MSACL 2019 EU Abstract

Detection of Vancomycin-Resistant Enterococcus faecium Using MALDI-TOF Mass Spectrometry

Manuel Arroyo (1), Adrián Ruiz (2), Luis Mancera (1), Lidia Quiroga (2), Emilia Cercenado (2), Belén Rodríguez-Sánchez (2)
(1) Clover Biosoft, Granada, Spain (2) Clinical Microbiology and Infectious Diseases Department, Hospital General Universitario Gregorio Marañón. Instituto de Investigación Sanitaria Gregorio Marañón, Madrid, Spain

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Belén Rodríguez-Sanchez (Presenter) Hospital General Universitario Gregorio Marañón

Presenter Bio: I am a molecular biologist leading a MS research group in the Hospital Gregorio Marañon in Madrid, Spain. During the last 8 years we have implemented MALDI-TOF in our hospital for the identification of bacteria, yeasts and filamentous fungi. We have also applyied MALDI-TOF for the detection of antibiotic-resistance mechanisms. We are now interested in bacterial typing using MALDI-TOF.

Relevant Financial Disclosures (within past 24 months) No relevant financial relationship(s) to disclose. Discussing Products? NO, presenter will NOT present educational content related to products/services of the ineligible company(ies) listed above. Conflict mitigation NOT REQUIRED.

Introduction: Differentiation between Vancomycin-Resistant and Vancomycin-Sensitive Enterococci (VRE and VSE) is of paramount important in the clinical microbiology laboratory for the correct management of infected patients.
Objectives: The goal of this study was to discriminate between Vancomycin-Resistant and Vancomycin-Sensitive Enterococcus faecium using MALDI-TOF Mass Spectrometry (MS).
Methods: Overall, 60 E. faecium isolates (20 isolates sensitive to vancomycin, 20 hosting the vanA and 20 the vanB gene clusters) were cultured overnight on Columbia agar + 5% sheep blood (bioMérieux, Marcy L'étoile, France). Small amounts of biomass from individual colonies were spotted on four spots of the MALDI plate and submitted to on-plate protein extraction with 100% formic acid. Two protein spectra from each spot was achieved and labeled as “Day 1”. Each isolate was subcultured again in the above-mentioned conditions and labeled as “Day 2” and “Day 3”. For these time points, colonies were again spotted x4 and 4 protein spectra were collected from each of them in the range of 2.000 to 20.0000 Da using Bruker Biotyper (Bruker, Bremen, Germany). Data was submitted to analysis using Clover Bacterial Analysis software (Clover Biosoft, Granada, Spain). All spectra were preprocessed by a pipeline of a) baseline subtraction, b) picking the 50 most representative peaks, c) peak alignment and d) TIC normalization. Finally, 3D PLS-DA analysis was applied to discriminate the different groups of isolates.
Results: The PLS discriminant analysis used was able to establish a clear separation between the three groups of Enterococci (Vancomycin-Sensitive, vanA- and vanB-gene cluster hosting isolates) providing a list of most relevant peaks, key for their discrimination. The 3D graph shows three well-discriminated groups, corresponding to each group of isolates. Furthermore, the PLS-DA analysis provides a metric to examine the relevance of each m/z value to differentiate the groups.
Conclusions: Our study shows that MALDI-TOF MS is a rapid, reliable and inexpensive tool for the detection of vancomycin-resistant E. faecium. Further studies are required in order to corroborate these preliminary results.