Performance of MALDI-TOF Mass Spectrometry for the Identification of the HACEK Group and Other Fastidious Gram-Negative Rods

Marisa Almuzara,Carlos Vay,Claudia Barberis,Karen C V Cárdenas,Maria S Ramirez,Angela Famiglietti

doi:10.2174/1874285801913010125

Abstract

Objective: The aim of this study was to determine the capacity of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) to identify 155 HACEK clinical isolates and other fastidious or infrequently isolated Gram-negative rods (e.g., Actinobacillus, Capnocytophaga, Pasteurella, Neisseria, Moraxella, Dysgonomonas, among others). Methods: All the isolates were identified by standard biochemical tests and MALDI-TOF MS. Two different extraction methods (direct transfer formic acid method on spot and ethanol formic acid extraction method) and different cut-offs for genus/specie level identification were used. MALDI-TOF MS identification was considered correct when the result obtained from the MS database agreed with the phenotypic identification result. When both the methods gave discordant results, the 16S rDNA gene sequencing was considered as the gold standard identification method. Results: Employing the score cut-offs suggested by the manufacturer, 93.55% and 69.03% isolates were correctly identified at the genus and species level, respectively. On the contrary , employing lower cut-off scores for identification, 98.06% and 92.09% isolates were properly identified at the genus and species level respectively and no significant differences between the results obtained with two extraction methods were observed . Conclusion: The accurate identification of 14 genera showed the reliability of MALDI-TOF MS as an optional methodology to the routine identification methods currently used in laboratories.

Highlights

The identification of the HACEK group (Haemophilus, Aggregatibacter, Cardiobacterium, Eikenella, Kingella) and other fastidious Gram-negative rods (e.g., Actinobacillus, Capnocytophaga, Pasteurella, Neisseria, Moraxella, Dysgonomonas, among others) by conventional phenotypic methodology is difficult, mainly because of their slow growth and low reactivity in biochemical tests
It is well known that conventional identification methods for the HACEK group and other fastidious Gram-negative rods are time-consuming, which contribute to the problems in their routine identification in the microbiology laboratory
All of the isolates were previously identified by conventional biochemical tests [7, 8]. 16S rRNA gene sequencing was applied to solve any discrepancies between the identification obtained via conventional methodology and the identification obtained by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF Mass Spectrometry (MS)). 16S rRNA gene sequencing was applied for definitive species identification in cases in which phenotypic tests were not reliable

Summary

Introduction

The identification of the HACEK group (Haemophilus, Aggregatibacter, Cardiobacterium, Eikenella, Kingella) and other fastidious Gram-negative rods (e.g., Actinobacillus, Capnocytophaga, Pasteurella, Neisseria, Moraxella, Dysgonomonas, among others) by conventional phenotypic methodology is difficult, mainly because of their slow growth and low reactivity in biochemical tests. Identification using molecular methods requires trained staff Because of these factors, we thought it would be valuable to assess the capacity of matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) Mass Spectrometry (MS) for the identification of HACEK group members and other fastidious or infrequently isolated Gram-negative rods In the past few years, this technology has been demonstrated to be very useful in clinical microbiology because it allows for the definitive identification of different bacterial species within a few minutes [1 - 6]

Methods

Results

Conclusion