Rapid identification and typing of listeria species by matrix-assisted laser desorption ionization-time of flight mass spectrometry

Abstract

Listeria monocytogenes is a food-borne pathogen that is the causative agent of human listeriosis, an opportunistic infection that primarily infects pregnant women and immunologically compromised individuals. Rapid, accurate discrimination between Listeria strains is essential for appropriate therapeutic management and timely intervention for infection control. A rapid method involving matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) that shows promise for identification of Listeria species and typing and even allows for differentiation at the level of clonal lineages among pathogenic strains of L. monocytogenes is presented. A total of 146 strains of different Listeria species and serotypes as well as clinical isolates were analyzed. The method was compared with the pulsed-field gel electrophoresis analysis of 48 Listeria strains comprising L. monocytogenes strains isolated from food-borne epidemics and sporadic cases, isolates representing different serotypes, and a number of Listeria strains whose genomes have been completely sequenced. Following a short inactivation/extraction procedure, cell material from a bacterial colony was deposited on a sample target, dried, overlaid with a matrix necessary for the MALDI process, and analyzed by MALDI-TOF MS. This technique examines the chemistry of major proteins, yielding profile spectra consisting of a series of peaks, a characteristic "fingerprint" mainly derived from ribosomal proteins. Specimens can be prepared in a few minutes from plate or liquid cultures, and a spectrum can be obtained within 1 minute. Mass spectra derived from Listeria isolates showed characteristic peaks, conserved at both the species and lineage levels. MALDI-TOF MS fingerprinting may have potential for Listeria identification and subtyping and may improve infection control measures.

FIG. 1.

MALDI-TOF MS spectra of whole-cell extracts of Listeria reference strains. Spectra representative of the MS profile were obtained from the German Collection of Microorganisms and Cell Cultures. The absolute intensities of the ions are shown on the y axis, and the masses (in Da) of the ions are shown on the x axis. The m/z value stands for mass to charge ratio. For a single positive charge, this value corresponds to the molecular weight of the protein.

FIG. 2.

Discriminating peaks of Listeria strains analyzed by MALDI-TOF MS. The absolute intensities of the ions are shown on the y axis, and the masses (in Da) of the ions are shown on the x axis. The m/z value stands for mass to charge ratio. For a single positive charge, this value corresponds to the molecular weight of the protein.

FIG. 3.

Dendrogram derived from a PFGE profile of ApaI macrorestriction showing restriction patterns among the L. monocytogenes serotypes. Isolate identification numbers and serotype are indicated to the right.