Direct detection of lipid A on intact Gram-negative bacteria by MALDI-TOF mass spectrometry

Larrouy-Maumus, Gérald; Clements, Abigail; Filloux, Alain; McCarthy, Ronan R.; Mostowy, Serge

doi:10.1016/j.mimet.2015.12.004

Cited by 51 publications

(47 citation statements)

References 17 publications

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“…After purification through Bligh–Dyer extraction, a detailed MALDI MS and MS 2 investigation has been performed on the isolated lipid A fraction. In parallel, a MALDI MS investigation was also performed directly on the bacterial pellet, following the approach reported by Larrouy‐Maumus et al., without any chemical treatment that could potentially cause the loss of modifications of the lipid A domain.…”

Section: Resultsmentioning

confidence: 99%

“…MALDI MS and MS 2 analysis : The MS investigation was performed on an ABSCIEX TOF/TOFTM 5800 Applied Biosystems mass spectrometer, equipped with an Nd:YLF laser ( λ =345 nm), with a pulse length of <500 ps and a repetition rate of up to 1000 Hz. The bacterial pellet for MALDI preparation was treated as previously described . The matrix solution was prepared by dissolving 2,5‐dihydroxybenzoic acid (DHB) to a final concentration of 10 mg mL −1 in chloroform/methanol (9:1, v / v ).…”

Section: Methodsmentioning

confidence: 99%

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The Lipid A Structure from the Marine Sponge Symbiont Endozoicomonas sp. HEX 311

et al. 2018

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Endozoicomonas sp. HEX311 is a Gram-negative bacterium known to establish a commensal interaction with the marine demosponge Suberites domuncula. The molecular bases of the sponge-microbe interaction events are still poorly defined. Nevertheless, it has been proved that S. domuncula possesses an innate immune system with similarities to the mammalian one and is able to recognize the main component of the Gram-negative bacteria cell wall: the lipopolysaccharide. Whether this recognition occurs in a structure-dependent manner, which is typical for mammalian immune system receptors, is still under investigation. Herein, we report the Endozoicomonas sp. HEX311 lipid A structure obtained by a combination of data attained from chemical, MALDI MS, and MS approaches. The lipid A is a complex family of species decorated by pyrophosphate and phosphate units and carrying (R)-3-hydroxydodecanoic acid, (R)-3-hydroxytetradecanonic acid, iso-2-hydroxyundecanoic acid, iso-(R)-3-hydroxyundecanoic acid, and iso-nonanoic acid as acyl chains.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

The Lipid A Structure from the Marine Sponge Symbiont Endozoicomonas sp. HEX 311

et al. 2018

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“…As the structure of Lipid A from a range of bacterial species (including Klebsiella pneumonia, Shigella spp. and Pseudomonas aeruginosa ) can be determined by MALDI-TOF mass spectrometry (18), this technique provides a broadly applicable basis for the development of new diagnostics in many species of Gram-negative bacteria. Indeed, the Lipid A of Salmonella spp., which have been reported to carry MCR-enzymes (19) is similar to that of E. coli and can be detected using the negative-ion mode of the MALDI Biotyper Sirius as a peak at m/z 1796.2 (data not shown).…”

Section: Discussionmentioning

confidence: 99%

Fast and robust detection of colistin resistance in Escherichia coli using the MALDI Biotyper Sirius mass spectrometry system

Furniss

Dortet

Bolland

et al. 2019

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11 3 EA7361 "Structure, dynamic, function and expression of broad spectrum β-lactamases", ABSTRACT 31 Polymyxin antibiotics are a last-line treatment for multidrug-resistant Gram-negative 32 bacteria. However, the emergence of colistin resistance, including the spread of mobile mcr 33 genes, necessitates the development of improved diagnostics for the detection of colistin-34 resistant organisms in hospital settings. The recently developed MALDIxin test enables 35 detection of colistin resistance by MALDI-TOF mass spectrometry in less than 15 minutes 36 but is not optimized for the mass spectrometers commonly found in clinical microbiology 37 laboratories. In this study, we adapted the MALDIxin test for the MALDI Biotyper Sirius 38 MALDI-TOF mass spectrometry system (Bruker Daltonics). We optimized the sample 39 preparation protocol using a set of 6 MCR-expressing Escherichia coli clones and validated the 40 assay with a collection of 40 E. coli clinical isolates, including 19 MCR producers, 12 41 chromosomally-resistant isolates and 9 polymyxin-susceptible isolates. We calculated 42 Polymyxin resistance ratio (PRR) values from the acquired spectra; a PRR value of zero, 43 indicating polymyxin susceptibility, was obtained for all colistin-susceptible E. coli isolates, 44 whereas positive PRR values, indicating resistance to polymyxins, were obtained for all 45 resistant strains independent of the genetic basis of resistance. Thus, we report a preliminary 46 feasibility study showing that an optimized version of the MALDIxin test, adapted for the 47 routine MALDI Biotyper Sirius, provides an unbiased, fast, reliable, cost-effective and high-48 throughput way of detecting colistin resistance in clinical E. coli isolates.49

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“…The demonstration that lipid A from several gram-negative bacterial species can be detected and its structure characterized by MALDI-TOF mass spectrometry using intact bacterial colonies [19] laid the groundwork for many of the subsequent studies tackling the development of MALDI-TOF-based diagnostics for the rapid detection of colistin resistance. Notably, unlike bacterial identification or detection of β-lactamase activity, which require sample analysis using the positive-ion mode, this approach relies on operating the mass spectrometer in the negative-ion mode, which was not possible on the instruments in routine clinical use until recently [18].…”

Section: Maldi-tof Mass Spectrometry Can Be Used To Characterize the mentioning

confidence: 99%

“…As modification of lipid A is the most common mechanism of colistin resistance in gram-negative bacteria [17], and the lipid A structure of several species can be characterized by MAL-DI-TOF mass spectrometry [19], the MALDIxin approach can be applied to other gramnegative pathogens in which colistin resistance is a great challenge. Table 1 summarizes the expected peaks corresponding to native and modified lipid A for organisms in which colistin resistance is prevalent and this approach has been tested.…”

Section: Maldi-tof Mass Spectrometry Detects Resistance In Other Grammentioning

confidence: 99%

The clue is in the lipid A: Rapid detection of colistin resistance

et al. 2020

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Matrix-assisted laser desorption/ionization-time of flight mass spectrometry is an emerging technology for the detection of antimicrobial resistance Antimicrobial resistance (AMR) is one of the biggest public health concerns of our time [1]. Currently, resistance has been reported for almost all available antibiotics, from first-line to last-resort drugs [2]. AMR restricts treatment options, leads to increased morbidity and mortality of hospitalized patients and imposes a substantial financial burden on healthcare systems worldwide [3]. The limited pipeline of new antimicrobials and the continuous emergence of multidrug-resistant (MDR) organisms needs to be countered by novel antibacterial strategies coupled with rapid diagnostics to detect resistance, particularly in the case of broad-acting agents such as β-lactams and polymyxins. Traditionally, antimicrobial susceptibility testing is performed using culture-based methods, like minimum inhibitory concentration (MIC) determination assays, a lengthy process that is prone to misleading results, for example, in the case of clinical isolates that produce large amounts of capsule or biofilm. By contrast, polymerase-chain-reaction (PCR)-based tests like multiplex PCR allow the rapid identification of specific resistance factors but cannot predict susceptibility and are entirely insensitive to uncharacterized mechanisms of resistance. At the same time, despite the extensive progress of genotypic sequence-based approaches, such as long-read nanopore sequencing, these cannot yet be used as first-line diagnostics in part because genotype-to-phenotype predictions remain challenging [4,5]. With matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry being used routinely for bacterial identification in clinical settings, a range of MAL-DI-TOF-based diagnostics for the rapid detection of AMR are starting to be implemented [6,7]. More specifically, it is possible to discriminate some lineages of methicillin-resistant Staphylococcus aureus, identify certain strains of enterobacteria carrying plasmid-encoded βlactamases from the Klebsiella pneumoniae carbapenemase (KPC) family of enzymes, and detect Bacteroides fragilis strains that are resistant to β-lactams at the same time as performing bacterial identification. Alongside these identification-based approaches, efforts to identify both aminoglycoside and quinolone resistance using MALDI-TOF mass spectrometry have also been made. As resistance to these agents can arise through enzymatic modification of the antibiotic, this modification can be directly detected following incubation of the drug of interest with resistant bacteria [8,9]. Nonetheless, probably the most successful application of MAL-DI-TOF mass spectrometry for AMR detection to date has been functional screening for βlactamase activity. Bacteria are mixed with a β-lactam compound, and an increase in the

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Direct detection of lipid A on intact Gram-negative bacteria by MALDI-TOF mass spectrometry

Cited by 51 publications

References 17 publications

The Lipid A Structure from the Marine Sponge Symbiont Endozoicomonas sp. HEX 311

The Lipid A Structure from the Marine Sponge Symbiont Endozoicomonas sp. HEX 311

Fast and robust detection of colistin resistance in Escherichia coli using the MALDI Biotyper Sirius mass spectrometry system

The clue is in the lipid A: Rapid detection of colistin resistance

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