Multicenter Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Study for Identification of Clinically Relevant Nocardia spp

Abstract

This multicenter study analyzed Nocardia spp., including extraction, spectral acquisition, Bruker matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) identification, and score interpretation, using three Nocardia libraries, the Bruker, National Institutes of Health (NIH), and The Ohio State University (OSU) libraries, and compared the results obtained by each center. A standardized study protocol, 150 Nocardia isolates, and NIH and OSU Nocardia MALDI-TOF MS libraries were distributed to three centers. Following standardized culture, extraction, and MALDI-TOF MS analysis, isolates were identified using score cutoffs of ≥2.0 for species/species complex-level identification and ≥1.8 for genus-level identification. Isolates yielding a score of <2.0 underwent a single repeat extraction and analysis. The overall score range for all centers was 1.3 to 2.7 (average, 2.2 ± 0.3), with common species generally producing higher average scores than less common ones. Score categorization and isolate identification demonstrated 86% agreement between centers; 118 of 150 isolates were correctly identified to the species/species complex level by all centers. Nine strains (6.0%) were not identified by any center, and six (4.0%) of these were uncommon species with limited library representation. A categorical score discrepancy among centers occurred for 21 isolates (14.0%). There was an overall benefit of 21.2% from repeat extraction of low-scoring isolates and a center-dependent benefit for duplicate spotting (range, 2 to 8.7%). Finally, supplementation of the Bruker Nocardia MALDI-TOF MS library with both the OSU and NIH libraries increased the genus-level and species-level identification by 18.2% and 36.9%, respectively. Overall, this study demonstrates the ability of diverse clinical microbiology laboratories to utilize MALDI-TOF MS for the rapid identification of clinically relevant Nocardia spp. and to implement MALDI-TOF MS libraries developed by single laboratories across institutions.

FIG 1

Average MALDI-TOF MS score per species- or species complex-level identification. The average MALDI-TOF MS score per isolate is represented for those species or species complexes with greater than three isolates represented within the study. Horizontal bars within the box-and-whisker plots represent the average for all species or species complex (cpx) isolates in that category.

FIG 2

Discrepant score agreement among centers. (A) Discrepancies in original MALDI-TOF MS scores submitted for each center for selected isolates; (B) discrepancies in MALDI-TOF MS scores from submitted protein extracts analyzed side by side at a single center. Light gray circles, center 1; black circles, center 2; dark gray circles, center 3.

FIG 3

Representative Nocardia mexicana spectral overlay. Library (black) and isolate (red) spectra display peak mismatches.