JOURNAL OF BACTERIOLOGY, Sept. 2011, p. 5057 0021-9193/11/$12.00 doi:10.1128/JB.05647-11 Copyright © 2011, American Society for Microbiology. All Rights Reserved.
Vol. 193, No. 18
Genome Sequence of the Newly Isolated Chemolithoautotrophic Bradyrhizobiaceae Strain SG-6C䌤 Stephen L. Pearce, Rinku Pandey, Susan J. Dorrian, Robyn J. Russell, John G. Oakeshott, and Gunjan Pandey* CSIRO Ecosystem Sciences, Clunies Ross Street, Acton, ACT 2601, Australia Received 23 June 2011/Accepted 29 June 2011
Strain SG-6C (DSM 23264, CCM 7827) is a chemolithoautotrophic bacterium of the family Bradyrhizobiaceae. It can also grow heterotrophically under appropriate environmental conditions. Here we report the annotated genome sequence of this strain in a single 4.3-Mb circular scaffold. Strain SG-6C (DSM 23264, CCM 7827) was isolated from imidacloprid-contaminated soil collected in Canberra, Australia, after enrichment using 6-chloronicotinic acid as the sole carbon source. SG-6C is capable of chemolithoautotrophic growth using CO2 as a carbon source, as well as growth using 6-chloronicotinic acid as a sole carbon source (G. Pandey et al., unpublished data). This bacterium is a member of the family Bradyrhizobiaceae, but polyphasic characterization indicates the possibility that it belongs to a new species, if not a new genus, within this family. Further investigations to determine its taxonomic position are under way. SG-6C genomic DNA was prepared by using Qiagen Genomic-tip 500/G (Qiagen, GmbH) and following the manufacturer’s protocol. Both standard 454 pyrosequencing and paired-end sequencing were performed on a Genome Sequencer FLX system. We obtained 469,219 single and 135,028 paired-end sequences and assembled them using the Newbler de novo assembler (454 Life Sciences, Branford, CT) into 38 contigs. Paired-end information was able to combine these contigs into a single 4.3-Mb circular scaffold. The sequence was automatically annotated using the RAST server (1), followed by manual curation of the assigned annotations. The genome has a GC content of 61.9% and is predicted to contain 4,248 protein-coding sequences, 48 tRNA genes (1 nonfunctional), and 3 rRNA genes. We functionally annotated 2,686 protein-coding sequences (63%) by comparison to known genes. The 16S rRNA gene sequence has 98% identity to the chemolithoautotrophs Oligotropha carboxidovorans and Bradyrhizobium japonicum. O. carboxidovorans and B. japonicum, of the family Bradyrhizobiaceae, fix carbon through the Calvin-Benson-Bassham (CBB) cycle, which requires the key enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) (4, 7). The genome of SG-6C contains genes encoding a nearly complete CBB cycle. No RubisCO gene has been identified in SG-6C, however, meaning that carbon dioxide is not fixed through this pathway. Five other CO2 fixation pathways are known to exist in
bacteria and archaea. These are the reductive citric acid cycle pathway (3), the Wood-Ljungdahl pathway (8), the 3-hydroxypropionate bicyclic pathway (5), the 3-hydroxypropionate/4hydroxybutyrate cycle pathway (2), and the dicarboxylate/4hydroxybutyrate cycle pathway (6). SG-6C contains many genes involved in all five pathways; however, no complete pathway is present. The pathway through which SG-6C fixes carbon dioxide is therefore unknown. The median size of the coding sequences in the SG-6C genome is 782 bp. According to the Newbler output, the median gap size in the assembly is 250 bp (maximum size, 1,298 bp). It is therefore unlikely that many genes are missing from the current annotation. Strain SG-6C may therefore fix carbon dioxide using a novel pathway. Further analysis of SG-6C will help clarify if this is the case. Nucleotide sequence accession numbers. This whole-genome shotgun project has been deposited at DDBJ/EMBL/ GenBank under accession no. AFOF00000000. The version described in this paper is the first version, AFOF01000000. We thank Stephanie Palmer, Stephen Ohms, and Peter J. Milburn from the ACRF Biomolecular Resource Facility of the Australian National University for sequencing and assistance in the assembly of the genome. We also thank Peter East and David Seddon of CSIRO for helpful discussions and technical assistance in setting up a computer for genome analysis, respectively. This work was funded by CSIRO. REFERENCES 1. Aziz, R. K., et al. 2008. The RAST Server: rapid annotations using subsystems technology. BMC Genomics 9:75. 2. Berg, I. A., D. Kockelkorn, W. Buckel, and G. Fuchs. 2007. A 3-hydroxypropionate/4-hydroxybutyrate autotrophic carbon dioxide assimilation pathway in Archaea. Science 318:1782–1786. 3. Evans, M. C., B. B. Buchanan, and D. I. Arnon. 1966. A new ferredoxindependent carbon reduction cycle in a photosynthetic bacterium. Proc. Natl. Acad. Sci. U. S. A. 55:928–934. 4. Fuhrmann, S., et al. 2003. Complete nucleotide sequence of the circular megaplasmid pHCG3 of Oligotropha carboxidovorans: function in the chemolithoautotrophic utilization of CO, H2 and CO2. Gene 322:67–75. 5. Herter, S., G. Fuchs, A. Bacher, and W. Eisenreich. 2002. A bicyclic autotrophic CO2 fixation pathway in Chloroflexus aurantiacus. J. Biol. Chem. 277:20277–20283. 6. Huber, H., et al. 2008. A dicarboxylate/4-hydroxybutyrate autotrophic carbon assimilation cycle in the hyperthermophilic archaeum Ignicoccus hospitalis. Proc. Natl. Acad. Sci. U. S. A. 105:7851–7856. 7. Masuda, S., S. Eda, C. Sugawara, H. Mitsui, and K. Minamisawa. 2010. The cbbL gene is required for thiosulfate-dependent autotrophic growth of Bradyrhizobium japonicum. Microbes Environ. 25:220–223. 8. Ragsdale, S. W., and E. Pierce. 2008. Acetogenesis and the Wood-Ljungdahl pathway of CO2 fixation. Biochim. Biophys. Acta 1784:1873–1898.
* Corresponding author. Mailing address: CSIRO Ecosystem Sciences, GPO Box 1700, Canberra, ACT 2601, Australia. Phone: (61) 2 6246 4244. Fax: (61) 2 6246 4000. E-mail:
[email protected]. 䌤 Published ahead of print on 8 July 2011. 5057
