasts when used with on-plate extraction.

“While both MALDI-TOF platforms rapidly and accurately identify medically relevant yeast compared to traditional phenotypic methods,

VITEK® MS identified significantly more isolates than Biotyper using the direct smear method.”

➔ In-tube formic acid extraction is not needed to identify yeasts with the VITEK® MS system in contrast to the Bruker Biotyper.

➔ Neither the incubation time nor the medium used to grow the strains had any impact on the identification results with the VITEK® MS.

Organism group Percentage of correct identification to the species level

Direct smear with 25% FA Direct smear with 100% FA

VITEK® MS V2.0 94.9% Not tested

Bruker Biotyper 3.1 30.8% 53%

VITEK® MS IVD V2.0 – MULTICENTER EVALUATIONS

27

KEY POINTS

VITEK® MS IVD V2.0 – MULTICENTER EVALUATION

26

KEY POINTS

VITEK® MS IVD V2.0 – MULTICENTER EVALUATION

KEY POINTS

26

➔ VITEK® MS v2.0 showed similar or better performance than conventional biochemical tests for the identification of fastidious gram-negative bacteria.

DIAGNOSTIC MICROBIOLOGY AND INFECTIOUS DISEASE2014;78(2):129-31

Multicenter validation of the VITEK® MS v2.0 MALDI-TOF mass spectrometry system for the

identification of fastidious gram-negative bacteria.Branda JA1, Rychert J2, Burnham CA3, Bythrow M4, Garner OB5, Ginocchio CC6, Jennemann R7, Lewinski MA5, Manji R4,

Mochon AB5, Procop GW8, Richter SS8, Sercia LF8, Westblade LF9, Ferraro MJ2.(1) Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.(2) Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.

(3) Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO, USA.(4) Department of Pathology and Laboratory Medicine, North Shore-LIJ Health System Laboratories, Lake Success, NY, USA.(5) Department of Pathology and Laboratory Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA, USA.

(6) Department of Pathology and Laboratory Medicine, North Shore-LIJ Health System Laboratories, Lake Success, NY, USA; Department of Pathology and Laboratory Medicine, Hofstra North Shore-LIJ School of Medicine, Hempstead, NY, USA.

(7) Barnes Jewish Hospital, St. Louis, MO, USA.(8) Department of Clinical Pathology, Cleveland Clinic, Cleveland, OH, USA.

(9) Barnes Jewish Hospital, St. Louis, MO, USA; Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO, USA; Department of Pathology and Laboratory Medicine, Hofstra North Shore-LIJ School of Medicine, Hempstead, NY, USA.

This multi-center study involving five US-based sites aimed to determine the performance of the VITEK® MS v2.0 MALDI-TOF mass spectrometry system in identifying fastidious gram-negative bacteria.

The study included 226 clinical isolates, representing 9 genera and 15 species of fastidious gram-negative bacteria. H. influenzae, H. parainfluenzae, C. jejuni, L. pneumophila, M. catarrhalis and N. gonorrhea were among the species included.

Compared with the reference method used (DNA sequencing), the VITEK® MS system provided a single, accurate, species-level identification for 96% (216/226) of the study isolates, and accurate genus-level identification for an additional 1%. This performance was comparable to – or better than – reported performance for traditional biochemical testing.

“The ability to identify and rapidly report a specific identification for these [fastidious gram-negative] bacteria will enhance the capabilities of

many laboratories and allow clinicians to use targeted therapy, which will aid antimicrobial stewardship.”

EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES2014;33(3):337-46

Multi-center evaluation of the VITEK® MS system for mass spectrometric identification of

non-Enterobacteriaceae Gram-negative bacilli.Manji R, Bythrow M, Branda JA, Burnham CA, Ferraro MJ, Garner OB, Jennemann R, Lewinski MA, Mochon AB, Procop

GW, Richter SS, Rychert JA, Sercia L, Westblade LF, Ginocchio CC.Department of Pathology and Laboratory Medicine, North Shore-LIJ Health System Laboratories, NY, USA

The performance of the VITEK® MS v2.0 system was evaluated for the identification of the non-Enterobacteriaceae Gram-negative bacilli (NEGNB); a diverse group of pathogens that cause a variety of infections primarily pulmonary, urinary, skin and soft tissue, catheter-related, bloodstream, and gastrointestinal. This study was a multicenter evaluation performed at five US sites that included 558 unique NEGNB clinical isolates, representing 18 genera and 33 species.

The VITEK® MS v2.0 provided an identification for 516 out of 558 of the NEGNB tested. VITEK® MS v2.0 correctly identified 507/558 (90.9%) of the NEGNB isolates, 77.8% to the species level and 13.1% to the genus level. Four isolates (0.7%) were incorrectly identified to the genus level and five isolates (0.9%) were incorrect to the species level. The remaining 42 isolates (7.5%) were either reported as no identification (5.0%) or called “mixed genera” (2.5%). All results obtained with the VITEK® MS v2.0 were compared with reference 16SrRNA gene sequencing and when indicated recA sequencing and phenotypic analysis.

With the VITEK® MS system, the rate of incorrect identifications was low, both at the genus and species levels. The results establish that the VITEK® MS v2.0 system provides accurate identifications for NEGNB.

“The VITEK® MS v2.0 system provides accurate results for the identification of a challenging and diverse group of Gram-negative bacteria.”

➔ For NEGNB, VITEK® MS v2.0 provided an identification for 516 out of 558 isolates (92.5%), correctly reporting 507 out of 558 (90.9%).

➔ VITEK® MS v2.0 does not require any pre-processing of NEGNB prior to inoculation of the target slide.

29

VITEK® MS IVD V2.0 – MULTICENTER EVALUATION

28

KEY POINTS

VITEK® MS IVD V2.0 – MULTICENTER EVALUATION

➔ The VITEK® MS is an accurate system for identifying clinically relevant anaerobic bacteria.

➔ Overall, the performance of the VITEK® MS was highly accurate (91.2% correct to species level).

CLINICAL MICROBIOLOGY AND INFECTION 2014;20(4):335-9

Multi-Center Evaluation of Mass Spectrometric Identification of Anaerobic Bacteria Using the VITEK® MS System.

Garner OB, Mochon AB, Branda JA, Burnham CD, Bythrom M, Ferraro MJ, Ginocchio CC, Jennemann R, Manji R, Procop GW, Richter SS, Rychert JA, Sercia L, Westblade LF, Lewinski MA.Department of Pathology and Laboratory Medicine, UCLA, Los Angeles, CA, USA.

A multicenter study, including 5 clinical sites in the United States, evaluated the performance of the VITEK® MS MALDI-TOF with VITEK® MS v2.0 identification database in the identification of clinically relevant anaerobic bacteria in a clinical microbiology laboratory.

TThe study consisted of 651 clinically relevant anaerobic isolates, representing 11 genera and 26 separate species [8 genera of Gram-positive anaerobes (265 isolates) and 3 genera of Gram-negative anaerobes (386 isolates)].

Out of the 651 isolates, 594 (91.2%) were correctly identified to the species level as confirmed by 16S rRNA gene sequencing. An additional 8 isolates were identified correctly to the genus level bringing the total number of isolates correctly identified to the genus level to 92.5%. Thirty six isolates produced a result of no identification.

Identification of anaerobes has classically relied upon phenotypic assays such as Gram staining, growth characteristics and biochemical reactivity patterns that can be both time consuming and laborious. With the implementation of MALDI-TOF mass spectrometry, a routine clinical microbiology laboratory is provided with the opportunity for inexpensive, rapid, and accurate identification of anaerobes. This allows for earlier selection of appropriate therapy for anaerobic infections.

“The implementation of this technology in the clinical microbiology laboratory will lead to decreased turnaround times for identification.”

JOURNAL OF CLINICAL MICROBIOLOGY 2013; 51(7):2225-31

Multicenter Evaluation of the VITEK® MS Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry System for Identification of Gram-Positive

Aerobic Bacteria.Rychert J, Burnham CD, Bythrow M, Garner OB, Ginocchio CC, Jennemann R, Lewinski MA, Manji R, Mochon A, Procop

GW, Richter SS, Sercia L, Westblade LF, Ferraro MJ, Branda JA.Department of Pathology, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts, USA.

MALDI-TOF mass spectrometry is gaining acceptance in clinical microbiology laboratories for the rapid identification of a wide range of microorganisms. A multicenter study was performed to evaluate the VITEK® MS System v2.0 for the identification of aerobic Gram-positive bacteria.

A total of 1,146 unique isolates (13 genera, 42 species) were analyzed, and results were compared to nucleic acid sequence-based identifications. For 1,063 of 1,146 isolates (92.8%), the VITEK® MS gave an accurate identification to the species level. For an additional 31 isolates (2.7%), the identifications proposed were all correct to the genus level with multiple possible identifications. Mixed genus or single choice incorrect identifications were given for 18 isolates (1.6%). No identification was obtained for 33 isolates (2.9%), but there was no specific species where the VITEK® MS consistently failed to provide an identification.

When considering a subset of organisms that represent commonly encountered, important pathogens, the VITEK®

MS correctly identified 95% of the isolates to the species level with no misidentifications. Also, the VITEK® MS was able to correctly differentiate Streptococcus pneumoniae from other viridans Streptococci. The current study shows the VITEK® MS System to be highly accurate for the identification of Gram-positive aerobic bacteria in the clinical laboratory setting.

“...the VITEK® MS v2.0 uses a “bin matrix” approach… a method that has been shown to aid in differentiating between very similar mass spectra

derived from closely related species or subspecies.”

KEY POINTS

➔ The VITEK® MS system is highly accurate for the identification of Gram-positive aerobic bacteria in the clinical laboratory setting.

➔ The VITEK® MS v2.0 was able to reliably distinguish between S. pneumoniae and other S. mitis group species.

➔ A stringent approach with strict rules was used on isolates tested. In particular, a single deposit rather than multiple deposits of each isolate was spotted on the target plate, and protein extraction was not performed.

2928

31

VITEK® MS IVD V2.0 – MULTICENTER EVALUATION

30

KEY POINTS

VITEK® MS IVD V2.0 – MULTICENTER EVALUATION

➔ MALDI-TOF MS is a rapid and highly accurate method for yeast identification.

➔ The major advantage in using MALDI-TOF MS for identification of yeasts compared to conventional methods is the noticeable decreases in cost and time to identification.

JOURNAL OF CLINICAL MICROBIOLOGY 2013; 51(7):2267-72

Multicenter Study Evaluating the VITEK® MS System for Identification of Medically Important Yeasts.

Westblade LF, Jennemann R, Branda JA, Bythrow M, Ferraro MJ, Gerner OB, Ginocchio CC, Lewinski MA, Manji R, Mochon AB, Procop GW, Richter SS, Rychert JA, Sercia L, Burnham CD.

Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

The performance of the VITEK® MS System and VITEK® MS v2.0 database was assessed for identifying medically important yeasts in a multicenter study that included five clinical microbiology laboratories located in five different geographical regions in the United States.

The study consisted of 852 yeast isolates including clinically relevant Candida species and non-Candida species. In total, 31 different species were evaluated.

Out of the 852 isolates, 823 isolates (96.6%) were identified to the genus level and 819 (96.1%) were identified to the species level. Twenty-four isolates (2.8%) were not identified, and five isolates (0.6%) were misidentified.

Organism identification of yeasts is often sufficient to initiate appropriate empirical antifungal therapy without wainting for antibiotic susceptibility results. Therefore, rapid and highly accurate identification of yeast isolates using MALDI-TOF MS will be able to enhance patient care and reduce hospital-associated costs due to fungal infections.

“Implementation of this methodology should streamline yeast identification in the laboratory, positively affect

patient care, and reduce health care-associated costs.”

EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES 2013;32(12):1571-8

Identification of Enterobacteriaceae by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

using the VITEK® MS system.Richter SS, Sercia L, Branda JÁ, Burnham CD, Bythrow M., Ferraro MJ, Garner OB, Ginocchio CC, Jennemann R, Lewinski

MA, Manji R, Mochon AB, Rychert JA, Westblade LF, Procop GWDepartment of Clinical Pathology, Cleveland Clinic, 9500 Euclid Avenue/LL1-2, Cleveland, OH

Identification of Enterobacteriaceae on the VITEK® MS system (database v2.0) was evaluated in this multicenter study including 5 participating study sites. The study consisted of 965 Enterobacteriaceae isolates, representing 17 genera and 40 species.

All isolates were analyzed on the VITEK® MS system in accordance with the manufacturer’s instructions, and accuracy of the mass spectrometric identifications was compared to16S rRNA gene sequencing as well as supplemental phenotypic testing for organisms unidentified by sequencing.

965 isolates tested, 96.7% of the VITEK® MS results agreed with the reference method identification (83.8% correct to the species level and 12.8% limited to genus-level identification). There was no identification for 1.7% of the isolates. The VITEK® MS system misidentified 7 isolates (0.7%) as different genera.

This is the first report of VITEK® MS system performance for identification of Enterobacteriaceae typically encountered in the clinical laboratory using database v2.0 and MYLA® software developed for in vitro diagnostic (IVD) use. The VITEK® MS IVD system provided accurate results for a large and diverse collection of Enterobacteriaceae clinical isolates in a more clinically relevant time frame compared to current biochemical identification systems.

“This multicenter study demonstrated that identifications provided by the VITEK® MS IVD system for Enterobacteriaceae are highly accurate in comparison to a molecular reference method.”

KEY POINTS

➔ This study used an efficient workflow (single spotting of isolates, no extraction step) that minimized the hands-on time required.

➔ The large number of Enterobacteriaceae isolates from different geographic regions of the US and the use of a reference molecular method for all isolates is a predominant strength of this study.

➔ The VITEK® MS result agreed with the reference method for 96.7% of the 965 isolates tested.

31

VITEK® MS RUO (RESEARCH USE ONLY)

35

KEY POINTS

VITEK® MS RUO (Research Use Only)VITEK® MS RUO (Research Use Only)

KEY POINTS

➔ VITEK® MS can be used to identify bacteria directly from positive blood cultures when combined with sample preparation based on lysis centrifugation.

➔ VITEK® MS identification of bacteria present in positive blood culture bottles is an efficient method for the rapid identification of bacterial isolates in the clinical microbiology laboratory.

JOURNAL OF INFECTION AND PUBLIC HEALTH2016 Jun 13. pii: S1876-0341(16)30068-5

Comparison among four proposed direct blood culture microbial identification methods using MALDI-TOF MS.

Bazzi AM1, Rabaan AA2, El Edaily Z3, John S3, Fawarah MM3, Al-Tawfiq JA3.(1) Microbiology Laboratory, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia. Electronic address: ali.bazzi@jhah.com.

(2) Molecular Diagnostic Laboratory, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia.(3) Microbiology Laboratory, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia.

(4) Specialty Internal Medicine, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia; Indiana University School of Medicine, Indiana, USA.

Sepsis is one of the leading cause of death worldwide and it is crucial to start appropriate antimicrobial treatment as quickly as possible. Current protocols for microbial identification rely on Gram staining of a positive blood culture followed by subculture to solid media, incubation for 18–24 h and then confirmatory tests to identify the causative organism. Use of a more rapid technology such as MALDI-TOF that can identify a broad range of bacterial species directly from positive blood cultures could help reduce mortality by facilitating early and appropriate antibiotic treatment.

In this study, four different methods using VITEK® MS and starting directly from the positive blood culture bottle were evaluated on 120 positive blood cultures. Three methods were based on lysis and centrifugation followed by washing and chemical treatment of the pellet, either with acetic acid (method 1), formic acid (method 2) or ethanol (method 3).The fourth method was based on short incubation on solid media. The results are summarized in the table below:

In conclusion, all four methods could identify the isolates directly from blood cultures. Methods 1 and 2 are more effective for Gram positive bacteria compared to methods 3 and 4.

“We have confirmed that our rapid identification methods which are based on extraction by lysis centrifugation are highly accurate and precise for a

range of Gram-negative and Gram-positive bacteria.”

34

Percentage of correct identification to the species level

Method 1 Method 2 Method 3 Method 4

Gram negative bacteria 90.9% 90.9% 90.9% 81.8%

Gram positive bacteria 90% 90% 76.7% 86.7%

EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY & INFECTIOUS DISEASES 2015;34(9):1787-95

Rapid urine preparation prior to identification of uropathogens by MALDI-TOF MS.

Veron L., Mailler S., Girard V., Muller B., LHostis G., Ducruix C., Lesenne A., Richez A., Rostaing H., Lanet V., Ghirardi S., Van Belkum A., Mallard F.

Innovation/Technology Research Department, bioMérieux S.A., Chemin de l’Orme, 69280, Marcy L’Etoile, France,

Complicated urinary tract infections (UTI), such as pyelonephritis, may lead to sepsis. Rapid diagnosis is needed to identify the causative urinary pathogen and to guide antimicrobial therapy. UTI are often caused by one bacterial species, Escherichia coli being the most prevalent. Typically, after culturing a specimen, the isolate’s identification and susceptibility are determined in about 18-24 hours. Recently, MALDI-TOF has revolutionized the world of microbiology by shortening the time needed to identify isolates grown on culture medium. But the time needed for culture remains a limiting factor in the time to deliver results.

This study describes a direct–from-sample identification of urinary pathogens using MALDI-TOF, combined with two different sample preparations: differential centrifugation and dual filtration. A short incubation period of urine on a plate was also evaluated. The results of the comparison between the three methods is summarized in the table below:

In conclusion, the most efficient method to identify pathogens from urine is the short growth of the urine on plate for 5 hours.

“The results shown in the present study demonstrate that a short culture is a very simple and efficient sample preparation method allowing fast and reliable identification of uropathogens

by MALDI[-TOF].”

Differential centrifugation Dual filtration Short growth

Correlation of MALDI-TOF MS with reference identification

68.4% 78.9% 84.2%

➔ Short incubation on plate followed by VITEK® MS identification is a reliable and rapid method for the identification of urinary pathogens.

➔ Combined with rapid AST, the use of this rapid method could significantly advance the diagnosis of UTI.

37

VITEK® MS RUO (Research Use Only)

PLoS ONE2014;9(2):e87870129-31

OPEN ACCESS

Same Day Identification and Full Panel Antimicrobial Susceptibility Testing of Bacteria from Positive Blood Culture Bottles Made Possible by a Combined Lysis-Filtration Method with MALDI-TOF VITEK® Mass

Spectrometry and the VITEK® 2 System.Machen A1, Drake T2, Wang YF3.

(1) Department of Pathology and laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia, United States of America.(2) Department of Pathology and Laboratory Medicine, Grady Health System, Atlanta, Georgia, United States of America.

(3) Department of Pathology and laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia, United States of America ; Department of Pathology and Laboratory Medicine, Grady Health System, Atlanta, Georgia, United States of America.

This proof-of-principle study evaluates a Lysis-Filtration Method for identification (ID) of bacteria and yeast directly from positive BACT/ALERT® blood culture bottles using the VITEK® MS RUO (SARAMIS® database) system, and direct antimicrobial susceptibility testing (AST) using the VITEK® 2 system. In addition, the average time to identification and antimicrobial susceptibility testing using this combined method was compared to conventional methods.

A total of 100 non-duplicated positive blood cultures were tested, and 1012 microorganism-antimicrobial combinations were assessed. An aliquot of non-charcoal blood culture broth was incubated with lysis buffer briefly before being filtered and washed. Microorganisms recovered from the filter membrane were first identified using the VITEK® MS. Then a suspension made from the filtered organisms was adjusted to the correct McFarland and tested on the VITEK® 2 system for antimicrobial susceptibility. (Figure 1)

Of 100 bottles tested, the VITEK® MS resulted in 94.0% correct organism identification to the species level and 2.0% correct identification to the genus level. Compared to the conventional AST methods, direct antimicrobial susceptibility testing from VITEK® 2 resulted in 93.5% (946/1012) category agreement of antimicrobials tested, with 3.6% (36/1012) minor error, 1.7% (7/1012) major error, and 1.3% (13/1012) very major error of antimicrobials.

The average time from a blood culture bottle flagging positive to ID and AST was 11.4 hours using the combined Lysis-Filtration method with VITEK® MS and VITEK® 2 compared to 56.3 hours using conventional methods (p<0.00001). (Figure 2)

This study demonstrates the effectiveness of the Lysis-Filtration method combined with VITEK® MS and VITEK® 2 for rapid, direct ID and AST of bacteria in a clinical setting. Furthermore, it indicates that same-day results for microbial ID and AST directly from positive blood culture can be achieved and used for appropriate antibiotic therapy and antibiotic stewardship.

“This proof-of principle study provides a practical clinical method and tools for rapid ID and AST, which have the potential to impact antibiotic

usage and clinical outcomes.”

KEY POINTS

➔ This combined method offers a simplified workflow (Figure 1) which does not require Gram staining, subculture or centrifugation.

➔ Use of the Lysis-Filtration method for both ID and AST (VITEK® MS RUO and VITEK® 2) was shown to be 5 times faster than conventional methods (11.4 hours vs. 56.3 hours) allowing for same-day reporting of AST results directly after the blood culture bottle turns positive (Figure 2).

36

Total

GP

GN

Total

Lysis-Filtration Method ID

Lysis-Filtration Method AST

Standard Methods ID

Standard Methods AST

GP

GN

a. Time (hrs) from when blood culture bottle was flagged as positive

0

b. Time (hrs) from when blood was collected from patient

24 48 72 96

0 24 48 72 96 120 144

Figure 1. Process flow diagram for identification and antimicrobial susceptibility testing of bacteria directly from positive blood culture bottles using the Lysis-Filtration Method with MALDI-TOF VITEK® MS and the VITEK® 2 System.

Blood culture turns positive

Process broth using Lysis-Filtration Method

10-15 minPrepare inoculum

with saline 1-5 min

Antimicrobialsusceptibility testing(AST) by VITEK®2

6-24 hr

Microorganism identification (ID)

by VITEK®MS15 min <1 hr

Prepare VITEK®MSslide with matrix

1-5 min

Figure 2. Average time in hours to results of ID or AST from (a) time when blood culture bottle was flagged as positive or (b) time when blood was collected from patient.

For the Lysis-Filtration Method, ID was performed using VITEK® MS, AST was performed using VITEK® 2.AST = antimicrobial susceptibility testing. GN = Gram-negative organisms; GP = Gram-positive organisms; ID = identification

39

KEY POINTS

VITEK® MS RUO (Research Use Only)

38

KEY POINTS

VITEK® MS RUO (Research Use Only)

MYCOSES 2016;59(8):535-8

Identification and typing of the emerging pathogen Candida auris by matrix-assisted laser desorption

ionisation time of flight mass spectrometry.Girard V1, Mailler S1, Chetry M1, Vidal C2, Durand G1, van Belkum A1, Colombo AL3, Hagen F4, Meis JF4,5, Chowdhary A6.

(1) R&D Microbiology, bioMérieux, La Balme les Grottes, France.(2) R&D Biomathematics, bioMérieux, Grenoble, France.

(3) Laboratório Especial de Micologia, Escola Paulista de Medicina, UNIFESP, São Paulo, Brazil.(4) Department of Medical Microbiology and Infectious Diseases, Canisius Wilhelmina Hospital, Nijmegen, The Netherlands.

(5) Department of Medical Microbiology, Radboud University Medical Center, Nijmegen, The Netherlands.(6) Department of Medical Mycology, Vallabhbhai Patel Chest Institute, University of Delhi, Delhi, India

In the last five years, Candida auris has emerged as an important pathogen capable of causing healthcare-associated infections with a high mortality rate. This yeast is often phenotypically misidentified as Candida haemulonii, Candida famata, Candida sake, Saccharomyces cerevisiae, and Rhodotorula glutinis by commercial identification systems. Currently its identification requires molecular sequencing which is not always available in many laboratories. Therefore, it is imperative for clinical microbiology laboratories to accurately identify this organism to aid in preventing healthcare-associated outbreaks.

In this study, the VITEK® MS database was generated using 12 strains of Candida auris using the on-plate extraction. A collection of 50 unique isolates of C.auris from different countries was analyzed for validation purposes using the same protocol. All isolates were correctly identified to the species level using the expanded database except for four strains that were also found discordant using 18S sequencing. A clear distinction could be made between Candida auris and other common Candida species such as C. haemulonii, C.albicans and C.glabrata. Morevover, C.auris spectra formed distinct groups based on the geographic origin of the strains, showing that MALDI-TOF can be used as a screening tool.

In conclusion, the optimized VITEK® MS database associated with a suitable on-plate sample preparation protocol, enables rapid, reliable and robust identification of the emerging pathogen Candida auris. This study also demonstrates the value of integrating MALDI-TOF MS technology into routine laboratory use to improve patient management.

“We here show that VITEK® MS combined with AFLP [Amplified Fragment Length Polymorphism] allows rapid and accurate identification and

subsequent typing of this emerging pathogen.”

➔ VITEK® MS RUO database can be quickly modified to implement spectra from emerging pathogens therefore allowing their rapid and accurate identification.

➔ VITEK® MS RUO in combination with a molecular typing method (ALFP) allows accurate species identification and epidemiological characterization of strains.

EUROPEAN JOURNAL OF CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASES2016;35:829–838

Comparison of MALDI-TOF MS and AFLP for strain typing of ESBL-producing Escherichia coli.

Veenemans J1, Welker M2, van Belkum A2, Saccomani MC2, Girard V2, Pettersson A3, Verhulst C4, Kluytmans-Vandenbergh M4,5, Kluytmans J4,5.

(1) Laboratory for Microbiology and Infection Control, Amphia Hospital, Breda, 4800 RK, The Netherlands.(2) BioMerieux, Microbiology R&D, La Balme Les Grottes, France.

(3) Department for Medical Microbiology and Infection Control, VU University Medical Centre, Amsterdam, The Netherlands.(4) Laboratory for Microbiology and Infection Control, Amphia Hospital, Breda, 4800 RK, The Netherlands.

(5) Julius Center for Health Sciences and Primary Care, UMC Utrecht, Utrecht, The Netherlands.

Bacterial typing is crucial to limit the spread of bacterial pathogens but current molecular methods are time-consuming and costly. MALDI-TOF Mass Spectrometry has been recognized worldwide as a rapid and powerful tool for identification of bacteria and fungi. Using MALDI-TOF MS for organism typing is interesting as this technology requires minimum sample preparation, analyzes data directly, is quick, automated, and inexpensive. In this context, the rapid differentiation between resistant and susceptible strains appears particularly attractive.

The aim of this study was to evaluate the discrimination power of VITEK® MS RUO to type 52 ESBL-producing E.coli strains involved in an outbreak and collected in four separate buildings of the same nursing home. All isolates were typed by Amplified Fragment Length Polymorphism (AFLP) and spectra were acquired in parallel in quadruplicate using the VITEK® MS RUO system. The four replicates per strain were used to create a consensus spectra in the RUO database by retaining the peaks that were found in three out of four spectra. Based on the spectra obtained, the majority of the isolates were classified into four clusters, MALDI type 1 to MALDI type 4 and some peaks could be clearly associated with one particular type. These clusters matched with different AFLP types. MALDI was also able to identify the strains associated with the outbreak which were gathered in one cluster.

In conclusion, the MALDI-TOF MS typing method is not as powerful as molecular methods but has the advantage of being quick and affordable and could be considered as a first-line epidemiological tool which allows rapid prospective typing.

“The performance of MALDI-TOF MS for the typing of E.coli is promising showing good agreement with typing results obtained by ALFP and other typing methods”

➔ MALDI-TOF MS is a promising, rapid, and inexpensive typing tool.

VITEK® MS RUO (Research Use Only)

40

KEY POINTS

VITEK® MS RUO (Research Use Only)

SYSTEMATIC AND APPLIED MICROBIOLOGY2015;38(5):315-22

Rapid and accurate identification of Streptococcus equi subspecies by MALDI-TOF MS.

Kudirkiene E1, Welker M2, Knudsen NR1, Bojesena AM3.

(1) Department of Veterinary Disease Biology, University of Copenhagen, Stigbøjlen 4, 1870 Frederiksberg C, Denmark.(2) BioMérieux SA, Unit Microbiology, R&D Microbiology, La Balme Les Grottes 38390, France.

(3) Department of Veterinary Disease Biology, University of Copenhagen, Stigbøjlen 4, 1870 Frederiksberg C, Denmark.

Strangles, a severe respiratory tract infection caused by Streptococcus equi spp equi, remains the most frequently diagnosed infection of horses. It is therefore important to distinguish Streptococcus equi spp equi (SEE) from the other two subspecies, Streptococcus equi spp zooepidemicus (SEZ) and Streptococcus equi spp ruminatorum (SER) which are considered to be opportunistic pathogens in horses. Identification of these three subspecies based on biochemical tests remains a challenge and molecular identification has been performed with limited success due to high sequence similarities within the species.

This study describes the use of MALDI-TOF technology to separate S.equi subspecies. Spectra were acquired on a collection of 17 isolates of SEE and SEZ using the VITEK® MS RUO database. Consensus spectra (Superspectra) were then made for each of the two subspecies based on peaks that are conserved for either of the subspecies. The Superspectra were then validated using 42 and 43 isolates of SEZ and SEE respectively. The analysis showed that all the isolates tested could be differentiated to the species level.

In conclusion, MALDI-TOF MS can identify the two important subspecies of Streptococcus equi and may represent a future rapid and cost effective alternative to traditional phenotypic methods.

“The analysis of all streptococci strains examined proved MALDI-TOF MS to be highly efficient in identifying strains belonging to the subspecies

of S.equi spp equi and S.equi spp zooepidemicus”

KEY POINTS

➔ VITEK® MS is a rapid and effective tool for the identification of bacteria in the veterinary diagnostic laboratory.

➔ Subspecies of Streptococcus equi can be differentiated using VITEK® MS.

➔ The RUO database can be exported for potential implementation into other VITEK® MS RUO systems allowing users to share data.

41

JOURNAL OF CLINICAL MICROBIOLOGY2016;54(7):1896-8

Influence of Culture Media on Detection of Carbapenem Hydrolysis by Matrix-Assisted Laser Desorption

Ionization-Time of Flight Mass Spectrometry.Ramos AC1, Carvalhaes CG2, Cordeiro-Moura JR3, Rockstroh AC4, Machado AM5, Gales AC3.

(1) Laboratório Alerta, Division of Infectious Diseases, Department of Internal Medicine, Escola Paulista de Medicina/Universidade Federal de São Paulo, São Paulo, SP, Brazil.

(2) Laboratório Alerta, Division of Infectious Diseases, Department of Internal Medicine, Escola Paulista de Medicina/Universidade Federal de São Paulo, São Paulo, SP, Brazil Laboratório Central, Hospital São Paulo, UNIFESP, São Paulo, SP, Brazil.

(3) Laboratório Alerta, Division of Infectious Diseases, Department of Internal Medicine, Escola Paulista de Medicina/Universidade Federal de São Paulo, São Paulo, SP, Brazil.

(4) bioMérieux, São Paulo, SP, Brazil.(5) Laboratório Central, Hospital São Paulo, UNIFESP, São Paulo, SP, Brazil.

Carbapenemase-resistant bacteria are spreading on a worldwide basis causing public health concerns due to their ability to elude antimicrobial treatment. Early identification of these bacteria is therefore of high importance. Currently, there is a push to develop rapid susceptibility test methods such as a MALDI-TOF MS assay capable of detecting β-lactamase activity in these organisms.

In this study, the influence of different culture media on detection of carbapenemase activity by MALDI-TOF using VITEK® MS was evaluated. A total of 61 carbapenemase producing and 32 non-carbapenemase producing Gram negative bacteria were grown on four different media [Muller Hinton (MHA), Blood agar (BA), Mac Conkey agar (MAC) and chromID CPS agar]. The MALDI-TOF MS assay was performed by incubating organisms from fresh colonies in a buffer solution containing ertapenem for 2 to 4 hours. A mass shift corresponding to carbapenem hydrolysis was then observed on the mass spectrum.

No false positive results were observed for KPC-2 producing Klebsiella pneumoniae grown on MHA, BA and CPS at 2 hours. An extended period of incubation was required for two BKC-1 producing K.pneumoniae and four resistant Acinetobacter baumanii isolates on the same three media. In contrast, carbapenem hydrolysis by 13 carbapenem-resistant isolates grown on MAC medium was not detected by VITEK® MS after 2 hours of incubation. After 4 hours of incubation, the hydrolysis was detected for 9 resistant strains but three remained undetected.

In conclusion, false negative results can be obtained for A.baumanii isolates grown on MAC medium showing that this MALDI-TOF method is sensitive to the growth medium.

“MALDI-TOF hydrolysis has been a useful tool for detecting carbapenemase producers. However, other mechanisms of carbapenem resistance such as

outer membrane protein alteration cannot be assessed by this methodology”

➔ The detection of ß-lactamase production by MALDI-TOF MS is interesting.

➔ The information is useful for telling physicians what they cannot use rather than provide any information about what they can use.

43

KEY POINTS

VITEK® MS RUO (Research Use Only)

42

KEY POINTS

VITEK® MS RUO (Research Use Only)

JOURNAL OF CLINICAL MICROBIOLOGY 2015;53(4):1399-402

Fusobacterium nucleatum subspecies identification by matrix-assisted laser desorption ionization-time of flight

mass spectrometry.Nie Sa,b, Tian Bc, Wang Xc, Pincus DHf, Welker Mg, Gilhuley Kb, Lu Xa, Han YWc,d,e, Tang YWb,h

(a) Department of Laboratory Medicine, Futian Hospital, Guangdong Medical College, Shenzhen, China Department of Laboratory Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.

(a) Department of Periodontics, Case Western Reserve University, Cleveland, Ohio, USA.(c) bioMérieux, Inc., Hazelwood, Missouri, USA.(d) bioMérieux SA, La Balme les Grottes, France.

(e) Department of Laboratory Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.(f) Department of Laboratory Medicine, Futian Hospital, Guangdong Medical College, Shenzhen, China.

(g) Department of Periodontics, Case Western Reserve University, Cleveland, Ohio, USA Department of Pathology, Case Western Reserve University, Cleveland, Ohio, USA Department of Reproductive Biology, Case Western Reserve University, Cleveland, Ohio, USA

(h) Department of Laboratory Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York, USA Department of Pathology and Laboratory Medicine, Weill Medical College of Cornell University, New York, New York, USA

Identification of Fusobacterium nucleatum to the subspecies level is of medical interest due to varying pathogenesis related to different levels of disease. The level of correct identification of thirty-four isolates of F. nucleatum to the subspecies level was studied using the VITEK® MS system and the SARAMIS® database version 4.14.

The commercially available database was modified using 6 American Type Culture Collection (ATCC®) strains and 9 well characterized clinical isolates representing 5 subspecies (animalis, fusiforme, nucleatum, polymorphum, and vincentii) of F. nucleatum. For each strain, 9 spots from 3 different cultures were analyzed and 9 independent spectra were recorded and used to create SuperSpectra for each subspecies.

SARAMIS® generated a confidence-based ranking of agreement to the stored spectra and a confidence level for the identification. A confidence level of ≥80% was considered a reliable identification.

Thirty-four clinical isolates were tested using the modified SARAMIS® database. All isolates were identified to the subspecies level with a confidence value of greater than 80%. For 28 of these 34 isolates, agreement with the 16S rRNA gene sequencing comparison was achieved. Discordant subspecies level identification was given for 4 isolates and 2 isolates were identified to the species level. This lack of subspecies identification may be due to limitations in either or both 16S rRNA sequencing and MALDI-TOF technologies and may be due to the limited number of isolates available for database expansion and validation.

“As accurate identification of F. nucleatum to subspecies level could improve knowledge about the pathogenicity, epidemiology, and clinical relevance, it is

predicted that MALDI-TOF MS can be used in routine practice in clinical laboratories to identify F. nucleatum at subspecies level.”

➔ Correct identification to the subspecies level was obtained for 28 of 34 isolates as compared to 16S rRNA.

➔ For the remaining 6 isolates, correct identification was achieved at the species level and discordant subspecies identification results were obtained for only 4 isolates.

JOURNAL OF CLINICAL MICROBIOLOGY2015;53(7):2355-8

Mycobacterium abscessus Complex Identification with Matrix-Assisted Laser Desorption Ionization-Time of

Flight Mass Spectrometry.Panagea Ta,b, Pincus DHc, Grogono Dd,e, Jones Mf, Bryant Jg, Parkhill Jg, Floto RHd,h, Gilligan Pb,f.

(a) Sismanogleio-A.Flemig General Hospital, Athens, Greece(b) University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA

(c) bioMérieux, Inc., Hazelwood, Missouri, USA(d) Cambridge Centre for Lung Infection, Papworth Hospital, Cambridge, United Kingdom

(e) University of Cambridge, Cambridge, United Kingdom(f) UNC Health Care, Chapel Hill, North Carolina, USA

(g) Wellcome Trust Sanger Institute, Hinxton, United Kingdom(h) Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom

Non-tuberculous mycobacteria are becoming more prevalent in industrialized countries as causes of respiratory disease in cystic fibrosis patients as well as pulmonary, skin and soft tissue and post-surgical infections in other patients. Although still being debated, recent studies with whole genome sequencing (WGS) suggest there are 3 subspecies of M. abscessus – M. abscessus subsp. abscessus (senso stricto), M. abscessus subsp. massiliense and M. abscessus subsp. bollettii. It is important to discriminate between these subspecies due to differences in resistance and treatment response rates.

This study compared the performance of the VITEK® MS PLUS System RUO database version 4.12 and IVD database version 3.0 for the identification Mycobacterium abscessus using a collection of strains previously characterized using WGS. Isolates were grown on both Middlebrook7H11 media and Burkholderia cepacia selective agar (BCSA).

Two MALDI-TOF databases were evaluated for the identification of 54 M. abscessus strains, 41 M. abscessus subsp. abscessus and 13 M. abscessus subsp. massiliense. An extraction protocol was performed and the isolates were tested using the VITEK® MS PLUS system. Each isolate was tested twice to avoid acquisition errors.

All 54 isolates were correctly identified as M. abscessus by the VITEK® MS PLUS system with both databases and both media types except for one isolate grown on Middlebrook7H11 media and one on BCSA in the RUO SARAMIS® database. M. abscessus was however given as the closest match.

At the time of this study, the databases tested were only able to identify the isolates to the species level. The ability to identify these organisms to the subspecies level was therefore not expected. As the taxonomy of this organism is further clarified, updates to the MALDI-TOF databases will be required. This study demonstrates the potential of this technology.

“In conclusion, MALDI-TOF is a powerful technique capable of accurate and rapid identification of M. abscessus isolates not only from

specific mycobacterial culture media but also from selective media used to detect Burkholderia cepacia complex isolates in routine

bacterial cultures of CF patients.”

➔ All 54 isolates of M. abscessus were correctly identified to the species level except for 2 isolates.

➔ Peak analysis revealed three peaks potentially able to differentiate between subspecies.

EVOLUTION OF THE VITEK® MS

Axima@SARAMIS® (Research Use Only)

VITEK® MS RUO (Research Use Only)SARAMIS® database

VITEK® MS IVD V1 Pre-commercialized databasebioMérieux developed database

VITEK® MS IVD V1.0CE-Marked

VITEK® MS IVD V2.0FDA cleared and CE-Marked

VITEK® MS Plus (IVD Plus RUO)

VITEK® MS IVD V3.0FDA cleared and CE-Marked

NOTES.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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bioMérieux S.A. • 69280 Marcy l’Étoile • France • Tel.: + 33 (0)4 78 87 20 00 • Fax: +33 (0)4 78 87 20 90www.biomerieux.com • www.biomerieux-diagnostics.com/vitekr-ms-0

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2

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