Draft Genome Sequence of Mycobacterium austroafricanum DSM 44191.

We announce the draft genome sequence of Mycobacterium austroafricanum DSM 44191(T) (= E9789-SA12441(T)), a non-tuberculosis species responsible for opportunistic infection. The genome described here has a size of 6,772,357 bp with a G+C content of 66.79% and contains 6,419 protein-coding genes and 112 RNA genes.

GENOME ANNOUNCEMENT

Mycobacterium austroafricanum was named after the initial isolation of a set of 23 strains from water in South Africa (1). Numerical taxonomy indicated that these isolates are representative of a new species related to Mycobacterium parafortuitum (1). Further genetic analyses indicated that, in fact, M. austroafricanum belongs to the Mycobacterium vaccae complex, which also contains Mycobacterium vanbaalenii, and is more distantly related to Mycobacterium aurum and Mycobacterium pyrenivorans (2). M. austroafricanum is an environmental organism isolated from soil, in particular, from hydrocarbon-polluted soils (3, 4). Indeed, M. austroafricanum has attracted much attention because it is able to degrade gasoline hydrocarbons (5, 6). M. austroafricanum has rarely been isolated from patients (7, 8), and while M. austroafricanum DNA has been detected in diseased joint fluids, the clinical significance of M. austroafricanum has not yet been established (9).

We therefore sequenced the whole genome of the M. austroafricanum DSM 44191 (E9789-SA12241T) strain in order to illustrate its phylogenetic relationship with closely related mycobacteria and to help depict its unique metabolic capabilities.

Genomic DNA was isolated from M. austroafricanum grown in Middlebrook 7H9 broth (Becton Dickinson, Sparks, MD) at 37°C. It was then sequenced using three high throughput NGS technologies: Roche 454 (Roche Diagnostics Corporation, Indianapolis, IN) (10), SOLiD version 4 (Life Technologies, Carlsbad, CA), and MiSeq Illumina (Illumina Inc., San Diego, CA). A 3.74-kb paired-end library was loaded on a picotiter plate and sequenced with the Roche-GS FLX Titanium Sequencing Kit XLR70. The run yielded 143.9 Mb with 435,968 passed filters and an average length of 329 bp. The bar-coded paired-end SOLiD library generated 975,705 reads of 50× 35-bp length. Finally, a paired-end Nextera library, fragmented at 800 bp, sequenced on MiSeq at 2× 151 bp, yielded 823,878 reads with an indexing of 6.57% on the flowcell.

Reads from these various sequencing technologies were first assembled separately. The 454 reads were then assembled into contigs and scaffolds using Newbler version 2.8 (Roche). Illumina reads were trimmed using Trimmomatic (11), then assembled using the Spades software (12, 13). The obtained contigs were combined by SSPACE (14) and Opera software v1.2 (15) complemented by GapFiller v1.10 (16). The genome was improved using CLC Genomics v5 software (CLC bio, Aarhus, Denmark).

The M. austroafricanum draft genome sequence consists of 20 scaffolds of 69 contigs containing 6,682,536 bp, with an estimated genome size including gaps of 6,772,357 bp. The G+C content of this genome is 66.79%. Noncoding genes and miscellaneous features were predicted using RNAmmer (17), ARAGORN (18), Rfam (19), and PFAM (20). Open reading frames were predicted using Prodigal (21), and functional annotation was achieved using BLASTp against the GenBank database (22) and the Clusters of Orthologous Groups (COG) database (23, 24). The genome was shown to encode at least 112 predicted RNAs, including 4 rRNAs, 85 tRNAs, 2 transfer-messenger RNAs, and 21 miscellaneous RNAs. Also, 6,419 genes were identified, which yields a coding capacity of 6,124,875 bp (coding percentage, 90.4%). Among these genes, 850 (12.77%) were found to encode putative proteins and 1,324 (16.88%) were assigned as encoding hypothetical proteins. Moreover, 6,549 genes matched at least one sequence in the COG database with BLASTp default parameters.

Nucleotide sequence accession numbers.

The Mycobacterium austroafricanum strain DSM 44191 (= E9789-SA12441T) genome sequence has been deposited at DDBJ/EMBL/GenBank under the accession no. HG964450 to HG964469. The whole-genome shotgun master numbers are CCAW010000001 to CCAW010000069.