Optimization of the MALDIxin test for the rapid identification of colistin resistance in Klebsiella pneumoniae using MALDI-TOF MS.

Dortet L1,2,3,4, Broda A1, Bernabeu S1,2,3,4, Glupczynski Y5, Bogaerts P5, Bonnin R3,4, Naas T2,3,4, Filloux A1, Larrouy-Maumus G1.

Author information

1: MRC Centre for Molecular Bacteriology and Infection, Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London, UK.
2: Department of Bacteriology-Hygiene, Bicêtre Hospital, Assistance Publique - Hôpitaux de Paris, Le Kremlin-Bicêtre, France.
3: EA7361 'Structure, Dynamic, Function and Expression of Broad Spectrum β-Lactamases', Paris-Sud University, LabEx Lermit, Faculty of Medicine, Le Kremlin-Bicêtre, France.
4: French National Reference Centre for Antibiotic Resistance, Le Kremlin-Bicêtre, France.
5: Laboratory of Clinical Microbiology, Belgian National Reference Centre for Monitoring Antimicrobial Resistance in Gram-Negative Bacteria, CHU UCL Namur, Yvoir, Belgium.

Abstract

BACKGROUND:

With the dissemination of carbapenemase producers, a revival of colistin was observed for the treatment of infections caused by MDR Gram-negatives. Unfortunately, the increasing usage of colistin led to the emergence of resistance. In Klebsiella pneumoniae, colistin resistance arises through addition of 4-amino-l-arabinose (l-Ara4N) or phosphoethanolamine (pEtN) to the native lipid A. The underlying mechanisms involve numerous chromosome-encoded genes or the plasmid-encoded pEtN transferase MCR. Currently, detection of colistin resistance is time-consuming since it still relies on MIC determination by broth microdilution. Recently, a rapid diagnostic test based on MALDI-TOF MS detection of modified lipid A was developed (the MALDIxin test) and tested on Escherichia coli and Acinetobacter baumannii.

OBJECTIVES:

Optimize the MALDIxin test for the rapid detection of colistin resistance in K. pneumoniae.

METHODS:

This optimization consists of an additional mild-acid hydrolysis of 15 min in 1% acetic acid. The optimized method was tested on a collection of 81 clinical K. pneumoniae isolates, including 49 colistin-resistant isolates (45 with chromosome-encoded resistance, 3 with MCR-related resistance and 1 with both mechanisms).

RESULTS:

The optimized method allowed the rapid (<30 min) identification of l-Ara4N- and pEtN-modified lipid A of K. pneumoniae, which are known to be the real triggers of polymyxin resistance. At the same time, it discriminates between chromosome-encoded and MCR-related polymyxin resistance.

CONCLUSIONS:

The MALDIxin test has the potential to become an accurate tool for the rapid determination of colistin resistance in clinically relevant Gram-negative bacteria.