MALDI-TOF Mass Spectrometry as a Rapid Screening Alternative for Non-tuberculous Mycobacterial Species Identification in the Veterinary Laboratory.

Víctor Lorente-Leal,Javier Bezos,Emmanouil Liandris,Marta Pérez-Sancho,Lucía De Juan,Beatriz Romero

doi:10.3389/fvets.2022.827702

Víctor Lorente-Leal, Javier Bezos + Show 4 more

Open Access

https://doi.org/10.3389/fvets.2022.827702

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Abstract

Non-tuberculous mycobacteria (NTM) are difficult to identify by biochemical and genetic methods due to their microbiological properties and complex taxonomy. The development of more efficient and rapid methods for species identification in the veterinary microbiological laboratory is, therefore, of great importance. Although MALDI-TOF Mass Spectrometry (MS) has become a promising tool for the identification of NTM species in human clinical practise, information regarding its performance on veterinary isolates is scarce. This study assesses the capacity of MALDI-TOF MS to identify NTM isolates (n = 75) obtained from different animal species. MALDI-TOF MS identified 76.0% (n = 57) and 4% (n = 3) of the isolates with high and low confidence, respectively, in agreement with the identification achieved by Sanger sequencing of housekeeping genes (16S rRNA, hsp65, and rpoB). Thirteen isolates (17.3%) were identified by Sanger sequencing to the complex level, indicating that these may belong to uncharacterised species. MALDI-TOF MS approximated low confidence identifications toward closely related mycobacterial groups, such as the M. avium or M. terrae complexes. Two isolates were misidentified due to a high similarity between species or due to the lack of spectra in the database. Our results suggest that MALDI-TOF MS can be used as an effective alternative for rapid screening of mycobacterial isolates in the veterinary laboratory and potentially for the detection of new NTM species. In turn, Sanger sequencing could be implemented as an additional method to improve identifications in species for which MALDI-TOF MS identification is limited or for further characterisation of NTM species.

Highlights

Mycobacteria are a diverse group of acid-fast Gram-positive bacilli that include more than 200 species differentiated into five newly emended genera [1]
Sequence identity was below 98% for the complementary sequencing targets for 12 isolates, but the 16S rRNA sequence was related to the M. avium (n = 8), M. terrae (n = 2), M. fortuitum (n = 1), or M. simiae (n = 1) complexes (Supplementary Table 1)
The implementation of MALDI-TOF Mass Spectrometry (MS) in the identification of mycobacteria has become an interesting alternative to genetic methods in human clinical microbiology laboratories

Summary

Introduction

Mycobacteria are a diverse group of acid-fast Gram-positive bacilli that include more than 200 species differentiated into five newly emended genera [1]. Molecular genetic methods greatly improved the capacity of clinical microbiology laboratories to identify NTM species with the development of several methods, such as genetic probe assays, including the INNO-LiPA Mycobacteria or the GenoType Common Mycobacteria assays [7,8,9]. These methods are still used in many laboratories, they are expensive, identify a limited number of mycobacterial species and misidentify less prevalent NTM species due to probe cross-reactivity [10]. Many laboratories still use a combination of probes and Sanger sequencing as a reference method to identify NTM species [8, 15, 16]

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