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Combined use of commercial and sequencing analysis methods for the identification of non-tuberculous mycobacteria in a tertiary Hospital of Athens: Seventeen years of experience 

F Kontos(1) G Mavromanolakis(2) S Pournaras(1)

1:Laboratory of Clinical Microbiology, Attikon University Hospital, Medical School, National and Kapodistrian University of Athens, Athens, Greece; 2:Department of Internal Medicine. General Hospital of Agios Nikolaos, Crete, Greece PC 72100

Background:   The clinical relevance and the optimal treatment regimens of Non-Tuberculous Mycobacteria (NTM) differs strongly by species.  We describe our experience regarding the molecular identification of NTM species by using commercial and more advantaged molecular identification methods as sequence analysis.

Material/methods:  Specimens submitted for mycobacterial culture between 12/2006-12/2023 in a University Hospital.  The recovered NTM were identified by the commercially available reverse hybridization-based assays Genotype Mycobacterium Common Mycobacteria and Additional Species (Bruker). Sequencing analysis of 16S rDNA (1500bp) and  hsp65 (440bp) genes was performed when necessary.  

Results: In total, 581 non-repetitive clinical isolates of NTM were recovered; belonged to 35 known Mycobacterium species with most frequent the M. avium (n= 113) and M. gordonae (n=82), while 6 strains were not belonged to any known species representing probably novel Mycobacterium species. The 91.2% of strains (18 species) were correctly identified at the species level by the commercial assays. Thirty-two (5.5%) strains identified by the commercial assays only to the genus level. Twenty-six belonged to  rare NTM species while six strains had unique sequences. Eleven M. marseillense   and one M. mantenii strains erroneously identified by the commercial assays as M. intracellulare.   Three M. kumamotonense and one M. nonchromogenicum strains were identified as M. celatum and 3 M. parascrofulaceum strains were identified as M. scrofulaceum.

Conclusions: Our findings suggest that the combined use of molecular commercial identification tests with sequencing analysis improve the ability to correctly identify   the common but also the rare NTM, as well as to detect probably novel mycobacterial species.

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