Draft Genome Sequence of Mycobacterium neworleansense Strain ATCC 49404T.

Mycobacterium neworleansense is a rapid growing nontuberculosis species belonging to the Mycobacterium fortuitum complex. The draft genome of M. neworleansense ATCC 49404(T) comprises 6,287,317 bp exhibiting a 66.85% G+C content, 5,997 protein-coding genes, and 89 predicted RNA genes.

GENOME ANNOUNCEMENT

Mycobacterium neworleansense is a rapidly growing nontuberculous mycobacterium formerly classified as Mycobacterium fortuitum sorbitol-negative third biovariant (1). Within the M. fortuitum complex, M. neworleansense clusterizes with Mycobacterium peregrinum, Mycobacterium septicum, and Mycobacterium porcicum (2, 3). The name M. neworleansense derives from New Orleans, USA, where this mycobacterium was first isolated from a scalp wound (1).

We performed the whole-genome sequencing of M. neworleansense ATCC 49404T in order to refine understanding of its taxonomic relationships with closely related species of the M. fortuitum complex.

From this perspective, M. neworleansense ATCC 49404T was cultured in MGIT Middlebrook liquid culture (Becton Dickinson, Le Pont-de-Claix, France) at 37°C in a 5% CO2 atmosphere. M. neworleansense genomic DNA was then sequenced by Illumina MiSeq runs (Illumina Inc., San Diego, USA) using a 5-kb mate-paired library. Reads were trimmed using Trimmomatic (4) and assembled using Spades v3.5 (5, 6). Contigs were combined together by SSPACE v2 (7), Opera v2 (8) helped by GapFiller v1.10 (9), and homemade tools in Python to refine the set. This resulted in a draft genome consisting of nine contigs without gap for a total of 6,287,317 bp and a G+C content of 66.85%. Noncoding genes and miscellaneous features were predicted using RNAmmer (10), ARAGORN (11), Rfam (12), PFAM (13), and Infernal (14). Coding DNA sequences were predicted using Prodigal (15), and functional annotation was achieved using BLAST+ (16) and HMMER3 (17) against the UniProtKB database (18). The genome was shown to encode at least 89 predicted RNAs, including 9 rRNAs, 59 tRNAs, 1 transfer-messenger RNA (tmRNA), and 20 miscellaneous RNAs. A total of 5,997 identified genes yielded a coding capacity of 5,837,343 bp (coding percentage, 92.84%). Among these genes, 246 (4.1%) were found to be putative proteins and 1,011 (16.86%) were assigned as hypothetical proteins. Moreover, 4,296 genes matched a least one sequence in the Clusters of Orthologous Groups (19, 20) with BLASTP default parameters.

In silico DNA-DNA hybridation (DDH) (21) was performed with the M. fortuitum complex species and other reference genomes selected on the basis of their 16S rRNA gene sequence proximity with M. neworleansense. The M. neworleansense genome was locally aligned 2 by 2 using a BLAT algorithm (22, 23) against each the selected genomes, and DDH values were estimated from a generalized linear model (24). The DDH was 35.9% ± 2.48% with Mycobacterium septicum DSM 44393 (25), 32.8% ± 2.46% with Mycobacterium senegalense CK2 M4421 (1), 32.8% ± 2.46% with Mycobacterium conceptionense (26), 32.6% ± 2.46% with Mycobacterium farcinogenes (27), 32.3% ± 2.46% with Mycobacterium peregrinum (28), 31.7% ± 2.46% with Mycobacterium fortuitum ATCC 6841 (29), 20.5% ± 2.32% with Mycobacterium chubuense NBB4 (30) and Mycobacterium gilvum Spyr1 (31), and 20.1% ± 2.31% with Mycobacterium aromaticivorans JS19b1 (32).

These data confirm M. neworleansense as a unique species more closely related to M. septicum in the M. fortuitum complex.

Nucleotide sequence accession numbers.

The M. neworleansense ATCC 49404T genome sequence has been deposited at EMBL under the accession numbers CWKH01000001 to CWKH01000009.