Fig 1.
(Left column) Structures of the native (top) and modified (middle and bottom) lipid A portion of the Escherichia coli LPS. L-Ara4N (marked in red) and PEtN (marked in blue) modifications to lipid A cause resistance to polymyxin antibiotics. L-Ara4N modification of the 4ˊ-phosphate of native lipid A is characteristic of colistin-resistant E. coli strains carrying chromosomal mutations; this modification often co-occurs with PEtN modification of the 1-phosphate of the lipid A structure. Lipid A from strains expressing MCR enzymes is only modified by PEtN. (Right column) Lipid A spectra obtained from intact E. coli colonies using the MALDI Biotyper Sirius system (Bruker Daltonics) and the adapted MALDIxin protocol [18]. Spectra obtained from colistin-susceptible strains have one major peak at m/z 1,796.2 (top), corresponding to native lipid A. Depending on the mechanism of colistin resistance, spectra from colistin-resistant strains have additional peaks at m/z 1,927.2 (middle, marked in red), due to L-Ara4N modification of lipid A, and/or at m/z 1,919.2 (bottom, marked in blue), due to PEtN modification of lipid A. LPS, lipopolysaccharide; L-Ara4N, 4-amino-4-deoxy-L-arabinose; PEtN, phosphoethanolamine; MCR, mobile colistin resistance; m/z, mass-to-charge ratio.
Table 1.
m/z values rounded to the nearest decimal for native and modified lipid A peaks obtained by MALDI-TOF mass spectrometry from intact colonies of gram-negative bacterial pathogens that are commonly resistant to colistin.
Fig 2.
Schematic diagram of the sample preparation process for the adapted MALDIxin test performed on the MALDI Biotyper Sirius system (Bruker Daltonics).
The total sample processing time using this approach is less than 15 minutes, allowing for immediate assessment of whether bacteria are resistant to polymyxin antibiotics, independent of the mechanism of resistance. In the published feasibility study [18], bacterial colonies were grown overnight on solid medium; in future clinical applications, bacteria could be obtained directly from biological samples, such as urine or blood. Figure created with BioRender.