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New method for fast and direct Mycobacteria identification from positive blood culture

A Camaggi(1) S Tonello(2) R Minisini(2) M Mantovani(1) M S Caroppo(1) S Andreoni(1)

1:AOU Maggiore della Carità, Laboratory of Microbiology and Virology - Novara, Italy; 2:Università del Piemonte Orientale, DIMET – Novara, Italy

Disseminated nontuberculous mycobacterial (NTM) infection is an infectious disease that occurs mostly in immunocompromised hosts. The use of a fully automated method, as BACTEC-MYCO/F-LYTIC (Becton Dickinson Sparks, MD, USA) bottles system, represents a sensitive method for mycobacteremia detection. Unfortunately, Line Probe Assay or other conventional commercial assays for mycobacteria identification could not be used for samples containing blood (subculture on liquid or solid media are required before). In this study we evaluate the MBT-Sepsityper Kit (Bruker Daltonik, GmbH, Germany) for the direct mycobacteria identification from positive blood cultures bottles (BCBs) using MALDI-TOF MS.

Seventy MYCO/F-LYTIC-bottles were inoculated with human blood (4.9 mL) and various NTM species suspension (0.1 mL at 0.5 McF): M. abscessus, M. avium, M. chelonae, M. chimaera, M. elephantis, M. fortuitum, M. hassiacum, M. intracellulare, M. llatzerense, M. lentiflavum, M. mucogenicum, M. nebraskense, M. parascrofulaceum, M. simiae. Once flagged positive and AFB documented, 5.0 mL of blood culture were centrifugated and 1.0 mL of pellet underwent Sepsityper preparation, following the manufacturer’s instructions (classical protocol) and our homemade inactivation/extraction protein steps (ethanol/formic acid-acetonitrile) for MALDI mycobacteria identification. All the samples were correctly identified at the species level except M. chimaera and M. intracellulare. Their mass spectra are more similar and by standard MALDI-TOF library they couldn’t be differentiated.

These data show how the employment of the Sepsityper-kit for the mycobacteria identification with MALDI-TOF in BCBs allows a rapid (~2 hours) and reliable identification leading to a significant decrease in timing to results (5-15 days depending on mycobacteria species).

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