MSACL 2019 EU Abstract

MBT FAST – Rapid Antimicrobial Susceptibility Testing by MALDI-TOF MS

Markus Kostrzewa1, Ilka D. Nix2, Evgeny A. Idelevich2, Katrin Sparbier1, Oliver Drews1, Karsten Becker2
1Bruker Daltonik GmbH, Bremen, Germany; 2University Hospital Münster, Institute of Medical Microbiology, Münster, Germany

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Markus Kostrzewa (Presenter) Bruker Daltonik GmbH

Presenter Bio: Study of biology at the Justus-Liebig-University, Giessen, Germany.

Diploma (1990) and Ph.D. thesis(1993) about molecular evolution of plastids.


1993 - 1997

Postdoc at the Institute of Human Genetics, JLU Giessen (AG Prof. U. Müller).

• Diagnostic of hereditary human diseases
• Research in the framework of the Human Genome Project

Joined Bruker in 1998

Head of the new R&D group “Bioanalytical Development“, establishment of molecular biology and biology related development at Bruker Daltonik.

Development of DNA analysis by MALDI-TOF mass spectrometry, “Clincal Proteomics” - methods, consumables and software for mass spectrometry profiling of body fluids and tissues

Development of microorganism identification by MALDI-TOF mass spectrometry (“MALDI Biotyper” system).

2005 Director Molecular Biology, R&D

2012 Vice President Clinical Mass Spectrometry R&D

2017 Vice President Micr

Relevant Financial Disclosures (within past 24 months) Stock/Bonds Bruker Daltonik GmbH Salary Bruker Daltonik GmbH Discussing Products? YES, presenter will present educational content related to products/services of one or more ineligible company(ies) listed above. Conflict mitigation IN PROCESS.

Introduction: Over the recent years, (multi-) drug resistant clinical isolates escalated resulting in a global threat that requires the fast and reliable detection of the corresponding resistance pattern. This is a major pre-requisite for an adequate and dedicated patient management and the prevention for further spread of those strains. We have developed a novel approach allowing MALDI-TOF mass spectrometry -based antibiotic susceptibility testing (AST), MBT FAST, within only a few hours.

METHODS: Twelve Enterobacteriaceae isolates comprising Escherichia coli, Klebsiella pneumoniae, Enterobacter spp. and Citrobacter spp. (each n = 3) and three Hafniaceae (Hafnia alvei) consecutive clinical isolates were investigated. A therapeutically relevant panel of seven antibiotics with anti-Enterobacterales activity including piperacillin, piperacillin/tazobactam, cefotaxime, ceftazidime, ciprofloxacin, gentamicin and meropenem was tested at EUCAST breakpoint concentrations to allow categorisation as either susceptible or resistant. 6 µl of bacterial suspensions (5x105 cfu/ml) with and without antibiotics in cation-adjusted Mueller-Hinton broth were placed onto a Biotarget 96 (Bruker Daltonik), each sample in duplicate. The inoculated targets were incubated in a humidity chamber at 36°C for 4.5 hours. Subsequently, medium was removed using absorptive pads (Bruker Daltonik) and matrix was spotted onto the dried spots. Measurements were performed in duplicate, acquired spectra were analysed by a newly developed prototype software (Bruker Daltonik). Each isolate was tested independently three times and median results were calculated. Broth microdilution was used as reference method.

RESULTS: Comparison of the results obtained by the MBT FAST assay with those obtained by standard broth microdilution approach revealed a categorical agreement of 93.0% for all seven antibiotics. In contrast to 20 h incubation time necessary to perform the microdilution, the MALDI-TOF MS based assay was ready for readout already after 4.5 h incubation. Overall, 2.0% of the tested isolates showed false-resistant results and 5.0% showed false-susceptible results. The validity of the approach, i.e. successful detection of growth control, was 96%.

CONCLUSION: This study indicated that simultaneous susceptibility testing of antibiotics by MALDI-TOF MS-based MBT FAST is feasible and accurate within 4.5 hours. The whole workflow consists of pipetting steps and a simple liquid removal and can be performed without any centrifugation, therefore an automation of the procedure will be possible. The read-out can of results can be performed with standard routine MALDI-TOF mass spectrometers as they are available in most clinical microbiology laboratories today. Therefore, this method will be an attractive for rapid routine AST alternative for the future.