Genome of Acetobacter pasteurianus DmelCS_004

Draft Genome Sequence for Acetobacter pasteurianus DmelCS_004 isolated from the Gut of Drosophila melanogaster

David Sannino, John M. Chaston, Peter D. Newell, Chun-nin (Adam) Wong and Angela E. Douglas

 Department of Entomology, Cornell University, Ithaca, NY

Acetic acid bacteria are have been identified as symbionts in insects of multiple orders. Here we report the draft genome sequence of Acetobacter pasteurianus DmelCS_004, isolated from the gut of Drosophila melanogaster. We assembled the genome with Velvet and annotated the genome using the RAST server, attaining a genome sequence of 137 contigs with 2,840 annotated features and a total of 2,845,508 bp.

Bacteria of the genus Acetobacter are Gram-negative obligate aerobes, which grow in sugary, acidic, and ethanol-rich environments, and are distinguished from other Acetic acid bacteria by their ability to oxidize acetate and lactate to CO2 (Kersters et al., 2006). Acetobacter species such as Acetobacter pasteurianus are economically important as they acetify wine, cider, rice, and other substrates to produce vinegar, and are commonly detected in wine and vinegar, as well as on grapes (Hidalgo et al., 2010; Kersters et al., 2006; Wu et al., 2010). Acetobacter species are also associated symbiotically with insects of the orders Diptera, Hymenoptera, and Hemiptera that feed on sugar-rich diets (Crotti et al., 2010). Drosophila melanogaster is attracted to rotting fruits, and feeds on the yeasts and bacteria, including Acetobacter, that utilize these substrates (Crotti et al., 2010). We previously demonstrated that a bacterium assigned to the species Acetobacter pomorum is one of the five most abundant bacterial species that comprise the gut microbial community of lab raised Drosophila melanogaster (Wong et al., 2011). After isolating the bacterium from the gut of D. melanogaster Canton-S, and sequencing the genome, we determined the isolate to be Acetobacter pasteurianus, which is closely related to A. pomorum.

Here we announce the sequencing of the Acetobacter pasteurianus DmelCS_004 genome.

Acetobacter pasteurianus DmelCS_004 was isolated from aseptically dissected guts of Drosophila melanogaster, and maintained at 30° C on M.R.S. medium (de Man et al., 1960).  Cultures were grown shaking at 200 rpm to late-log phase and genomic DNA prepared with the Qiagen DNeasy Blood and Tissue Kit as per the manufacturer’s recommendations.  The Cornell Life Sciences Core Facility performed library preparation for Illumina sequencing.

We acquired 22,306,666, 100-bp paired-end read-pairs using Illumina HiSeq 2000. 21,082,160 read-pairs passed quality filtering, providing ~1500X genome coverage, and we used a random subset of 17,500,000 quality filtered read pairs for the assembly. We assembled the genome using Velvet 1.2.03 (Zerbino and Birney, 2008) by randomly allocating each sequence to one of 7 subset files, each containing 2,000,000 read pairs (141X coverage per subset file). Sequences from each subset file were assembled into contigs using a range of kmer lengths of 77-DmelCS_004, aided by predictions of the Velvet Advisor tool (Seeman) and tests with multiple kmer lengths. The output contig file from each subset was used as input in a second velvet run with a kmer length of DmelCS_004 to create a final assembly that was 2,834,454 bp long and represented by 137 contigs, with a maximum contig size of 466,505-bp and N50 of 90,072-bp.

Annotation and subsequent analyses were performed using the Rapid Annotation using Subsystem Technology (RAST) server (Aziz et al., 2008) to create an annotated genome sequence with 2,845,508 bp, 137 contigs, 2840 features, and an estimated 7 missing genes.

The annotated genome sequence and predicted gene functions of A. pasteurianus DmelCS_004 was analyzed in SEED. The A. pasteurianus DmelCS_004 genome contains a complete predicted ammonia assimilation pathway, in which the organism assimilates ammonia through an NADPH dependent glutamate synthase. A. pasteurianus DmelCS_004 also contains a complete pathway for urea decomposition, containing a urea ABC transporter for the uptake of urea, and a urease that degrades urea to carbamine acid and ammonia. These two components are not present in the genomes of the other Acetobacters and Lactobacilli isolated from the guts of D. melanogaster, suggesting A. pasteurianus may function in assisting host urea decomposition. There are no predicted genes for motlility found in the genome. Interestingly, as Acetobacter comprises obligate aerobes, a predicted arsenate reductase was found in the genome. The genome has the genes for a complete Pentose Phosphate Pathway, and an incomplete, non-functional glycolysis pathway, indicating that A. pasteurianus DmelCS_004 utilizes the pentose phosphate pathway as an alternative to glycolysis

The sequence file is available as Download (see sidebar on left).  If you need the data in a different format, please contact us.

Acknowledgements

This work was supported by a NIH grant 1R01GM095372 to AED and Ruth L. Kirschstein NRSA postdoctoral fellowship to PDN (1F32GM099374-01). We thank Peter Schweitzer for assistance with filtering reads and Madeline Galac for assistance installing Velvet.

 

References

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